Light-harvesting chlorophyll a/b binding proteins are important in plant photosynthesis and abiotic stress response.To study the characteristics of GhLhcb2A1 and its expression patterns and functions in low temperature and drought response in upland cotton,full-length CDS of GhLhcb2A1 gene was cloned from the leaf cDNA of Jimian 262 by PCR.Bioinformatic analysis was conducted to learn the basic characteristics of the gene.The expression patterns and functions in low temperature and drought response were evaluated by qRT-PCR and virus-induced gene silencing.It was shown that the length of GhLhcb2A1 CDS was 798 bp,encoding 265 amino acids.GhLhcb2A1 was highly expressed in leaves and was significantly up-regulated in leaves and roots under low temperature and drought treatment.Compared with the control,its expression maximized at 3 h under low temperature and drought in leaves with 17.42 and 30.03 folds increase respectively,whereas maximized at 6 h under low temperature and 12 h under drought in roots with 11.65 and 65.04 folds respectively.Subcellular localization assay verified that GhLhcb2A1 was expressed in the chloroplasts of cells.Compared with the control plants,GhLhcb2A1 silenced plants showed a more severe phenotype of water loss and dryness under both low temperatures and drought.The accumulated malondialdehyde content in the leaves of the silenced plants was significantly higher than that of the control,while the proline content and superoxide dismutase activity were significantly lower than those of the controls,suggesting that GhLhcb2A1 silenced plant reduced the resistance to low temperature and drought.The above results implied that this gene played a positive role in regulating low temperature and drought response.

To explore candidate genes for resistance of Aphis gossypii to the insecticide imidacloprid,the transcriptome data of the two strains were obtained and compared with imidacloprid indoor resistant and relatively sensitive lines of Aphis gossypii by using Illumina HiSeq 2500 high-throughput sequencing technology.The gene annotation was performed using the NCBI database,and bioinformatics analysis of the differential genes at the transcriptional level included GO function,KEGG metabolic pathway,and other analyses.The relative expression of eight candidate differentially expressed genes(CYP6a2,CYP6a13,CYP6k1,CYP6j1,CYP4c1,AChE2,CarE and ALP3)was detected using qRT-PCR technology,and the evolutionary relationships of resistance related genes were analyzed.After sequencing and sequence splicing,a total of 70 101 Unigenes were obtained,with an average length of 654.37 bp.29 131,27 861 and 2 993 Unigenes were annotated in NR,GO and KEGG databases,respectively.According to the NR annotation analysis of the differential genes of insecticide resistance and sensitivity strains,a total of 22 differential genes that may be related to insecticide resistance were found,including 9 detoxification enzyme genes(CYP6a13,CYP6k1,CYP6j1,CYP4c1 and ALP3 each,two CYP6a2,two CarE),8 cuticle protein genes(CP)and their precursors(CPP),target enzyme genes(AChE2),2 transcription factors(WRKY1,leucine zipper transcription factor-like protein 1 gene,LZTFL1),one pancreatic lipase-related protein 2 gene(PLRP2)and one multidrug resistance-associated protein gene(MRP).The results of qRT-PCR indicated that the expression levels of CYP6a2,CYP6a13,CYP6k1,CYP6j1,CarE and ALP3 genes in insecticide resistance strains were significantly higher than those in sensitive strains.The phylogenetic tree analysis of CYP,ALP,GST and CP genes obtained by NR annotation indicated that the genetic relationship between the Aphis gossypii and Aphis glycines and Acyrthosiphon pisum was relatively close.We found that 87.50% of the candidate differentially expressed genes in the two strains showed consistent changes in expression levels at the transcriptional and mRNA levels.

Currently,traditional breeding methods cannot fully meet the demands of the cotton market and production. Therefore, molecular biotechnology can be employed to accelerate the development of new cotton varieties. Transcription factors have emerged as significant tools in the study of cotton gene function, genetics, and breeding during this period. Among them, the MYB transcription factor family is one of the largest transcription factor families present in many plants, and it performs multiple roles in plant growth and development. Hence, the MYB transcription factor holds high research value. While the function of MYB transcription factors has been extensively studied in model plants, research in non-model plants, particularly cotton, remains limited and is mostly concentrated in upland cotton. To further understand MYB transcription factors, this paper reviews the research progress on MYB transcription factors in various plants and cotton. It covers their classification basis, structural characteristics, evolution, roles in response to biotic and abiotic stress, cotton fiber development, and secondary metabolism. Additionally, it provides statistics on the known functions of cotton MYB transcription factors. This review aims to deepen our understanding of cotton MYB transcription factors and serve as an important reference for future studies on the functions and mechanisms of MYB transcription factors in different cotton species.

In order to investigate the physiological regulation mechanism of exogenous H₂O₂ on the cotton seedlings under NaCl stress,the cotton variety Xinluzao 48 was used as the test material in an outdoor potting method.Two-factor random combinations of salt stress (NaCl,concentration gradients of 0,100,200 mmol/L) and H₂O₂ (concentration gradients of 0,0.005,0.010,0.020,0.050 mmol/L) were set,to study the change rule of fresh weight,dry weight,chlorophyll content,chlorophyll fluorescence parameters,photosynthetic gas parameters,antioxidant enzyme activities and osmotic regulation system of cotton seedlings.The results showed that exogenous H₂O₂ effectively alleviated the inhibitory effect of salt stress on the growth of cotton seedlings,increased chlorophyll content,photosynthetic gas parameters and chlorophyll fluorescence parameters of cotton seedlings,maintained the normal operation of photosynthesis of cotton seedlings and ensured the accumulation of dry matter.Meanwhile,exogenous H₂O₂ could increase the activity of antioxidant enzymes (POD,APX,CAT),accelerated the removal of ROS from cotton seedlings.Exogenous H₂O₂ reduced the electrolyte leakage rate,MDA content,the content of osmoregulation substances such as pro,free amino acid and SS,and improved the salt resistance of cotton seedlings.Among all treatment,0.020 mmol/L exogenous H₂O₂ had the best effect in alleviating the salt stress suffered by cotton seedlings.In summary,exogenous H₂O₂ improves the adaptation of cotton seedlings to salt stress by improving photosynthetic performance,keeping stable photosynthesis in cotton seedlings,and maintaining the dynamic balance between ROS production and elimination in cotton seedlings.

GhENODL6 has been discovered to be involved in the resistance of Gossypium hirsutum to Verticillium dahliae.To further reveal its role in cotton resistance against Verticillium wilt,GhENODL6 open reading frame was inserted into the yeast expression vector pNC-GBKT7 using Nimble Cloning technology to construct the recombinant plasmid pNC-GBKT7-ENODL6.The resulting plasmid was successfully transferred into yeast Y2HGold and exhibited normal growth on DDO medium,while failing to grow on QDO/X/A medium,indicating that the GhENODL6 protein did not exhibit toxicity towards the yeast host and lacked autoactivating activity.Subsequently,the Y2HGold yeast carrying the pNC-GBKT7-ENODL6 decoy vector was subjected to hybridization with a cDNA library and screened for colonies displaying blue coloration.Through PCR amplification,a 526 bp non-vector insert was obtained in these blue yeasts,which showed high sequence consistency with the WRKY47 gene within G.hirsutum genome.By employing homologous amplification techniques,we successfully cloned the WRKY47 open reading frame from upland cotton,which spanned 1 587 bp in length and encoded 528 amino acid residues.Furthermore,a yeast two-hybrid assay confirmed an interaction between WRKY47 and GhENODL6.To summarize our findings,the study constructed the recombinant vector pNC-GBKT7-ENODL6 and identified WRKY47 as an interacting protein of GhENODL6.

MYB transcription factors play an important role in the growth and development of cotton,and GhMYB42 is one of the transcription factors of the MYB family,which also has certain research value,so this study cloned the coding sequence of GhMYB42 gene from cotton and constructed a prokaryotic expression vector.The nucleotide sequence and amino acid sequence of GhMYB42 were analysed using bioinformatics methods, and the coding sequence of the GhMYB42 gene was constructed into the prokaryotic expression vector pGEX-4T-1 by the Gataway BP and LR reactions, and the optimal conditions for the induction of the protein by IPTG were determined by setting up different IPTG-inducing conditions, and finally the recombinant protein was identified by Western Blot.The results showed that the full-length sequence of GhMYB42(XP_016732693.1) was 1 508 bp,the coding region length was 792 bp,the coding area was 263 amino acids,the predicted molecular weight was about 29.534 ku,and the isoelectric point was 5.18.Amino acid sequence comparison analysis revealed that the sequence similarity of the MYB transcription factors was 80.62%, and that the N-terminus of the GhMYB42 protein contained two tandem SANT structural domains, making it an R2R3 transcription factor.Phylogenetic tree analysis showed that the MYB42 protein in Gossypium hirsutum had the highest homology with another MYB protein (XP_012439547.1) in Gossypium hirsutum and was on one branch.Since the results of each experimental gradient were not significantly different during protein induction,it chose the conditions for IPTG to be 0.2 mmol/L at a final concentration of 0.2 mmol/L,a temperature of 37 ℃ for 3 h,and a temperature and time for protein solubility to be induced at 37 ℃ for 3 h.The Western Blot results showed that the size of the recombinant protein was correct,and finally the GhMYB42 recombinant protein with a size of 55.54 ku was successfully obtained,and it will purify the recombinant protein and further study the function of the transcription factor GhMYB42.

In order to explore the effect of salt stress on the physiological mechanism of cotton,saline-tolerant cotton Zhong J0710 was used as a control,two new cotton distantly hybrid germplasms HL2 and A₂H cultivated in cotton breeding research group were treated with salt solution for 15 d(concentration of 200 mmol/L)at seedling stage,and the relative plant height,relative root length and relative survival rate of the test materials were analyzed,and the salt damage index was calculated.The salt tolerance of the test materials was comprehensively evaluated by combining the measurement results of physiological indicators such as primary light energy conversion efficiency (Fv/Fm),antioxidant enzyme activity,proline content,and soluble sugar of seedlings at different treatment times (0,5,12,24,48 h).The results showed that after 15 d of salt stress,the damage degree of the growth of seedlings of the three test materials was as follows:Zhong J0710<HL2<A₂H,and salt stress significantly inhibited the relative plant height,relative root length and relative survival rate of A₂H,indicating that the salt concentration treatment had less effect on the growth and development of HL2 and Zhong J0710,but had a greater effect on A₂H.Under different treatment times,salt stress had different degrees of influence on the maximum light energy conversion efficiency of the three test materials,and the inhibition degrees from weak to strong compared with before stress were HL2<A₂H<Zhong J0710.Compared with the non-stress period, the peroxidase (POD) activity of the tested materials was increased by salt stress, and the POD activity of HL2 during the stress process was significantly higher than that of A2H and Zhong J0710 in the salt-tolerant control, and reached a very significant level at 12—48 h of stress,and the change law of superoxide dismutase (SOD) activity was not obvious,and the role in salt stress was low.The contents of proline in HL2 and the salt-tolerant control increased with the increase of salt stress time,and the soluble sugars and soluble proteins of germplasm HL2 were significantly higher than those in the salt-tolerant control and A₂H during the entire salt stress period.In summary,in salt adversity,salt-tolerant materials have a more developed root system;higher light energy conversion efficiency,higher reactive oxygen species clearance capacity in cells and higher accumulation of osmoregulatory substances are the physiological basis for strong salt tolerance.

The aim of this study was to evaluate the genetic diversity of upland cotton germplasm resources and screen representative germplasm,so as to provide theoretical basis for scientific evaluation and efficient utilization of upland cotton germplasm resources.Based on the SNP markers of 367 upland cotton resources and the phenotypic data of 353 resources,genetic diversities were analyzed,phylogenetic tree and primary core germplasm were constructed,and effects of primary core germplasm construction were evaluated.The results showed that the polymorphism information content(PIC)of the original upland cotton population was 0.24,and the genetic diversity index of each phenotypic trait was close to 2.00,which were similar to the results of previous studies.The upland cotton germplasm could be divided into three groups based on SNP markers,and 73 genotypic primary core resources were constructed.The Nei's genetic diversity index,PIC and other genetic diversity indexes of primary core germplasm were higher than those of original germplasm.Based on the phenotypic data,the upland cotton germplasm could be divided into three groups,and the mean value of each trait showed the trend that the group Ⅰ was the smallest,the group Ⅱ was in the middle,and the group Ⅲ was the largest.Seventy phenotypic primary core resources were constructed.Compared with the original germplasm,the mean values of all traits were not significantly different,the range and genetic diversity index were similar to that of the original germplasm,and the coefficient of variation was higher than that of the original germplasm.Fifteen primary core resources constructed based on SNP markers overlapped with those constructed based on phenotypic data.Most of these resources were genotypic group Ⅱ and phenotypic group Ⅱ.These results showed that the genotypic genetic diversity of upland cotton germplasm resources was low,but the phenotypic variation was rich and the phenotypic genetic diversity was high.The genotypic and phenotypic primary core germplasm constructed in this study had a good representation of genetic diversity and could be used as a typical germplasm for preservation and utilization.

Genomic-wide identification of systematic work on the SKS gene family in cotton has not been reported.Identifying SKS (Skewed5 similar) gene family members and analyzing evolutionary relationship in Gossypium hirsutum,and analyzing the mechanism of GhSKS13 gene regulating cotton disease resistance are expected to provide a new direction for the cultivation of resistant plants.Here,the members of upland cotton SKS family were identified by bioinformatics at the genome-wide based on published genomic data of upland cotton genetic standard line TM-1,and G.hirsutum variety CRI-14 was used as the material.And chromosome distribution,evolutionary relationships,gene structure,and collinearity of SKS family members were predicted.Then,the expression pattern of GhSKS13 was analyzed by Real-time Quantitative PCR and virus-induced gene silencing was employed for preliminary investigation of the function of GhSKS13 in cotton resistance to Verticillium dahliae.A total of 48 upland cotton SKS genes were identified,unevenly distributed on 19 chromosomes,clustered into 5 subgroups with highly conserved gene sequences.Collinearity analysis revealed that the upland cotton SKS gene family was subject to purifying selection.Expression pattern analysis showed that GhSKS13 was predominantly expressed in upland cotton root tissues and significantly up-regulated by V.dahliae infection. GhSKS13-silenced reduced plants resistance to V.dahliae and suppressed the expression of the pathogen-related genes including GhPR1, GhPR2,GhPR3 and GhPR5 compared to control plants. GhSKS13-silenced plants invaded by V.dahliae showed significantly lower hydrogen peroxide (H₂O₂) deposition compared to the control,suggesting that GhSKS13 promotes the formation of reactive oxygen species (ROS).In conclusion,this study clarified the phylogenetic relationships,chromosomal distribution characteristics and gene structure characteristics of upland cotton SKS family members,and elucidated the involvement of GhSKS13 in upland cotton plant resistance to V.dahliae.

To investigate the effects of Mn treatment on the growth,development and cadmium adsorption capacity of cotton under Cd stress.Different concentrations of Mn treatment(0,5,10,50 μmol/L)were applied to exploit changes of superoxide dismutase(SOD),catalase(CAT),malondialdehyde(MDA),free proline(Pro),Cd content,dry matter quality,and main root length of roots of cotton seedlings of Xiang C178 and Xiang FZ001 under Cd stress.The stress of 10 μmol/L Cd had a significant effect on the growth of cotton root.The physical barrier and antioxidant defense mechanism were constructed to reduce the damage of Cd to root cells,resulting in a decrease in SOD activity and an increase in CAT activity,MDA content and Pro content.And, adding Mn was observed to effect the physiological response to cadmium of cotton seedling roots. Low concentration of Mn treatment resulted in the decrease of Cd content,the increase in SOD activity and Pro content,the decrease in CAT activity and MDA content in the root of cotton seedlings,which effectively prevented the inhibition of Cd stress on the root growth of cotton seedlings.High concentrations of Mn could increase Cd content and SOD activity in cotton seedling roots, which affected the root growth of cotton seedlings.When using cotton to repair Cd-contaminated farmland,it can be less than 10 μmol/L Mn ion treatment is used to improve the Cd tolerance of cotton seedlings and ensure the normal growth of cotton plants.

To investigate the function of GhFUL1 gene in branching development of cotton,the GhFUL1 gene in the MADS-box family and its promoter were cloned from cotton,and the function and promoter activity of the gene were analyzed.The results indicated that the open reading frame (ORF) of GhFUL1 gene was 726 bp and encoded 241 amino acids.Phylogenetic tree analysis showed that the amino acid sequence of FUL subclade was conserved,and GhFUL1 owned the highest similarity to Cacao TcAGL8-1 except for the homologous proteins in cotton.Subcellular localization result showed that GhFUL1 was localized in nucleus and membrane.The expression level of different cotton tissues was examined,and the result showed that GhFUL1 was highly expressed in shoot apical meristem.Overexpressing GhFUL1 in Arabidopsis thaliana increased the number of lateral branches by decreasing the expression of cytokinin oxidation/dehydrogenase genes AtCTK1 and AtCTK6.Compared with the wild type,transgenic plants did not result in significant changes of the flowering time and floral organ development.The present results suggested that GhFUL1 might affect the number of branches by regulating the cytokinin synthesis pathway.The promoter of GhFUL1 was predicted to include the TATA-box and CAAT-box elements,and cis-acting elements related to light response,auxin response and stress response by PlantCARE website.Then the promoter of GhFUL1 was cloned and constructed into expression vector pBI121 to detect promoter activity.The GUS staining of transgenic Arabidopsis lines showed that GhFUL1 promoter was specifically expressed in shoot apical meristem at seedling stage and flower organ at maturity.This study revealed that the gene and promoter of GhFUL1 played an important role in the Arabidopsis branching development process after transformation.

In order to study the function of GhAOP2-like in cotton,based on the previous proteomic data from the laboratory of Institute of Cotton, Hebei Academy of Agriculture and Forestry Sciences, this study obtained the gene sequence of GhAOP2-like using homologous cloning strategy.The physicochemical properties,structure and subcellular localization of GhAOP2-like protein were analyzed by bioinformatics method.The tissue-specific expression of GhAOP2-like and the change of expression level under drought,salt and hormone treatment were detected by quantitative real-time polymerase chain reaction(qRT-PCR).The results showed that the CDS sequence length of GhAOP2-like was 972 bp and encoded 323 amino acids.GhAOP2-like was located on chromosome D13.GhAOP2-like protein did not contain signal peptides and transmembrane domains,and was located in the cytoplasm with the molecular formula of C₁₆₅₄H₂₅₂₅N₄₂₉O₄₇₈S₁₉ and the theoretical isoelectric point was 5.20.The clustering analysis results showed that the AOP sequences in cotton and other species were clearly divided into two groups,but had the closest relationship with the AOP sequences of Hibiscus syriacus.qRT-PCR results showed that GhAOP2-like was expressed in roots,stems,leaves,and developing seeds,but the expression level of GhAOP2-like was the highest in roots.GhAOP2-like was up-regulated under drought,salt,GA₃,MeJA and ABA treatments,but the expression level changed most in MeJA treatment,indicating that GhAOP2-like might improve the resistance of upland cotton by participating in JA signaling pathway.

In order to explore the function of GhERF105-like in the development of pigment glands or gossyol metabolism of cotton,the GhERF105-like gene of Zhongmiansuo 12 was cloned by RT-PCR,and the bioinformatics analysis and expression patterns were analyzed.The results showed that the coding region of GhERF105-like was 711 bp,encoding 236 amino acids.The molecular weight of GhERF105-like was 26.3 ku,and the theoretical pI was 7.72.The predicted secondary structure of GhERF105-like was 22.88% α-helix,13.14% extended strand,3.81% β-turn and 60.17% random coil.GhERF105-like had an AP2 domain at 91—155 aa.Two threonine,one tyrosine and one serine phosphorylation sites were predicted to be located in the AP2 domain.Phylogenetic analysis showed that motifs of GhERF105-like and its homologous proteins were highly conserved in different species.GhERF105-like motifs were consistent with homologous proteins in Gossypium hirsutum,G.mustelinum,G.darwinii,G.raimondii and G.barbadense.The expression of GhERF105-like in each tissue of glanded cotton Zhongmiansuo 12 was significantly or extremely significantly higher than that of dominant glandless Zhongmiansuo 12.The expression levels of GhERF105-like in leaves of different glanded cotton varieties/lines were different,and were all significantly higher than the expressions of dominant or recessive glandless cotton varieties/lines.GhERF105-like was significantly up-regulated by ABA and BR,but significantly or extremely significantly down-regulated by MeJA,Eth,NaCl and PEG treatment.In conclusion,GhERF105-like is closely related to the development of pigment glands in cotton,and may regulate gland development or gossypol metabolism through ABA,BR,Eth and JA pathways.

In order to analyze a transgenic event of an upland cotton germplasm strain with low expression of a Bt gene,specific primers were used to identify the Bt gene.The PCR amplification result showed a fragment length of the Bt gene was 310 bp,which conformed characteristic primer fragment length to Bt gene of the United States.Using the germplasm strain to cross with island cotton Shengli No.1 showed that the Bt gene was dominant in F₁,and Bt-harboring plants to non-harboring plants displayed a 3∶1 segregation ratio in F₂ population.The result proved that the Bt gene was a quality trait controlled by a pair of dominant gene,which meant the foreign Bt gene was inserted into upland cotton as a locus.Furthermore,F₂ population and SSR markers were used in the gene mapping.The result revealed that the Bt gene was mapped on the 10th chromosome of cotton,and 14 pairs of primers were linked to the target gene,such as NAU5166,NAU3574,NAU456,BNL256,cgr6745,cgr5406,cgr6546,NAU7110,HAU3201,BNL1665,dPL0468,NAU5316,BNL2960,NAU3122.The Bt gene was located between the molecular markers NAU7110 and HAU3201,and its genetic distance was 0.9,4.4 cM,respectively.

To explore the biological function of cotton cytochrome P450 gene GhP450-94C1 in cotton Verticillium wilt response, and lay a foundation for cotton Verticillium wilt resistance gene mining and disease resistance breeding. A cytochrome P450 gene, GhP450-94C1, was cloned through transcriptome screening. The physicochemical properties of the gene were analyzed by bioinformatics methods. The expression pattern of GhP450-94C1 under Verticillium wilt induction was analyzed by Real-time quantitative polymerase chain reaction (qRT-PCR). Virus-induced gene silencing (VIGS) technology was used to preliminarily explore its biological function in cotton resistance to Verticillium wilt. The main results were as follows: upland cotton cytochrome P450 gene, GhP450-94C1, was obtained by cloning. The open reading frame (ORF) was 1 503 bp, encoding an acidic, hydrophilic and unstable transmembrane protein with 500 amino acids. The molecular formula was C₂₅₉₇H₄₀₂₅N₆₉₁O₇₂₅S₂₂ with a molecular weight of 57.23 ku, which was located in the endoplasmic reticulum membrane and contained a P450 domain. There was 86.45% probability of signal peptide; the secondary structure prediction showed that the protein contained 24 α-helixes and 8 β-sheets. This gene responds to Verticillium wilt infection, and after inhibiting its expression, the sensitivity of plants to Verticillium wilt is enhanced. GhP450-94C1 is a positive regulator of cotton resistance to Verticillium wilt.

To eradicate Verticillium wilt of cotton,identification of resistant genes and breeding of resistant cotton varieties are the best methods at present.We used MEGA 5.2 and other related software to construct phylogenetic trees of the proteins encoded by WRKY7 genes in Gossypium hirsutum and Arabidopsis thaliana and Oryza sativa.The induction and response of GhWRKY7 gene to Verticillium dahliae and its mechanism were elucidate by subcellular localization,virus-induced gene silencing,qPCR and GUS reporting system analysis.The results showed that GhWRKY7 and AtWKRY7 were highly homologous and belonged to Group Ⅱ.GhWRKY7 was located in plant nucleus,and the relative expression of GhWRKY7 in cotton leaves and root organs was significantly higher than that in stems.GhWRKY7 gene expression was significantly upregulated after 24 h of inoculation.The GhWRKY7-silenced plants showed higher susceptibility to Verticillium dahliae infection compared to the control(Expression TRV empty vector plants),suggesting that GhWRKY7 gene positively regulated cotton disease resistance.Compared to the control,the expression levels of disease-resistance related genes,including GhPR1,GhPR3,GhPR4,GhPR5,GhPDF1.2,GhPAL1 and GhCYP71B36 in GhWRKY7-silenced plants significantly decreased after inoculation,indicating that GhWRKY7 improved the disease resistance of the plants due to increased the expression levels of disease-resistance related genes.Transient expression analysis of GhWRKY7 gene by constructing GUS reporter vector in tobacco cells revealed that GhCYP71B36 gene could specifically bound to cis-element of GhCYP71B36 promoter and transcriptionally activated downstream GhCYP71B36 expression,thus improved the disease resistance of cotton.In conclusion,GhWRKY7,as a transcription factor that positively regulates Verticillium wilt resistance in cotton,is involved in the expression of downstream disease-resistance related genes such as Camalexin synthesis,thereby improving plant disease resistance.Therefore,GhWRKY7 can be used as a candidate gene for cotton resistance breeding and provide security for cotton production.

In order to analyze the salt tolerance of ScALDH21 transgenic cotton at seedling stage,T₄ transgenic ScALDH21 cotton lines and its recipient Xinnongmian 1 was used as the research material,and the research were executed under field and greenhouse conditions.In field condition,the plant materials were planted with 0.4%,0.5% and 0.6% salt soil in stripe pots,respectively.The growth phenotype,the chlorophyll content and survival rate of each transgenic lines and its recipient under different salt concentrations were observed and monitored,and their net photosynthetic rate,stomatal conductance,transpiration rate and instantaneous water use efficiency were measured.In greenhouse condition,the contents of MDA,POD activity,lignin,Na⁺,K⁺ and Na⁺/K⁺ were determined after 150 mmol/L NaCl and water treatment respectively.Compared with the recipient cotton,the survival rate,chloroplast content,net photosynthetic rate,stomatal conductance,transpiration rate and instantaneous water use efficiency of ScALDH21 transgenic cotton were increased under salt stress.MDA content decreased,POD enzyme activity increased,lignin content increased.From the aspect of morphology and physiological indexes, it was verified that the transgenic ScALDH21 cotton improved the salt tolerance of the plant at the seedling stage.

Gossypium barbadense L.,has superior cotton fiber quality,good abiotic stress tolerances,and high resistance to verticillium wilt and other biotic stress.It has always been one of the critical research subjects of germplasm improvement and utilization of Gossypium hirsutum L.by recombining exogenous DNA from G.barbadense L..The conventional genetic hybridization technology at genome level,biolistics transformation and pollen tube method for introducing exogenous G.barbadense DNA,and substitution line technology at chromosome level were reviewed.The assessment of attempts from geneticists and breeders in various genetic recombination strategies to deal with the genetic barriers of genetic recombination for these two species were conducted.Chromosome substitution line with G.hirsutum background carrying individual G.barbadense chromosome established by breeders is a prospective solution to develop stable germplasm with elite agronomy traits and/or biotic and abiotic stress tolerances,which will facilitate upland cotton germplasm innovation harboring gene resource from G.barbadense,then accelerate the cotton germplasm improvement.

Appropriate reduction of nitrogen fertilization is an important way to deal with serious nitrogen loss and low fertilizer-nitrogen use efficiency in field ecosystem in China.However,it is unclear whether reducing nitrogen fertilizer input can maintain the yield of densely planted cotton after oil.Taking the post oil direct seeding cotton field in Dongting Lake area as the research object,compare the yield,photosynthetic characteristics of main stems and leaves,nitrogen use efficiency and other differential characteristics of cotton under two-year nitrogen fertilizer reduction and deep application,and analyze the feasibility of nitrogen fertilizer reduction and deep application,so as to provide a scientific basis for the formulation of fertilization management measures in direct seeding cotton fields after oiling.From 2018 to 2019,the deep application test of nitrogen fertilizer reduction was carried out in Maowan base of Hunan Institute of Cotton Science,Changde City,Hunan Province.Five nitrogen fertilizer application treatments(0,90,180,270,360 kg/ha)and two deep application treatments(5,15 cm)were set up.Samples were taken during the main growth period of cotton to measure the net photosynthetic rate,soluble sugar and starch content of functional leaves of main stems,the aboveground plant biomass and NPK content of each part.To study the effects of deep application of nitrogen fertilizer reduction on the yield,photosynthetic characteristics,aboveground plant biomass,NPK accumulation and distribution of densely planted cotton after oil.With the increase of N-fertilizer application rate,the Pn,soluble sugar and starch content of functional leaves of cotton main stem and the accumulation of NPK in cotton plant increased first and then decreased in the middle and late stages,all of which were the highest at 270 kg/ha;the apparent utilization rate of nitrogen fertilizer,agronomic utilization rate,partial productivity and production efficiency are opposite;in addition,under the same nitrogen application rate,the indicators of deep 15 cm treatment were higher than that of shallow 5 cm treatment.Reasonably reducing the amount of nitrogen fertilizer combined with deep application treatment is conducive to the accumulation and balance of biomass in vegetative and reproductive organs of cotton plants,as well as the absorption and distribution of nitrogen,phosphorus and potassium,so as to promote the improvement of yield and quality.Based on the dense planting mode after oiling,it is recommended that the amount of nitrogen applied to cotton should be 180-224 kg/ha and the depth of fertilization should be 15 cm.

To clarify the function of the receptor gene for Gossypium hirsutum L. cytokinin receptor (GhCRE1) in the regulation of plant growth and development, and to provide material for studying the molecular mechanism of the GhCRE1 gene in regulating seed germination in Gossypium hirsutum, ultimately improving seed germination rates at the molecular level. The GhCRE1 gene was cloned and the fragment was ligated to the pCUbi1390 vector containing the Ubiquitin promoter using gene overexpression technology to construct GhCRE1 overexpression vector, while the sgRNA fragment of the Arabidopsis CRE1 gene was designed by CRISPR/Cas9 gene editing technology and the sgRNA nucleotide sequence was synthesized to construct the Arabidopsis CRE1 gene targeting knockout vector. The above two vectors were transformed into Agrobacterium and the inflorescence dip method was used to create GhCRE1 overexpression strains and gene editing loss-of-function mutants using Arabidopsis thaliana as the recipient material. Screening and identification by PCR and qPCR techniques, then, two strains of GhCRE1 Arabidopsis over-expressed and the CRE1-deficient Arabidopsis strains were obtained. After comparing the germination potential of harvested seeds with that of wild-type seeds on 1/2 MS medium, further clarify the function of GhCRE1 gene in affecting seed germination of Gossypium hirsutum. The results showed that the Arabidopsis GhCRE1 overexpression strains and CRISPR gene editing loss-of-function mutant strain were successfully obtained, after compiling the data related to seed germination potential, it was found that on the 2nd day, the germination potential of the two overexpression transgenic strains increased by about 15% and 11%, respectively, compared with the wild type, while the Arabidopsis CRE1 loss-of-function mutant germination potential decreased by about 15%. This study revealed that GhCRE1 gene overexpression significantly increased germination speed in Arabidopsis.

LMI1 is a key gene in the regulation of serrated leaf development.In order to study the mechanism of okra leaf development in cotton,we cloned GaLMI1-like gene and GaLMI1-like promoter by PCR amplification technology from Shixiya 1,which is an Asia cotton species of a genome.The sequence lengths were 681,1 439 bp,respectively.Domain analysis found that GaLMI1-like contained a homeobox domain.Then,we constructed a reorganization overexpression vector p6MYC-GaLMI1-like.The transgenic Arabidopsis showed incised leaf shape when GaLMI1-like was overexpressed in Arabidopsis.Moreover,we analyzed the cis-acting elements of the promoter sequence of GaLMI1-like.It was found that it had basic acting elements,such as CACA-box and TATA-box,and also had photoresponsive element,root,stem and mesophyll-specific expression elements.GUS fusion expression vector of GaLMI1-like promoter was constructed and transformed into Arabidopsis thaliana.GUS staining results showed that the promoter could drive GUS gene expression in root column,stem and leaf,and the expression was especially high in leaf.These results indicated that GaLMI1-like had the function of regulating the formation of incised leaves,and its function was realized by the strong expression of GaLMI1-like regulated by GaLMI1-like promoter in cotton leaves.

The aim of this study was to explore the effect of applying microbial fertilizer on salt reduction and the response of cotton growth in coastal saline cotton field. The mild and moderate saline cotton fields in the coastal area of the Yellow River Delta were selected. Cotton field experiments were conducted with different kinds of biological products and different amounts of microbial fertilizer. Soil salinity, cotton plant height, SPAD value, canopy NDVI and other data were collected. The effect of microbial fertilizer on reducing salt in saline cotton field and its response to cotton growth were systematically analyzed. The results showed that different kinds of biological products had a certain effect on salt reduction, and the salt content in surface and underlayer, rhizosphere and between ridges decreased. In mildly saline cotton field, soil salt content decreased between 7.03% and 35.06%, plant height increased by 9.76% to 15.40%, SPAD value increased by 12.97% to 22.64%, and NDVI value increased by 12.58% to 19.85%. Microbial fertilizer treatment had the best effect. Soil salt content decreased by 6.18% -31.85%, plant height increased by 10.02% -17.12%, SPAD value increased by 13.67% -15.55%, and NDVI value increased by 9.22% -18.69% in moderate salinized cotton fields. The effect of microbial fertilizer on reducing salt was the most obvious. In general, the salt reduction and growth promotion effects of the application of microbial fertilizer were better than that of organic matter+beneficial microbial treatment, and better than that of commercial organic fertilizer treatment. The recommended application amount of microbial fertilizer was 1 500 kg/ha. This study proved that the application of microbial fertilizer in coastal saline cotton field had an obvious effect of reducing salt, and had a positive effect on the growth of cotton. The results provided a reference for the biological improvement of coastal saline farmland.

In order to explore the effects of water and planting density on nitrogen use efficiency of cotton, a field experiments was carried out in Qingyuan Experimental Station of Agricultural University of Hebei between 2019 and 2020. Nongdamian 601 was taken as the experimental material, and two-factor split zone experiment was adopted, with water treatment as the main area:W1(60%-70% soil relative water content) and W2(40%-50% soil relative water content), the secondary areas were planting densities D6(60 000 plants/ha), D9(90 000 plants/ha) and D12(120 000 plants/ha). The nitrogen accumulation, distribution and yield of cotton organs under different water and planting densities were measured.The results showed as follows:Under different water treatments, the total nitrogen accumulation and reproductive organs in cotton was the highest under high density condition, and the trend was consistent during the two years. Compared with the drought treatment, the nitrogen accumulation and total nitrogen accumulation in the reproductive organs of the control group were significantly increased, but there was no significant difference in the proportion of nitrogen distribution in the reproductive organs among different water treatments. The yield of seed cotton increased with the increase of planting density.Under the same density treatment, the seed cotton yield under the drought condition in 2019 was 13.74% lower than that of the control group, and the seed cotton yield under the drought condition in 2020 was only 2.54% lower.Correlation analysis showed that the accumulation and distribution of nitrogen in various organs of cotton were significantly positively correlated with the yield of seed cotton and boll number per unit area.In conclusion, reducing irrigation amount and increasing planting density(90 000-120 000 plants/ha) are effective ways to save water and increase nitrogen uptake in local cotton.

In order to elucidate the mechanism of thousand kernel weight (TKW) reduction in ims-15, the transcriptomes of 30 days post anthesis (DPA) embryos of ims-15 and its near isogenic line Ji737 were compared, and the sequencing results were verified by Real-time quantitative PCR. A total of 4 239 differentially expressed genes (DEGs) were screened, including 2 229 (52.6%) up-regulated genes and 2 010 down-regulated genes(47.4%) in ims-15. GO functional enrichment analysis showed that the DEGs were significantly enriched in seven items of biological processes such as cell wall macromolecular catalysis, photosynthesis-light harvesting, cell wall macromolecular metabolism and photosynthesis, and in four items of molecular functions such as chlorophyll binding, tetrapyrrole binding and iron ion binding, and in the nine items of cellular components such as photosystem I, thylakoid part, photosynthetic membrane and thylakoid. three categories of biological processes, molecular functions, and cellular components. In the category of biological processes, seven items such as cell wall macromolecular catalysis, photosynthesis-light harvesting, cell wall macromolecular metabolism and photosynthesis were enriched. And in the category of molecular functions, four items such aschlorophyll binding, tetrapyrrole binding, and iron ion binding were enriched. And in the category of cellular components, nine items such as photosystem I, thylakoid part, photosynthetic membrane and thylakoid were enriched. Down-regulated genes annotated in photosynthesis-related items accounted for 85.11% of all DEGs in these items.And all of the DEGs annotated in photosynthesis-light harvesting, chlorophyll binding and photosystem were down regulated in ims-15. KEGG enrichment analysis showed that 20 metabolic pathways, including photosynthesis antenna protein, flavonoid biosynthesis, phytohormone signal transduction, phenylpropanoid biosynthesis, MAPK signaling pathway, glycolysis/gluconeogenesis, starch and sucrose metabolism, were significantly enriched.The enrichment of photosynthesis-antenna protein pathway was the highest and the 18 DEGs enriched in this pathway were down regulated in ims-15. The number of DEGs enriched in the plant hormone signal transduction pathways was the most. Transcription factor analysis obtained a large number of transcription factors that affect kernel weight formation, such as WRKY, Dof, AP2/EREBP et al. The correlation coefficient between qRT-PCR and RNA-seq data was 0.955 9, which verified that the results of RNA-seq were accurate and reliable. Based on the results of GO functional enrichment analysis, KEGG enrichment analysis and transcription factor analysis, some metabolic pathways closely related to the development of grain weight had been clarified. Especially, the embryonic photosynthetic pathway played an important role in the formation of kernel weight. It was discovered that plant hormone signal transduction pathways and transcription factors played an important regulatory role in kernel weight formation.

In order to deeply analyze the molecular genetic mechanism of cotton yield related traits and explore effective molecular markers and genes, a high yield cotton cultivar Jifeng 914 was crossed with a super fiber quality inbred line Jifeng 817. Four traits including boll weight (BW), lint percentage (LP), seed index (SI) and fruiting branch number (FBN) were investigated in F₂ and F₃ populations, and QTL mapping was conducted. The results showed that the four traits of Jifeng 914 were larger than that of Jifeng 817, and normal distribution were observed in the offspring populations. Significant positive correlation was observed between SI and BW, and significant negative correlation existed between SI and LP. A total of 50 QTL were mapped on 22 chromosomes, including 8 QTL for BW, 20 QTL for LP, 15 QTL for SI and 7 QTL for FBN. The largest phenotypic variation explanation rate (PVE) reached 12.96%. Two QTL, qBW-A11-1 and qLP-A6-1 could be repeatedly detected in F₂ and F₃ populations, which were named as stable QTL. In addition, 41 genes were annotated in major effect (PVE ≥ 10%) or stable QTL regions, mainly participating in plant cell wall synthesis or cellulose biosynthesis. Based on the transcriptome information, Ghir_D03G005440.1 enconding Cinnamoyl-CoA reductase 2-like protein expressed higher level in TM-1 than that in Hai 7124. This gene located in the major QTL qSI-D3-1, indicating that it may be involved in the regulation of SI. This study provides more genetic basis for the detection of high cotton yield traits.

In order to study the molecular genetic basis of the interaction among the earliness, yield and fiber quality of upland cotton, a Recombinant inbred line(RIL) population was constructed with CCRI36 as the female parent and G2005 as the male parent. For 6 consecutive years, 13 characters related to the earliness, yield and fiber quality of the RIL population were investigated, and the correlation among the three types of characters was analyzed. The genetic basis was analyzed based on the results of QTL mapping. The results showed that the materials with better earliness had lower boll weight and poorer fiber quality;the materials with better fiber quality had a relatively poor yield character. A total of 494 QTL were detected, 68 QTL overlap regions affected at least two types of characters at the same time, including 11 regions that affected earliness, yield and fiber quality, 11 regions that affected earliness and yield, 39 regions that affected earliness and fiber quality, and 7 regions that affected yield and fiber quality. Only 24 of the 68 QTL-overlap regions had the same additive effect direction. In addition, 99 multi environment epistatic QTLs were detected, 45 of which overlapped with additive QTLs.

To explore the regulating effect and potential of yield increase of prohexadione calcium on cotton,and to provide scientific basis for high-yield cultivation in Xinjiang cotton area. This experiment was conducted in Dafeng Town,Hutubi County,Xinjiang,and Sidaohezi Town,Shawan County,Xinjiang in 2019. A randomized block test design was used. The test used DPC as a control(CK)and four prohexadione calcium doses were set:T1(750 g/ha),T2(1 350 g/ha),T3(1 950 g/ha)and T4(2 550 g/ha), at the bud stage,early flowering stage,topping spray in divided doses before and after topping. Dry matter accumulation,agronomic characters,yield and quality of cotton were investigated. The daily growth of main stem increased first and then decreased with time after spraying prohexadione calcium,and reached the peak after two applications(the early flowering period).The regulation effect of prohexadione calcium on plant height had a certain dose effect. With the increase of the dose,it increased first and then decreased. The inhibitory effect was most significant at the T3 level. Boll opening period of cotton plant height at the two test sites increased by 3.35%,7.15% compared with the control. At the same time,prohexadione calcium increased the biomass accumulation of cotton above ground parts,and the biomass of T3 reproductive organs was increased by 35.07%,34.35% at the two test points respectively;the leaf branches treated with prohexadione calcium were 3.89-14.34 cm longer than the control,and the first fruit branch were higher than the control length 4.83-10.16 cm,the fourth fruit branch were higher than the control length 3.34-12.34 cm; compared with the control, the number of bolls per plant of fruit branches of cotton treated with prohexadione calcium increased by 4.90%-23.04%,and the yield of seed cotton increased by 226.14-907.87 kg/ha,both of which were the same T3 has the highest yield,with lint cotton yields of 2 885.66,2 879.53 kg/ha,while prohexadione calcium has the effect of optimizing fiber quality. Spraying 1 950 g/ha of 5% prohexadione calcium during the whole growth period could significantly control cotton plant height,shape high-yield plant types,effectively coordinate the relationship between vegetative growth and reproductive growth,significantly increase cotton yield and improve fiber quality.

The purpose of this study was to investigate the relationship between the changing of chemical detopping cottons' endogenous hormones and spatial distribution of boll branch. The present objective of this study was to measure four hormones,diurnal variation by Elisa inverse fourth leaf of cotton after detopping,North Xinjiang Early maturing cotton No.45 was the cotten variety,set the chemical(C)and two contrast group:no(CK)and artificial detopping(M). The results indicated that the IAA contention after chemical detopping(C)has decreased trend more obviously and rapidly than the no detopping(CK)and artificial detopping(M),and remained at a low level after 168 h;the GA₃ contention had no much difference about chemical topping(C)and no topping(CK),which provides a theoretical basis for the cotton "Secondary growth";the CTK contention of cotton after chemical(C)had slower rising speed than no topping(CK)and artificial topping(M),but it had maximum peak value,and remained at a higher level after 168 h;the ABA contention after chemical(CK)has maintained a high level until the end of the experiment. The experimental indexes'peak and bottom peak appeared in processing that proved chemical detopping could change the cotton endogenous hormone significantly inverse fourth leaf of cotton. Meanwhile chemical detopping could effectively control the cotton boll branch length,which were distributed mainly in the middle and lower branch,at the top of ball has obvious advantages compared to artificial detopping. The research provides certain theoretical basis for chemical detopped cottons'spatial distribution of boll and branch is regulated by the endogenous hormones.

In order to discover the genes related to the infection response of Verticillium dahliae in upland cotton and to reveal the emergency response mechanism of upland cotton to Verticillium dahliae infection,the proteome of cotton roots under Verticillium wilt stress was studied by iTRAQ(Isobaric tags for relative and absolute quantification,iTRAQ),using Ji 2658 as the material,and cotton roots were infected with Verticillium dahliae Vd991 for 1 h as the sample,and water treatment as the control.Forty-six differentially expressed proteins(DEPs)were screened,of which 37 were up-regulated and 9 were down-regulated.Gene ontology annotation analysis showed that the functions of DEPs were divided into 24 categories,with molecular functions accounting for 7 categories,biological processes accounting for 8 categories and cell components accounting for 9 categories.Pathway analysis revealed that DEPs were involved in 32 metabolic pathways.The metabolic P value of phenylalanine metabolism(0.025 308 8)was the smallest.By analyzing the functions of DEPs,it was found that energy metabolism-related proteins,tissue metabolism-related proteins,protein synthesis-related proteins,antioxidant enzymes,nucleoside diphosphate kinase(NDPK)were involved in the process of cotton resistance to Verticillium wilt in the early stage of Verticillium wilt stress(1 h).Moreover,the release of cytochrome c(Cyt-c)indicated that cotton root initiated programmed cell death(PCD)to inhibit the transmission of Verticillium wilt.

MYB transcription factor proteins are ubiquitous in plants and play an important role in biological and abiotic stresses. To explore the function of cotton MYB gene, the paper cloned a MYB gene from cotton leaves using homologous cloning technology, and analyzed its bioinformatics and expression under different stresses. The results showed that a new MYB transcription factor gene GhMYBPA1 (gene entry site XM_016869420) was successfully cloned from Gossypium hirsutum cv. Zhongmian 35, the full of cDNA was 825 bp in length with a 630 bp ORF, which encoded a 210 amino acids peptide. Bioinformatics analysis results showed that the molecular weight of GhMYBPA1 was 20.183 ku, GhMYBPA1 contained two conserved DNA-binding domains at N-terminal, which belonged to R2R3-MYB transcription factors. Amino acid homology analysis showed that GhMYBPA1 had higher identification with GaMYB12-like from Gossypium arboreum. Based on qRT-PCR analysis, GhMYBPA1 was constitutively expressed in cotton roots, stems, leaves, and it was dominantly expressed in flowers and then was leaves. Moreover, the results of plant treated with various stresses showed that the expression of GhMYBPA1 gene changed under high salt, low temperature and drought stress, it was suggested that GhMYBPA1 might play an important regulatory role in the abiotic stress process of cotton. The results could lay a theoretical foundation for further researches on the function of GhMYBPA1 gene.

In order to elucidate the cotton GhMADS43 gene in shoot apical meristem and floral meristem development and to screen for proteins that interact with this geneby yeast two hybrid system.This study cloned the coding sequences of GhMADS43 and analyzed the expression of this gene in different tissues and shoot apical meristem periods.The results showed that the GhMADS43 gene was 696 bp in length and encoded 231 amino acids, which was similar to FUL in other plant species. According to specific expression analysis, GhMADS43 gene had higher expression in roots and apical buds, and the expression of the gene was higher in infinite fruit branches cotton CCRI50 than that of one fruit branches cotton Zaoling No.1.We also constructed bait vectorsp pGBKT7-GhMADS43 and transformed it into the yeast strain Y2HGold. The results showed that all the Y2HGold strains containing the recombinant plasmid grew well on the SD/-Trp media. It suggested that the recombinant bait proteins were no toxic to yeast. The transformed yeast strains developed blue plaques on the SD/-Trp/X-α-Gal media. And they could not grow on the SD/-Trp/-Ade/-His media with different concentration of 3-AT. The results indicated that the GhMADS43 gene had no self-activating activity. The constructed bait vector can be used for further screening of proteins interacting with GhMADS43 from yeast two-hybrid cDNA library in cotton. The study plays the foundation for analyzing of the gene function and regulatory pathways in shoot apical meristem and floral meristem development in the future.

Pyruvate dehydrogenase phosphatase GhPDP is closely related to the regulation of cotton male sterility energy. To obtain a large number of active GhPDP proteins, the prokaryotic expression conditions the target protein were optimized in this experiment. The cDNA sequence encoding the GhPDP gene was inserted into pET-22b to construct a recombinant expression vector pET-22b-GhPDP, which was transformed into Escherichia coli Transetta(DE3) and induced expression by IPTG. Single factor test and orthogonal experiment were used to optimize the induction temperature, induction time, final IPTG concentration and bacterial cell density OD₆₀₀, using SDS-PAGE and GeneTools gel analysis software to analyze the expression of the target protein and explore the best conditions for soluble protein expression. The results showed that successfully constructed the recombinant expression vector pET-22b-GhPDP. The target protein was expressed in Transetta(DE3) strain, mainly in the form of inclusion body. The results of single factor experiments showed that the soluble expression level of the target protein was the highest under the conditions of induction temperature of 20℃, induction time of 5 h, IPTG final concentration of 0.10 mmol/L and cell density of OD₆₀₀ of 0.8. Through orthogonal analysis, it was concluded that the intensity of the factors affecting the soluble expression of the target protein from high to low as induced temperature, final IPTG concentration, induction time, and bacterial cell density OD₆₀₀. Based on the results of single factor experiment and orthogonal experiment, the optimal induction conditions were determined to be a bacterial cell density of OD₆₀₀ of 0.8, an induction temperature of 20℃, an induction time of 5 h, and IPTG final concentration of 0.10 mmol/L.

PP2C is an important group of protein phosphatase, which is widely involved in different adverse stress responses. In order to understand the functions of cotton PP2C genes in drought response, GaPP2C24 gene was cloned from G. arboreum cultivar Shixiya-1 by RT-PCR. Bioinformatics analysis of this gene were carried out and its expression patterns in different tissues and in drought stress response were studied by quantitative Real-time PCR. The results showed that the CDS sequence of GaPP2C24 gene was 1 251 bp in length and encoded 416 amino acids. Multiple sequence alignment showed that GaPP2C24 had a high similarity with amino acid sequences of other plants. The similarity between protein GaPP2C24 and cacao tree (EOY33930.1) and jute (OMO91132.1) was 85% and 84%, respectively. Phylogenetic tree analysis revealed that GaPP2C24 was much closer to jute that of the same genus malva, followed by cacao tree. The results of Real-time quantitative PCR showed that the expression of GaPP2C24 could be detected in cotyledons, hypocotyls, radicles, leaves, stems and roots. Its expression level was the highest in stems, followed by roots. The expression of GaPP2C24 was induced by drought stress. The expression of GaPP2C24 in roots and leaves reached the highest level after 1 h of drought treatment, with 53.9 and 6.9 times of the control, respectively. The results suggested that GaPP2C24 gene played an important role in cotton drought response.

To investigate the function of GhNAC201 in cotton pigment gland development,RT-PCR was performed to isolate the coding sequence(CDS)of GhNAC201 from glanded cotton Zhongmiansuo 12. Gene expressions of glanded and glandless cotton at different developmental stages,in different tissues and under various hormones treatment were analyzed by qRT-PCR. CDS of GhNAC201 was 948 bp in length and encoded 315 aa,with a theoretical pI 5.97. 27.94% alpha helix,5.40% beta turn,18.10% extended strand and 48.57% random coil were predicted in protein secondary structure. 3D structure of GhNAC201 had been modelled by rice stress-responsive NAC1 with model dimensions of X:55.229 Å,Y:36.150 Å,Z:46.112 Å. GhNAC201 contained a NAM domain at 30-163 aa,and 5 threonine,6 serine and 4 tyrosine phosphorylation sites were predicted within the NAM domain. Subcellular localization prediction showed that GhNAC201 had a certain probability to locate in secretory pathway or other subcellular structures other than mitochondrion and chloroplast. The homology of GhNAC201 homologous proteins in different species was relatively high,and 5 motifs were highly conserved.The motif pattern of GhNAC201 was consistent with its homologous proteins in Gossypium arboretum and G. raimondii. Expression levels of GhNAC201 in the glanded cotton Zhongmiansuo 12 were extremely significantly higher than those in dominant and recessive glandless cotton and could be extremely significantly up-regulated by ABA,BR,GA and MeJA. The spatial-temporal expression pattern of GhNAC201 in Zhongmiansuo 12 tended to be consistent with the spatial-temporal distribution of gland density. In conclusion, GhNAC201 was closely related to the development of pigment glands in cotton,and might regulate gland development or gossypol metabolism through ABA,BR,GA and JA pathways.

The amount of miRNA identified in cotton is still limited and the role of miRNAs in early-maturing cotton remains to be proved and revealed. In order to enrich the amount of miRNA in cotton and reveal the important roles of miRNA in the development of early-maturing cotton, two small RNA libraries were constructed using Shizao 1, a early-maturing cotton variety and Jimian 958, a medium cotton variety by high-throughput sequencing technology. As a result, a total of 131 miRNAs were identified, including 64 known miRNAs belonging to 41 miRNA family and 67 novel miRNAs; 54 of the 67 novel miRNAs belonged to known miRNA family and 13 novel miRNAs belonged to novel miRNA family. The expression of miRNAs were compared in the two cotton varieties, it found that 39 miRNAs showed obviously different expression(>2 fold) between the two varieties, which accounted for 29.8% and focused on low- and mid-expressed miRNAs. In the top 15 highly expressed miRNAs, only ghr_21(miR398 family) showed obviously different expression(3.4 fold). The expression of miRNAs belonging to the same family were compared and found different expression pattern in the two varieties. Above results revealed that miRNAs had important roles of earliness in the development of early-maturing cotton, which provide a basis for further study on the role of miRNA in the regulation of cotton prematurity.

Germplasm resources are the basis of genetic breeding research. Analyzing the genetic diversity of germplasm resources can provide a certain theoretical basis for parental selection. Genetic diversity, correlation, principal component and cluster analysis were performed on 11 phenotypic traits of 500 cotton germplasm resources materials. The results showed that the average coefficient of variation of the 11 traits was 9.69%. And there were 6 trait which coefficient exceeded 10.0%. The variation coefficient of the number of bells (20.14%) was maximum, and the variation coefficient of the fiber uniformity (1.81%) was minimum. The genetic diversity index of each trait was in the range of 1.71-2.06. The genetic diversity of boll number was highest and the genetic diversity index of growth period was lowest. Correlation analysis showed that there were different correlations between the various traits. Fiber length was significantly positively correlated with fiber uniformity, fiber strength and growth period but it was significantly negatively correlated with the micronaire value. The fiber strength was significantly positively correlated with the micronaire value and growth period. The lint percent was significantly positively correlated with the fiber length, fiber uniformity, and micronaire value but significantly negatively correlated with the seed index. The boll weight was significant positive correlated with lint percent, fiber length, fiber uniformity and growth period. By the principal component analysis, 5 principal components were extracted which cumulative contribution rate was 73.677%. The first principal component mainly related to fiber length, fiber strength, and fiber uniformity; the second was mainly related to the lint percent; the third was mainly related to the micronaire value; the fourth was mainly related to the number of fruit branches and the fifth was mainly related to the boll weight. Cluster analysis classified 500 cotton germplasm materials into four clusters at genetic distance 1.33. Cluster Ⅰ contained only one material, which character was high plant height, high boll weight, low lint percent, high seed index. Cluster Ⅱ contained 73 materials, which characters were few boll, low lint percent, and poor fiber quality. The Cluster Ⅲ contained 122 materials, which characters were high boll weight, high lint percent, long fiber and high fiber strength. Cluster Ⅳ contained 304 materials, which boll number, boll weight, lint percent, fiber length, and fiber strength were moderate.50 germplasm resources were extracted to construct the core collection. The variable coefficient and variance of each trait in core collection were higher than ones in the original population. Indicating that the core germplasm has better heterogeneity. The core collection heterogeneity was better than the original population. The core collection retains the genetic information of germplasm resources with the least number of resources, which not only reduces the workload of germplasm conservation, but also facilitates the selection of breeding parents.

Experiment was carried out to study the effects of five different nitrogen rates(120, 150, 180, 210, 240 kg/ha) applied once fertilization when seeing first flower on cotton yield and quality saw in summer in five trials where are in the main cotton producing areas of Hubei Province in 2018.The results showed that the cotton lint yield increased firstly and then decreased with the increase of nitrogen application rate. And the highest lint yield treatment was N150 by 1 498 kg/ha without significant difference compared with the five nitrogen treatments. That means a relatively stable yield could be obtained within large range nitrogen fertilizer level; There was no significant difference in fiber quality among the five different nitrogen treatments, did also in cotton growth characteristics and field growth. Therefore, under the conditions of cotton planting in summer, it can be obtained stable yield and quality when nitrogen application rate is among 120-240 kg/ha. In order to improve the efficiency of cotton planting according to the strategy of ″reducing investment without reducing production″, and it is feasible to apply nitrogen rate without exceeding 150 kg/ha in Hubei Province.

The LEA genes often exist as a large gene family in higher plants, and have the pivotal role in plant growth and development, and various stress response. To further study the function of cotton LEA genes, two LEA genes were isolated from a cotton fiber-normolized cDNA library, which were named GhLEA and GhLEAG1 (GenBank accession number KF906314 and KF906316). GhLEA contained an open reading frame(ORF) of 948 bp, encoding a protein of 315 amino acids, with a molecular weight of 35 ku, and a theoretical isoelectric point of 4.4. GhLEAG1 contained an open reading frame(ORF) of 756 bp, encoding a peptide of 251 amino acids, with a molecular weight of 27 ku, and a theoretical isoelectric point of 10.76. Phylogenetic tree showed that GhLEA and GhLEAG1 had different motif elements, belonging to LEA2 and LEA3, separately. Quantitative RT-PCR analysis showed that both GhLEA and GhLEAG1 had the expression peak at 20DPA, and the expression of GhLEA in root was the highest, but the expression of GhLEAG1 was the highest in stem. The expression patterns of the two genes were significantly different under salt stress and PEG. In addition, the two genes were also induced by ABA, the expression level was increased first and then decreased. The data indicated that GhLEA and GhLEAG1 might play an important role in the abiotic stress response of cotton. This study will provide an important reference for further research on the molecular function of LEA in cotton.

In order to excavate the role of exogenous silicon(Si) in salt stress response and explore the regulation mechanism of salt stress response, the cotton variety Xinluzao 45 was used as test material, and was treated with 3 levels of salt(NaCl:0, 100, 200 mmol/L) and 2 levels of silicon(K₂SiO₃:0, 262.3 mg/L). The changes of seedling growth, osmotic regulation, reactive oxygen, malondialdehyde(MDA) content and electrolyte leakage rate were measured after the treatment of exogenous Si, and the effect of silicon in relieving the inhibitory effect of NaCl on cotton seedlings were studied. The results showed that, compared to CK, the fresh leaf weight, dry leaf weight, stem fresh weight, stem dry weight, root dry weight, root cap ratio, root activity, malic acid and citric acid of cotton seedlings all showed a downward trend along with the increase of NaCl levels, while the contents of free proline(Pro), hydrogen peroxide(H₂O₂) and malondialdehyde, the rate of electrolyte leakage and the oxygen free radical(O₂^·) production presented a upward trend, and SS and free amino acid contents increased first and then decreased. The biomass and osmotic regulator of cotton seedlings increased obviously in the same salt treatment after application of exogenous silicon, while the rate of O₂^· production, electrolyte leakage rate, the contents of hydrogen peroxide and malondialdehyde in cotton seedlings decreased obviously. Comprehensive analysis showed that the salt stress could inhibit the growth of cotton seedlings, and with the increase of salt concentration, the inhibition effect and osmotic stress degree aggravated. The exogenous silicon increased the accumulation of osmotic regulation substances and reduced the accumulation of active oxygen in cotton seedlings, so it could alleviate the inhibiting effect of salt stress on cotton seedling growth and improve the salt resistance of cotton seedlings.

Since Verticillium wilt causes great loss in cotton production every year, it is an economical and effective method to identify resistance genes for breeding new varieties to control Verticillium wilt. A WRKY transcription factor gene was cloned from upland cotton, named GhWRKY48. The full-length of GhWRKY48 cDNA was 882 bp,encoding 293 amino acid residues with a predicted molecular mass of 32.68 ku and an isoelectric point (pI) of 6.10. qPCR analyses showed that GhWRKY48 gene constitutively expressed in root, stem and leaf, and preferentially done in stem. In addition, GhWRKY48 expression could be induced by V. dahliae infection, jasminic acid and salicylic acid. To dissect the defense function, GhWRKY48 -silenced plants were developed by virus-induced gene silencing techniques, which were then inoculated with V. dahliae. The results showed that the resistance of these silenced plants significantly increased compared to the control. The rates of diseased plants and disease index in GhWRKY48 -silenced plants were lower than in the control. All together, GhWRKY48 was a negative transcription factor participating in plant resistance to V. dahliae. Therefore, the GhWRKY48 gene could be used as an ideal candidate gene for cotton defense breeding.

In order to study the response of cotton phosphatidylethanolamine-binding protein GhTFL1b gene to stress and physiological function, we cloned the GhTFL1b gene from upland cotton, and then analyzed its expression pattern and protein interaction. The results of promoter structure prediction using PlantCARE software revealed that GhTFL1b promoter contained MeJA responsiveness element, abscisic acid(ABA) responsiveness element, MYB binding sites involved in drought inducibility and light responsiveness elements. Meanwhile, Quantitative Real time PCR result indicated that the expression profile of GhTFL1b was induced by salicylic acid(SA) negatively, while it could be regulated by salt(NaCl) positively. Yeast two hybrid approach were under taken to elucidate the protein interaction of GhFD and GhTFL1 subgroup, and the experiments showed that GhFD could interact with GhTFL1b but not with GhTFL1a, GhTFL1b and GhTFL1c in cotton. Taken together, the result suggested that GhTFL1b might be involved in the regulation of response to abiotic stresses(SA and salt).

To explore the effect of the application of cotton biochar on cotton field in grey desert soil in arid area, the effects of application methods of cotton biochar (annual cotton biochar NPKC, constant nitrogen+1.5 cotton biochar NPKBc1.5, low nitrogen+1.5 cotton biochar N_low PKBc1.5, constant nitrogen+3.0 cotton biochar NPKBc3.0, low nitrogen+3.0 cotton biochar N_low PKBc3.0, low nitrogen) on soil physical and chemical properties of desert cotton were analyzed through the design of random field experiment. The results showed that compared with CK, the leaf area of NPKBc1.5 cotton increased significantly, and the number of buds boll increased by 45.67%.The effects of increasing nitrogen application amount and cotton biochar were different, and the cotton biochar was increased to 22.5 t/ha, and the leaf area of a single plant increased by 24.27%.The increase of nitrogen fertilization did not increase the leaf area significantly. Under normal nitrogen application, cotton stalk carbon increased from 22.5 t/ha to 45 t/ha, root canopy ratio increased by 13.3%, leaf area decreased by 11.84%. The number of buds boll increased significantly (P<0.05) while the amount of nitrogen was decreased but the amount of cotton biochar was increased. Soil structure and physical and chemical properties were improved. Compared with CK, when 45 t/ha cotton biochar was applied, the soil pH value dropped to 7.09, down 1.16%.On the basis of N low PKBc1.5 and NPKBc1.5, cotton biochar was increased by 1 times respectively, with natural water content increasing by 1.7 and 0.3 respectively, field water holding capacity increasing by 3.5 and 2.4 respectively, and saturated water content increasing by 6.2 and 1.6, respectively. The content of soil organic carbon reached the maximum value of 17.23 g/kg, an increase of 79.72% compared with the control group. The boll number and leaf area of flower buds and individual plants were significantly affected by organic carbon and bulk density, but not significantly affected (P>0.05). Water content had a positive effect on the number of buds bells. Therefore, it could be concluded that the application of cotton biochar in desert soil could make up for the deficiency of nitrogen and improve the physical and chemical properties of soil.

To explore the causes of germination during different cotton varieties to low temperature stress,used two type materials (strong chilling resistant of SCRC37,SCRC36,and chilling sensitive of Stoneville 2B,Stoneville 5A) to study the difference of physiological indexes and gene expression to low temperature stress during different germination stages. The results showed that the MDA content of sensitive varieties increased sharply,to 12℃ stress for 36 h. The content of MDA in Stoneville 2B,Stoneville 5A respectively increased by 48.17% and 50.65%,compared with SCRC 37,to 12℃ stress for 48 h. Therefore,It was suggested to be used as a selective index for seed germination of cotton to low temperature stress,which the change of MDA content to 12℃ stress for 36 h. The SOD activity and POD activity were maintained at a high level of SCRC 37 and SCRC 36 at all levels of treatment. To low temperature stress,the CAT activity of tolerant varieties was gradually surpassing that of chilling sensitive varieties. To 12℃ stress for 48 h,the content of free proline in SCRC37 from Stoneville 2B and Stoneville 5A,respectively was 33.32% and 23.27%. These results indicate that the physiological indexes were relatively stable in response to the differences of low temperature tolerance and germination of different varieties. The gene of CBF1, CBF2, CBF4, CBF6 were up-regulated more rapidly in strong chilling varieties,and the expression was reached peak when stressed to 12 h. These may be the reason why the strong chilling varieties are more sensitive and rapid in response to low temperature stress.

The purpose of the experiment is to screen suitable planting spacing distribution patterns of different cotton varieties under reasonable density planting conditions,and to provide certain technical support for the development of mechanized cotton harvesting in the Yellow River basin. Three main cultivars in central and Southern Hebei were selected:Jimian 958,Shikang 126 and Ji 863. The test set four different strains of row spacing:80(Ⅰ),75(Ⅱ),70,(Ⅲ),(100+50) cm (Ⅳ) under 75 000 plants/ha planting density,at the same time set the conventional planting density of 45 000 plants/ha to control (CK,spaced 80 cm),to study the influence of different strains of planting spacing the under reasonable density planting condition on cotton growth,yield and quality. The results showed that the effects of different planting spacing on plant height,stem width and number of fruiting branches of different varieties were different under reasonable dense planting. There was no significant difference between the bolls No. per plant in Jimian 958 and Shikang 126,but the numbers were significantly lower than CK which was in low density. The ratios of the boll-setting per plant in treatments Ⅱ and Ⅳ were significantly higher than treatments Ⅰ and Ⅲ. The bolls No. per plant in treatment Ⅳ was the highest in Ji 863,and then was treatment Ⅱ. The seed cotton yield in processing Ⅳ was the highest in Jimian 958. Which in treatments Ⅱ and Ⅳ were significantly higher than treatments Ⅰ and Ⅲ in Shikang 126. And the yields in treatments Ⅱ and Ⅳ were higher than treatment Ⅲ in Ji 863,and which in treatment Ⅲ was higher than treatment Ⅰ,but there was no significant difference. It was recommended that the suitable planting spacingn of three cotton varieties should be 75 cm and (100+50) cm row spacing.

The molecular markers related with Verticillium wilt-resistance were explored by association analysis, which could provide candidate markers for disease resistance for use in marker-assisted breeding (MAS). In 214 upland cotton materials, 237 SSR markers were assayed for polymorphism, and cluster analysis was performed by PowerMarker v3.25, population structure was analyzed by Structure 2.2, an association analysis between SSR markers and Verticillium wilt-resistance traits were performed using the Tassel 2.1 GLM(general linear model) program. The results showed that the 237 SSR markers included 695 alleles belong to 280 loci,with an average of 2.479 0 alleles per SSR marker(range, 2-6). Structure analysis showed that all the accessions could be divided into 2 subgroups. Of the SSR loci, 27 were significantly(P<0.01) associated with Verticilium wilt-resistance under GLM program in different years, among which 2 marker loci (BNL3442 and BNL1064) were found to be consistent associated with Verticillium wilt-resistance in 3 years, and the ratio of explanation on the phenotype variation of above markers was 12.10% and 8.02% respectively. These stable SSR markers in different years are likely to be closely linked to disease resistance genes and are expected to be used in the screening of cotton Verticillium wilt-resistance material and resistance gene mining.

The development of plant disease-resistant cultivars by disease-resistant gene mining becomes a most available measure. The bioinformation,virus induced gene silencing,qPCR and fungus inoculation were employed to analyze cotton WRKY resistance to Verticillium dahliae. Eight cotton orthologous WRKY genes corresponding to 8 Arabidopsis defense-related WRKYs was selected by phylogenetic tree analysis. And their expression was induced by V. dahliae infection,indicating that they participate in cotton defense to this fungus. Among these genes,six GhWRKYs showed up-regulation expression, and expression levels of GhWRKY70 and GhWRKY48 exhibited up-or down-regulation over time. Five cotton WRKY genes including showed two expression peaks at 24,72 h after pathogen inoculation. GhWRKY22 gene was chosen for further charactering its function in cotton resistance to V. dahliae infestation.GhWRKY22 expressed predominantly in stem,and its expression level was obviously increased upon pathogen infestation and responded positively to the defense signaling molecules such as salicylic acid and jasmonic acid hormones. Knockdown of GhWRKY22 gene increased susceptibility of plants to V. dahliae infection. Additionally,the expression level of defense marker genes,including PR1,PR3,PR4,PR5,PAL and PDF1.2,significantly decreased in GhWRKY22-silenced plants compared to the control plants,which suggesed that GhWRKY22 participates in cotton defense against Verticillium wilt through SA and JA signaling pathways. Taken together,these results showed that 8 GhWRKYs were identified and characterized in regulation of resistance to V. dahliae. Among them,GhWRKY22 was a positive regulation component in cotton defense,which could be regarded as a candidate gene to breed plant defense cultivars.

GhTGA1 and GhTGA9.2 of transcription factor genes were isolated by RT-PCR technology from Zhongmiansuo 12 cultivar of resistance to Fusarium oxysporum. Sequence analysis showed that the ORF of GhTGA1 and GhTGA9.2 were 1 281 bp and 1 461 bp,respectively.They encoded 405 and 486 amino acids,respectively. GhTGA1 and GhTGA9.2 all contained two conserved bZIP and DOG1 domains. They belonged to the TGA transcription factor gene of bZIP subfamily. The GhTGA1 and GhTGA9.2 expression feature were detected by Real-time quantitative PCR(qRT-PCR),the results showed that GhTGA1 gene was up-regulation expressed with the duration of the treatment of Fov,ethylene (ET) and jasmonic acid (JA). In each treatment time point,the expression of GhTGA1 gene was lower than that of Mock with the treatment of salicylic acid (SA). We speculated that GhTGA1 gene might be involved in ethylene (ET) and jasmonic acid (JA) signaling pathway and regulated resistance to Fusarium oxysporum in cotton; After the treatment of Fusarium oxysporum,the expression of GhTGA9.2 gene was lower than Mock at different treatment time points,and was down-regulated expression. In other hand,it was up-regulation expressed with the duration of the treatment of hormone (ET,JA,SA).It suggested that GhTGA9.2 might participate in stress response through hormone (ET,JA and SA) pathways. The analysis of gene expression characteristics of GhTGA1 and GhTGA9.2 provides a basis for further study.

To reveal the relationship between the key enzyme gene expression of gibberellin synthesis pathway and different tissues or stages of growth, the CCRI49 cotton was selected as material, and relative quantification Real-time PCR was developed to determine the gibberellin key gene expression of different cotton organizations in different stages of development. The results showed that, in seedling, the expression level of all gibberellin synthesis pathway genes was higher in stem, except GA2ox1 gene. In full-blooming stage, the expression of GA2ox1, GA3ox1, GID1B in root, CPS1, GA20ox1, GA3ox1, GID1B in stem and GA3ox1 in leaf had an up-regulation. In boll opening stage, the expression levels of GA2ox1 and GA3ox1 were up-regulated in root, while the remaining genes were down-regulated. The expression levels of KS and GID1B in stem were down-regulated, while the other genes were up-regulated, with GA2ox1 rise in maximum. And GA2ox1 in the leaf was down-regulated and the rest were up-regulated. In full-blooming stage, the high expression of GA20ox1 gene provided the necessary active gibberellin level for the elongation and growth of the plant stem, and the GA3ox1 gene might be associated with flowering and bell. In boll opening stage, the expression of GA2ox1 gene in root and stem gradually increased to reduce the synthesis of active, gibberellic acid. The gene expression levels of GA3ox1 and GA20ox1 in leaf tissues were raised to provide the necessary hormone levels for cotton boll growth and dehydration. The results indicated that cotton plant by changing the key enzymes gene expression of gibberellin synthesis metabolic pathway to regulate plant growth and development, and the purpose gene expression volume trends exist a certain difference along with the growth and development. It provides a theoretical basis for studying the regulation mechanism of gibberellin on upland cotton, which can help accelerate the improvement of variety and germplasm innovation.

It is of great significance to investigate the response of arginase gene to abiotic stress and its physiological function in cotton. Bioinformatics analysis showed that the ORF sequence of GhARG1 was obtained by In silico cloning and RT-PCR technique. Salt stress, PEG6000 stress and ABA, SA and MeJA treatment cotton seedlings, and GhARG1 was expressed in E. coli prokaryotic. The ORF sequence contained 1 023 bp, and encoded 340 amino acids with typical conserved domains of arginase. Quantitative RT-PCR analysis showed that the expression of GhARG1 in leaves was up-regulated under stress condition of NaCl, PEG6000, and signal molecule of ABA, SA, respectively, but down-regulated by MeJA. The coding sequence of GhARG1 was fused to prokaroytic expression vector pCold-TF, and the combinant vectors were induced by IPTG and detected by SDS-PAGE. The molecule weight of protein GhARG1 was detected to be the expected 37 ku. The activity of arginase in the recombinant strain was 6 times that of the control. The results suggested that GhARG1, which coding product has arginase activity, might be involved in the response to salinity and drought stress through ABA and SA signaling pathway in cotton, and could lay foundation for further physiological functional identification.

In order to provide references for nitrogen fertilization in field production, the effects of different NH₄⁺/NO₃^- ratio of nitrogen nutrition (NH₄⁺-N/NO₃^--N were 0/100, 25/75, 50/50, 75/25, 100/0)on morphological characteristics and dry matter accumulation, hydroponic experiments were conducted using Nongda 601 as material. The results showed that the plant height, stem diameter, leaf area and main root length of cotton increased gradually with the growth stages changed. In addition to the main root length showed a continuous elongation, the plant height, stem diameter, leaf area and dry weights of cotton increased firstly and then decreased with the increasing of NH₄⁺ concentration in the ammonium nutrient solution. The plant height of cotton seedlings was the highest at the NH₄⁺/NO₃^- ratio of 50/50 and 25/75 respectively 7, 14, 21 d after enhanced ammonium nutrient solution treatment;both stem diameter and leaf area were the largest at the NH₄⁺/NO₃^- ratio of 50/50 after enhanced ammonium nutrition treatment 7, 14, 21 d, indicating that the concentration of NH₄⁺ more than 50% could inhibit significantly the growth of cotton seedlings. In terms of dry matter, total dry weight, stem dry weight and leaf dry weight were the highest at the NH₄⁺/NO₃^- ratio of 75/25, while there was no significant difference in root dry weight and root to shoot ratio, indicating that the effect of enhanced ammonium nutrition on the dry matter weight was positive mainly in the aboveground, while the effect in the underground was not obvious. Moreover, the ratio of root to shoot was the highest, the main root length was the longest when supplied with only NH₄⁺, indicating that single NH₄⁺ could promote root growth. All results showed that ammonium and nitrate nitrogen had different mechanisms for regulating growth and development in cotton underground and aboveground organs, mixed nitrogen was more favorable to the aboveground growth of cotton seedlings.