The aim was to study whether xenia effect existed in Chinese cabbage.Taking 14S116 as female parent and 14S443,14S375,14S120,14S502,14S393,14S536 as male parent,and taking 14S193 as female parent and 14S443,14S125,14S116,14S375,14S531 as male parent,and taking 92S24A as female parent and 8407,72M,DaT511,Da164-2-1,14S126,14S375,Shandong number 4,Jiecai,Da qin 3 as male parent,hybrids were prepared,and silique length,silique width,beak length,shape of seed,seed number of per pod,1000-grain weight,from which mid-parent,heterosis and male effect were acquired.Another,JY3 and JY4,JY5 and JY6,JY19 and JY20,Bre and Xiasheng,92S24 and Xiasheng,14S375 and 14S116 were respectively intercrossed,1000-grain weight seed were determined,and shope of seed and color of seed from some hybrids were observed,from which mid-parent,heterosis and male effect were acquired.The results showed that length and width of the silique,beak length,shape of seed,seed number of per pod,1000-grain weight of F₀ affected by pollen from different male parent were detected.Over-parent heterosis and heterosis were presented.

To explore the root disease resistance genetic regularity,by root disease resistance difference material,G57 and G70,the F₂ segregation population containing 500 per plant was built.Through artificial inoculation of the parents,F₁ and F₂ segregation population phenotypic characterization results were statistically analyzed. The results showed that root disease resistance of materials was controlled by a single dominant gene. In order to further localization of resistance genes in test materials,we used 678 molecular markers of parents,F₁ and F₂ population screening verification,resistance genes was preliminarily located in molecular marker KBRH129J18 and TCR02-F.This test was based on two chain tags,chain development design closer chain of molecular markers,ultimately with resistance genes chain 5 of SSR molecular markers,respectively,Bra0345-1,Bra0235-2,Bra0235-1,Bra19317,Bra019392. Their genetic distance with resistance genes were 2.4,2.4,2.4,2.5,3.3 cM.Validated,development and design of polymorphism markers in Chinese cabbage had good applicability.

To explore the reasons for difference in mechanism of cold resistance of winter rapeseed of Brassica rape and Brassica napus.The method of combining field and pot culture was adopted,and using 8 different cold resistance of winter rapeseed varieties as materials,before wintering period with 5 to 6 true leaves,field experiments researched botanical morphology and accumulation of dry matter,the physiological characteristics were measured by pot experiment after the treatment of 24℃→10℃→5℃→0℃→-5℃→-10℃ 48 h.The results showed that winter Brassica rape was creeping growth,growth point was under the ground,accumulation of dry matter was mainly concentrated in the underground part,such as fresh matter of underground parts and dry matter of underground parts of winter Brassica rape had increased,which average increased 236.1%,263.0% than winter Brassica napus respectively.It was showed that the photosynthetic organic product of strong cold resistant varieties was preferentially allocated to the underground part during the vegetative growth stage,to establish a large root system to provide metabolic energy for the safe winter.With the change of temperature,the physiological and biochemical indexes of different varieties was vary greatly,SOD activity of Longyou 7 increased 10.7% than CK at low temperature of -5℃,CAT and POD activity of Longyou 7 increased 24.7%,28.6% than CK respectively at 0℃,SP content of winter Brassica rape average increased 32.3% than winter Brassica napus at 0℃,SS content of winter Brassica rape average increased 71.4% than winter Brassica napus,and MDA content of winter Brassica napus average increased 52.8% than winter Brassica rape at low temperature of -10℃,this indicated that the strong cold resistant varieties could protect themselves from damage of low temperature condition,the CAT,POD,SP were protective substance of against chilling injury,but SOD,SS were protective substance of against freezing.Therefore,the winter Brassica rape had obvious advantages in the morphology and physiological level than winter Brassica napus,the morphological advantages was that to withstand cold weather of extreme,to provide metabolic energy of during the wintering period and regreen next year;At physiological levels,the activities of protective enzyme and content of osmotic regulation of winter Brassica rape increased significantly at low temperature stress to protect the structure of cell membrane,and the accumulation of MDA was decreased,to alleviate hurt of winter rapeseed leaves at low temperature,so the over wintering was ensured highly,it was important to provide a theoretical basis for the research on the cold resistance of winter rapeseed of Brassica rape and Brassica napus in the North of China.

Rapid and efficient DNA extraction is the key step in large-scale molecular breeding of crop. To construct a method for rapid extraction of genomic DNA from Chinese cabbage, the Chinese cabbage leaves were used as experimental materials, the quality of DNA extracted by the CTAB method, two-step CTAB method and four alkaline lysis methods was compared, the PCR amplification effects of DNA extracted by different methods were analyzed, and the preservation time and preservation conditions of genomic DNA extracted by different methods were compared, the optimal method was selected for the application of molecular marker-assisted selection against clubroot disease.The results showed that genomic DNA extracted by the CTAB and three kinds of alkaline lysis methods could be used as a template for PCR, and the PCR amplification products could be detected by 8% non-denaturing polyacrylamide gel electrophoresis to clear bands. Among them, the alkaline lysis method Ⅲ, not only the extraction quality was good and the extraction process was simple and fast, but also it could meet the needs of high-throughput DNA extraction of Chinese cabbage, the extracted DNA was stored at 4℃ and -20℃, and the genomic DNA stored for 30 days was used as a template for PCR amplification, the products was still detected by polyacrylamide gel electrophoresis to clear bands, it illustrates that this method has a long preservation time and it has been proved that this method has a good effect on the application of molecular marker-assisted selection against clubroot disease. Alkaline lysis method Ⅲ has significantly improved the efficiency of Chinese cabbage molecular marker-assisted selection, and can be widely used in Chinese cabbage molecular marker-assisted selection breeding.

The products obtained by long range PCR based on objective sequences were the effective probes for FISH to carry out molecular cytogenetic study.However,it is very difficult to amplify more than 5 kb-long fragments effectively by traditional PCR techniques.Suitable reaction conditions and reaction system are the necessary prerequisite for running effective LR-PCR amplification.In order to obtain LR-PCR products based on the objective sequences as FISH probes,80 pairs of LR-PCR primers were designed based on the sequences of repeat-free regions on the top of Chinese cabbage chromosome A03,and PCR technique system was optimized from the following aspects containing the template quality of Chinese cabbage genomic DNA,dNTPs concentration,annealing temperature and extended time.Results indicated that genomic DNA of seedling leaves and LA Taq polymerase enzyme could improve the amplification quality and efficiency of long-range PCR.And 5-15 kb PCR products could be amplified by the reaction system with 20 μL of total volume,comprising of 50 ng/μL DNA 2 μL,2.5 mmol/L dNTPs 1.6 μL,10 μmol/μL of each primer 1 μL,10×LA PCR Buffer Ⅱ (including Mg²⁺) 2 μL,5 U/μL LA Taq enzyme 0.2 μL.PCR reaction procedure was followed as:98℃ for 15 s;58-64℃ for 10 s,68℃ for 5 min,35 cycles;final extension at 68℃ for 10 min kept at 4℃.Under these conditions,60 fragments with 5-15 kb length were obtained from Chinese cabbage genome.This research would establish the basis for carrying out long range PCR-FISH on meiotic pachytene chromosomes of Chinese cabbage and for clarifying evolutionary by comparing chromosome painting among close related species.

In order to explore the mechanism of super cold resistant varieties of Winter turnip rape,select super cold resistant varieties of Winter turnip rape Longyou No.7 leaves as materials,using two different extraction liquids(Extraction A:0.1 mol/L Tris-HCl (pH=7.6),0.2 mol/L KCl,1 mmol/L PMSF,Extraction B:0.05 mol/L Tris-HCl (pH=7.6),0.01 mol/L EDTA-Na₂,0.02 mol/L Vc,1 mmol/L PMSF) to compare the extracted effect.The experiment showed that the extraction A liquid protein yield reached (0.073±0.004 6) mg/g,and the extraction ratio increased by 42.88% compared to liquid B.Through the Glucose-6-phosphate dehydrogenize (G₆PDH) activity detection and the extraction A was lower than B 0.91 percent;not only the extraction A extract protein was shorter than B 4.5 hours in the first oelectric focusing electrophoresis of two-dimensional electrophoresis,but also was get clear gel map;through the calculation of abundance,there were total 339 differentially expressed proteins discovered in Longyou No.7 under low temperature stress(4℃,48 h)later,changes in expression levels of more than two times the protein spots of 46,which 18 up-regulated protein expression spots,21 down-regulated protein expression spots,7 specific protein expression spots,these differential protein spots were suggested to be related to low temperature stress.This laid the foundation for the research of cold resistant varieties in Winter turnip rape Longyou No.7 proteomics aspect.

In order to analyze the gene expression more accurately in Chinese cabbage by quantitative Real-time PCR,Chinese cabbage translocation lines added No.4 chromosome fragments from cabbage under Chinese cabbage background were used as materials in this experiment. The leaves of Chinese cabbage-cabbage translocation lines in different growth stages,including seedling stage,rosette stage,folding stage and heading stage,floral buds of different sizes and leaves of folding stage treated with auxin(Indole-3-acetic acid,IAA)and auxin inhibitor(2,3,5-triiodobenzoic acid,TIBA)were selected to evaluate the expressions stability of genes. Expression stability of twenty one candidate reference genes were detected by qRT-PCR,including Apr, BcTIP41, U34559, EF1α, TUB4, CYP, DNAJ, HIS, TUA5, UKN1, SKIP16, CAC, ACTIN, ACTIN-1, ACTIN-2, GAPDH, UBC30, UBQ, PPR, PP2A, MDH. geNorm and NormFinder were used to analyze above these results,which showed that the qRT-PCR most suitable reference genes of Chinese cabbage-cabbage translocation lines had some differences in different development periods,different tissue of the material,and under condition of different hormone treatment. The UKN1 and TUB4 genes were most stable expression in vegetative growth stage(from seedling to heading)of Chinese cabbage-cabbage translocation lines. In folding stage of Chinese cabbage-cabbage translocation lines,the most stable expression reference genes were BcTIP41 and ACTIN after treatment with auxin IAA,and the most stable expression reference genes were UKN1 and BcTIP41 after treatment with auxin inhibitor TIBA. DNAJ, ACTIN and PP2A genes showed the most stable expression in the floral buds with six size levels of Chinese cabbage-cabbage translocation lines. These results will lay the foundation for the accurate analysis of the gene expression of Chinese cabbage-cabbage translocation lines,and further provided a reference for the selection of the reference gene in other plants of Brassica at different developmental stages and hormone treatments.

For the disease-resistant breeding of Qingmaye Chinese cabbage,five kinds of inoculation methods(Indigenous bacteria,Seal of indigenous bacteria,Dip method,Injection method,Seed soaking method),different soil pH,different inoculation concentration,illumination and root secretion were compared by use Qiulü 60 as material in this article.The results showed that the effect of soil inoculation was more excellent and stable.In contrast,seed-immersion method was the poorest.The acid soil pH was fit to clubroot break out and the disease index reached the maximum when pH value was 6.5.1 g soil inoculated 0.050 g root was the most suitable inoculation concentration.After light treatment,the disease index was increased,it showed that light had a promoting effect on diseases.Root secretion of tomato,resistant and susceptible hosts could promote the happening of disease.Therefore,in the artificial inoculation test of Qingmaye Chinese cabbage can be the above optimum inoculation method and conditions so as to improve the effect of inoculation.

In order to expand the new purple material resources and breed our own purple Chinese cabbage varieties in China,we focused on a new purple germplasm 15NG28,its F₁ back-crossing F₁ and self-bred progenies to study characteristics,distribution of anthocyanins in leaf,ploidy modes in genome,chromosome patterns and differentially expressed genes of these progenies.Results showed F₁ hybrids crossed by 15NG28 and Chinese cabbage could be obtained,which segregated purple and green colors.At the seedling and adult stages,15NG28 and purple F₁ plants had purple colors in adaxial leaf,abaxial leaf,vein,while the F₁ plants owned purple colors in adaxial,abaxial blade of leaf and 15NG28 had purple color in adaxial blade,green color in abaxial blade.Meanwhile purple F₁ hybrids had different ploidy modes,i.e.,3n,4n,6n,etc.and chromosome patterns,i.e.,24,28,36,etc.Among them,16M-170-21 was similar as B.juncea.16M-170-21 self-bred seeds could be gotten,which showed distinctly different characteristics.While 16M-170-22 was similar as Chinese cabbage.Characteristic of 31 BC₁F₁ progenies of 16M-170-22 were the same as Chinese cabbages,and purple to green ratio was 14:17.Especially self-bred seeds of 14 purple lines were better.RNA-seq results showed twelve genes involved in flavonoid,flavone,flavonol and anthocyanin biosynthesis pathway were apparently up-regulated in purple lines.Interestingly Bra013652 (as LDOX gene) and Bra027457 (as DFR gene) were structural genes,which were up-regulated apparently in purple lines compared to green lines.Especially the two genes were differently from c3563g1i2,a regulator factor R2R3-MYB reported before.Therefore 15NG28 and its progenies could be new germplasm resources and 15NG28 might origin from B.juncea.

In order to identify the clubroot resistance gene of Chinese cabbage in G6 and develop linkage markers, high-generation inbred line G4 of Chinese cabbage with high-susceptible to clubroot disease, high-generation inbred line G6 of Chinese cabbage with high-resistant to clubroot disease, F₁ obtained by crossing G4 and G6, and the F₂ generation separation populations constructed by F₁ selfing were used as materials. By artificial inoculation, phenotypic identification and genetic analysis, it was found that clubroot disease resistance in the disease resistance materials were controlled by a dominant single gene. Further expanding the number of F₂ generation populations to map the genes of clubroot disease and screening molecular markers linked to disease resistance genes, the linkage analysis of polymorphic markers was performed using JoinMap 4.0 software, and five InDel markers linked to the resistance gene of Chinese cabbage were obtained. The most recent markers on both sides were BrID10727 and BrID10867, the genetic distance from the disease resistance genes were 4.6,2.5 cM, respectively, and the disease resistance gene was located on the chromosome A08 of Chinese cabbage. In addition, it was found that the newly developed polymorphic marker BrID10381 based on the Crr1 gene sequence was located outside the initial localization range of the newly located clubroot resistance gene, so it could be inferred that the newly located clubroot resistance gene locus was not the same locus as Crr1, and the marker BrID10381 could be used for molecular marker assisted selection of Crr1 gene.

To clarify the resistant relationship between physiological races and Chinese cabbage(Brassica rapa L.ssp. pekinensis) variety, physiological races 4, 7, 10 and 11 were inoculated by injecting. The disease resistance of the 25 Chinese cabbage and CR markers were detected. The results showed that 8 cultivars such as Shandiwang 2 were resistance to only 4 physiological races; 7 cultivars such as Degao CR117 were susceptible to only 4 physiological races; 4 cultivars (including C1) were susceptible to 2 physiological races, and 6 cultivars (including Huakang 301) were only susceptible to 4 physiological race. All materials contained Crr1, CRk was detected in samples except the Degao CR Tiejia 1 and the Dadi CR118, the disease resistance locus CRa and CRb were detected in 21 and 16 samples respectively, 11 cultivars (including Degao CR117) contained Crr2 resistance locus, Crr3 resistance locus was detected only in Jingchun CR3, CRc resistance locus was contained in Xingguan and CR75.In short,different Chinese cabbage materials have different resistance to the same physiological race,and the same material responds differently to different physiological races. The physiological race No.4 was the most serious, followed by the physiological race No.7. The results of molecular marker identification showed that the resistance sites of Chinese cabbage materials were not necessarily resistant to disease. The resistance of Crr1 had little effect on material resistance.

Different tipburn resistance materials of Chinese cabbage were used as materials to study the inheritance of resistance gene to tipburn and to conduct QTL mapping so as to provide a theoretical basis for the molecular assistant selection (MAS) breeding and resistance mechanism. This research adopted significantly different resistance tipburn line high inbred Qingmaye type Black 227 (resistant lines) and high inbred Baotou type B120 (susceptible lines) as materials to obtain DH group by microspore culture F₁ generation. We classified degrees of the disease and combined the molecular genetic map of Chinese cabbage. According to disease severity in the field of DH group, QTL analysis on tipnurn resistance genes were conducted by MapQTL 5.0 software. Results showed that the inheritance of the resistance gene to Chinese cabbage tipburn in tested materials fit to the inheritance law of quantitative trait.This study detected two InDel markers BrID10343 and BrID10349 which link closelyed with tipburn resistance gene of Chinese cabbage. Two markers were on the Chr.7,in which the genetic distance of 1.031 cM and genetic contribution rates were above 40%.InDel markers BrID10343 and BrID10349 linked closelyed with tipburn resistance gene of Chinese cabbage. The results would be of great benefit to fine mapping for the major QTL of tipburn resistance gene, also the results will lay a good foundation for Chinese cabbage MAS resistance breeding.