Comparative Transcriptome Analysis of Resistant and Sensitive Strains to Imidacloprid of Aphis gossypii(Hemiptera:Aphisidae)

doi:10.7668/hbnxb.20195087

Abstract

Abstract:

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.

Key words: Aphis gossypii, Imidacloprid, Insecticide resistance, Insecticide sensitive, Transcriptome, Differentially expressed gene

WANG Xinxia, ZHANG Huibin, LIU Yunxiang, LIU Siyu, LAI Youpeng, LI Qiurong. Comparative Transcriptome Analysis of Resistant and Sensitive Strains to Imidacloprid of Aphis gossypii(Hemiptera:Aphisidae)[J]. Acta Agriculturae Boreali-Sinica, 2025, 40(1): 177-188. doi: 10.7668/hbnxb.20195087.

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References 48

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基因名称 Gene name	引物名称 Primer name	序列(5'—3') Sequence(5'—3')	退火温度/℃ Annealing temperature
18S rRNA	上游引物	ATTGACGGAAGGGCACC	57
	下游引物	CGCTCCACCAACTAAGAACG
CYP6a2	上游引物	ACCTTGGAGGAACCGATACAC	58
	下游引物	TAAGATGTTGACAGACGGATGAGT
CYP6a13	上游引物	CTTTTGGAATGTTTGCTGCTG	58
	下游引物	GGAGGATACTTTCGGAGTGTTT
AChE2	上游引物	TCGGTTTCACTACACTTGCTATCT	58
	下游引物	TGCTTTTGCTAGTTTAAGTCCTCT
CarE	上游引物	AGGCATACCCTACGCTCAAC	57
	下游引物	TTCTGCTGTGGCACCGATA
ALP3	上游引物	GAATCGGAAGTGGGAATGC	57
	下游引物	TGCTTGTCGTCTATCCACTCG
CYP6k1	上游引物	AGAACTCGGACATTGAAGAT	61
	下游引物	TGAACCAGCAGTGAACAT
CYP6j1	上游引物	ACCCAACTCAGATTTCACCCT	58
	下游引物	TTCCGAACCGCATACCAA
CYP4c1	上游引物	TAAACATTCAGCAGGTGGGC	59
	下游引物	TGCGGTTCCATTTGTAGTGAC

基因名称 Gene name	引物名称 Primer name	序列(5'—3') Sequence(5'—3')	退火温度/℃ Annealing temperature
18S rRNA	上游引物	ATTGACGGAAGGGCACC	57
	下游引物	CGCTCCACCAACTAAGAACG
CYP6a2	上游引物	ACCTTGGAGGAACCGATACAC	58
	下游引物	TAAGATGTTGACAGACGGATGAGT
CYP6a13	上游引物	CTTTTGGAATGTTTGCTGCTG	58
	下游引物	GGAGGATACTTTCGGAGTGTTT
AChE2	上游引物	TCGGTTTCACTACACTTGCTATCT	58
	下游引物	TGCTTTTGCTAGTTTAAGTCCTCT
CarE	上游引物	AGGCATACCCTACGCTCAAC	57
	下游引物	TTCTGCTGTGGCACCGATA
ALP3	上游引物	GAATCGGAAGTGGGAATGC	57
	下游引物	TGCTTGTCGTCTATCCACTCG
CYP6k1	上游引物	AGAACTCGGACATTGAAGAT	61
	下游引物	TGAACCAGCAGTGAACAT
CYP6j1	上游引物	ACCCAACTCAGATTTCACCCT	58
	下游引物	TTCCGAACCGCATACCAA
CYP4c1	上游引物	TAAACATTCAGCAGGTGGGC	59
	下游引物	TGCGGTTCCATTTGTAGTGAC

品系名称 Name of strains	分子功能 Molecular function	生物学过程 Biological process	细胞组分 Cellular component
抗药性品系	GO:0042302角质层的结构成分	GO:0014075胺反应	GO:0000323裂解液泡
ApRS	GO:0004197半胱氨酸型内肽酶活性	GO:0071260细胞对机械刺激的反应	GO:0005764溶酶体
	GO:0030984激肽原结合	GO:0070670对白细胞介素-4的反应	GO:0016324顶端质膜
	GO:0043621蛋白质自结合	GO:0009612机械刺激反应	GO:0042383肌膜
	GO:0008234半胱氨酸型肽酶活性	GO:0045471乙醇的反应	GO:0005773液泡
	GO:0015020葡糖羧酸基转移酶	GO:0007519骨骼肌组织发育	GO:0009897质膜外侧
	GO:0042277肽结合	GO:0071214细胞对非生物刺激的反应	GO:0045177细胞顶端
	GO:0008194 UDP糖基转移酶活性	GO:0104004细胞对环境刺激的反应	GO:0042470黑素体
	GO:0005198结构分子活性	GO:0009749葡萄糖反应	GO:0048770色素粒
	GO:0033218酰胺结合	GO:0009611伤害反应	GO:0098552薄膜侧面
	GO:0070011肽酶活性,作用于L-氨基酸肽	GO:0009746对己糖的反应	GO:0044421胞外区部分
	GO:0008233肽酶活性	GO:0060538骨骼肌器官发育	GO:0005576胞外区
	GO:0016758转移酶活性,转移己糖基	GO:0034284单糖的反应	GO:0005615胞外区
	GO:0004175内肽酶活性	GO:0097305酒精反应	GO:0098590质膜区
	GO:0008239二肽基肽酶活性	GO:0009743碳水化合物反应	GO:0044425膜部分
	GO:0016757转移酶活性,转移糖基	GO:0009636对有毒物质的反应	GO:0005968蛋白-香叶基转移酶复合物
	GO:0004177氨肽酶活性	GO:0014706横纹肌组织发育	GO:0005739线粒体
	GO:0003968 RNA导向的5'—3'RNA聚合酶活性	GO:0060537肌肉组织发育	GO:0035060布拉复合体
	—	GO:0043434肽激素反应	GO:0044444细胞质部分
	—	GO:0034097对细胞因子的反应	GO:0031982囊泡
敏感性品系	GO:0003676核酸结合	GO:0090304核酸代谢过程	GO:0005634细胞核
ApSS	GO:0003968 RNA导向的5'—3'RNA聚合酶活性	GO:0039694病毒核糖核酸基因组复制	GO:0044428核部分
	—	GO:0006139含碱基化合物代谢过程	GO:0043229胞内细胞器
	—	GO:0046483杂环代谢过程	GO:0031981核腔
	—	GO:0006725芳香族化合物的细胞代谢过程	GO:0043226细胞器
	—	GO:0019079病毒基因组复制	GO:0044446胞内细胞器部分
	—	GO:1901360有机环状化合物代谢过程	GO:0043227膜结合细胞器
	—	GO:0006259DNA代谢过程	GO:0044422细胞器部分
	—	—	GO:0043231细胞内膜结合细胞器
	—	—	GO:0070013细胞器内腔
	—	—	GO:0031974膜封闭腔
	—	—	GO:0043233细胞器腔
	—	—	GO:0005730核仁
	—	—	GO:0043232细胞内无膜细胞器
	—	—	GO:0043228无膜细胞器
	—	—	GO:0015630微管细胞骨架
	—	—	GO:0044424胞外部分

品系名称 Name of strains	分子功能 Molecular function	生物学过程 Biological process	细胞组分 Cellular component
抗药性品系	GO:0042302角质层的结构成分	GO:0014075胺反应	GO:0000323裂解液泡
ApRS	GO:0004197半胱氨酸型内肽酶活性	GO:0071260细胞对机械刺激的反应	GO:0005764溶酶体
	GO:0030984激肽原结合	GO:0070670对白细胞介素-4的反应	GO:0016324顶端质膜
	GO:0043621蛋白质自结合	GO:0009612机械刺激反应	GO:0042383肌膜
	GO:0008234半胱氨酸型肽酶活性	GO:0045471乙醇的反应	GO:0005773液泡
	GO:0015020葡糖羧酸基转移酶	GO:0007519骨骼肌组织发育	GO:0009897质膜外侧
	GO:0042277肽结合	GO:0071214细胞对非生物刺激的反应	GO:0045177细胞顶端
	GO:0008194 UDP糖基转移酶活性	GO:0104004细胞对环境刺激的反应	GO:0042470黑素体
	GO:0005198结构分子活性	GO:0009749葡萄糖反应	GO:0048770色素粒
	GO:0033218酰胺结合	GO:0009611伤害反应	GO:0098552薄膜侧面
	GO:0070011肽酶活性,作用于L-氨基酸肽	GO:0009746对己糖的反应	GO:0044421胞外区部分
	GO:0008233肽酶活性	GO:0060538骨骼肌器官发育	GO:0005576胞外区
	GO:0016758转移酶活性,转移己糖基	GO:0034284单糖的反应	GO:0005615胞外区
	GO:0004175内肽酶活性	GO:0097305酒精反应	GO:0098590质膜区
	GO:0008239二肽基肽酶活性	GO:0009743碳水化合物反应	GO:0044425膜部分
	GO:0016757转移酶活性,转移糖基	GO:0009636对有毒物质的反应	GO:0005968蛋白-香叶基转移酶复合物
	GO:0004177氨肽酶活性	GO:0014706横纹肌组织发育	GO:0005739线粒体
	GO:0003968 RNA导向的5'—3'RNA聚合酶活性	GO:0060537肌肉组织发育	GO:0035060布拉复合体
	—	GO:0043434肽激素反应	GO:0044444细胞质部分
	—	GO:0034097对细胞因子的反应	GO:0031982囊泡
敏感性品系	GO:0003676核酸结合	GO:0090304核酸代谢过程	GO:0005634细胞核
ApSS	GO:0003968 RNA导向的5'—3'RNA聚合酶活性	GO:0039694病毒核糖核酸基因组复制	GO:0044428核部分
	—	GO:0006139含碱基化合物代谢过程	GO:0043229胞内细胞器
	—	GO:0046483杂环代谢过程	GO:0031981核腔
	—	GO:0006725芳香族化合物的细胞代谢过程	GO:0043226细胞器
	—	GO:0019079病毒基因组复制	GO:0044446胞内细胞器部分
	—	GO:1901360有机环状化合物代谢过程	GO:0043227膜结合细胞器
	—	GO:0006259DNA代谢过程	GO:0044422细胞器部分
	—	—	GO:0043231细胞内膜结合细胞器
	—	—	GO:0070013细胞器内腔
	—	—	GO:0031974膜封闭腔
	—	—	GO:0043233细胞器腔
	—	—	GO:0005730核仁
	—	—	GO:0043232细胞内无膜细胞器
	—	—	GO:0043228无膜细胞器
	—	—	GO:0015630微管细胞骨架
	—	—	GO:0044424胞外部分

表达量变化趋势(抗药性/敏感性) The tendency of the expression quantity (ApRS/ApSS)	KEGG序号 KEGG ID	KEGG 通路 KEGG pathway	P值 P value
上调	ko04210	细胞凋亡	3.59×10^-7
Up-regulation	ko04142	溶酶体	9.76×10^-5
	ko01220	芳香化合物降解	0.012 663
	ko02024	群体感应	0.015 806
	ko04145	吞噬体	0.017 341
	ko00930	己内酰胺降解	0.018 940
	ko00430	牛磺酸和亚牛磺酸代谢	0.025 182
	ko00591	亚麻油酸代谢	0.040 634
下调	ko03450	异源末端连接	0.001 476
Down-regulation	ko03008	真核生物核糖体生物合成	0.003 058
	ko03015	mRNA监控通路	0.033 431
	ko04111	酵母细胞周期	0.034 403

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