田间灰飞虱抗药性相关细胞色素P450基因和酯酶基因的筛选  

Screening of cytochrome P450 and esterase genes related to insecticide resistance in field population of Laodelphax striatellus ( Homoptera: Delphacidae)

作　　者：张林林[1] 徐鹿[1] 韩召军[1] 

机构地区：[1]南京农业大学植物保护学院/农业部作物病虫害监测与防控重点开放实验室,江苏南京210095

出　　处：《南京农业大学学报》2010年第5期29-34,共6页Journal of Nanjing Agricultural University

基　　金：国家科技支撑计划项目(2006BAD02A16)

摘　　要：利用毒死蜱、吡虫啉和溴氰菊酯等3类不同药剂,通过毒力生测首先确定了田间灰飞虱对杀虫剂的低水平多重抗性特性;其次通过解毒酶活力测定,发现抗性品系的细胞色素P450氧化酶和酯酶明显升高;进而根据灰飞虱转录组数据进行引物设计,通过PCR克隆对转录组中的细胞色素P450及酯酶基因进行验证,在得到53条细胞色素P450基因序列及72条酯酶基因序列的基础上,利用半定量PCR比较这些基因在灰飞虱田间抗性品系和敏感品系间的表达差异。结果发现:细胞色素P450的CYP6家族中有10条,CYP4家族中有6条,这16条基因序列在抗性品系中的表达量明显高于敏感品系;酯酶中也有12条基因系列在抗性品系中的表达量明显高于敏感品系,它们中3条来自羧酸酯酶家族,8条来自磷酸酯酶家族,1条来自其他酯酶家族。提示:解毒酶基因的过量表达是灰飞虱田间低水平多重抗性的主要机制,也是抗药性发展初期的关键因素,利用分子生物学技术进行田间监测,有利于了解田间灰飞虱的抗药性发展。Low level of multiple insecticide resistance in a field resistant strain of Laodelphax striatellus was first identified by bioassay with chlorpyrifos,imidacloprid and deltamethrin. The enhancement of cytochrome P450 oxidase and esterase activity in this strain was also confirmed by analysis of detoxification enzymes. Then,the genes of cytochrome P450 and esterase were surveyed by PCR cloning with the primers design on the transcriptome data of this pest,and totally 53 of cytochrome P450 genes and 72 of esterase genes were obtained. Finally,semi-quantitative PCR was employed to compare the difference in expression of these genes between susceptible and resistant hoppers. The results showed that 16 cytochrome P450 genes including 10 from CYP6 and 6 from CYP4 were expressed significantly more highly in the resistant strain. So did 12 of esterase genes,of which 3 come from carboxylesterase,8 from phospho-esterase and 1 from other esterases. The discussion came to the conclusion that excessive expression of detoxification enzymes identified above was the main mechanism of the low level multiple resistances in the field population of L. striatellus and the key factors during the initiation of insecticide resistance development. Monitoring of the expression of these genes in field populations should be significant to knowing the development of the insecticide resistance of L. striatellus.

关 键 词：灰飞虱 抗药性 细胞色素P450 酯酶 半定量PCR 

分 类 号：Q965.9[生物学—昆虫学]