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利用cDNA-AFLP方法鉴定寄生线虫捻转血矛线虫中的左旋咪唑抗性标记。

Identification of levamisole resistance markers in the parasitic nematode Haemonchus contortus using a cDNA-AFLP approach.

作者信息

Neveu C, Charvet C, Fauvin A, Cortet J, Castagnone-Sereno P, Cabaret J

机构信息

INRA, IASP, 213, UR 1282, F-37380 Nouzilly, France.

出版信息

Parasitology. 2007;134(Pt 8):1105-10. doi: 10.1017/S0031182007000030.

Abstract

cDNA-AFLP (cDNA-Amplified Fragment Length Polymorphism)-based strategy has been used to identify levamisole (LEV) resistance markers in the nematode Haemonchus contortus. Transcript profiles of adult nematodes from two LEV-resistant and two susceptible isolates were compared. Among the 17 280 transcript-derived fragments (TDFs) amplified, 26 presented a polymorphic pattern between resistant and susceptible nematodes: 11 TDFs were present in both resistant isolates and absent from both susceptible isolates whereas 15 TDFs were present in both susceptible isolates and absent from both resistant isolates. 8 TDFs specifically present in resistant isolates were cloned and sequenced. Some of these TDFs could represent novel genes, as their sequences presented no homologies in databases. Interestingly, specific expression of one candidate (HA17) in resistant nematodes from different isolates was confirmed by RT-PCR experiments. The finding that HA17 expression correlates with LEV resistance in three H. contortus isolates vs five susceptible isolates strongly suggest that we identified a new potential marker of LEV resistance. This differential approach at the transcriptome level could be of great interest for the identification of the molecular mechanism involved in this phenotype.

摘要

基于cDNA-AFLP(cDNA扩增片段长度多态性)的策略已被用于鉴定捻转血矛线虫中左旋咪唑(LEV)抗性标记。比较了来自两个左旋咪唑抗性分离株和两个敏感分离株的成年线虫的转录谱。在扩增的17280个转录衍生片段(TDF)中,有26个在抗性和敏感线虫之间呈现多态性模式:11个TDF在两个抗性分离株中均存在,而在两个敏感分离株中均不存在,而15个TDF在两个敏感分离株中均存在,而在两个抗性分离株中均不存在。对8个特异性存在于抗性分离株中的TDF进行了克隆和测序。其中一些TDF可能代表新基因,因为它们的序列在数据库中没有同源性。有趣的是,通过RT-PCR实验证实了一个候选基因(HA17)在不同分离株的抗性线虫中的特异性表达。HA17表达与三个捻转血矛线虫分离株对五个敏感分离株的左旋咪唑抗性相关,这一发现强烈表明我们鉴定出了一个新的潜在左旋咪唑抗性标记。这种转录组水平的差异方法对于鉴定参与该表型的分子机制可能非常有意义。

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