利用抑制消减杂交技术筛选猪蛔虫性别差异表达基因

 为了筛选线虫的性别特异性基因，为线虫乃至寄生虫的性别控制提供新的工具，本研究选择猪蛔虫（Ascaris suum）为模型，分别提取雌性和雄性成虫的mRNA后，采用Clontech公司的PCR-selectTM试剂盒进行反转录合成cDNA并进行抑制消减杂交（SSH），构建猪蛔虫雌、雄虫性别差异表达的消减cDNA文库，并采用地高辛（DIG）标记的cDNA探针进行Southern 杂交检验所构建文库的消减效率。结果表明，雌、雄虫性别差异表达的消减cDNA文库均具有很强的性别特异性。随机从各库中各抽取25个克隆进行测序及在线BLAST分析，发现在25个雄虫ESTs中有20个已知ESTs，5个新的ESTs；25个雌虫ESTs中有11个已知ESTs，还有8个可能为新基因。猪蛔虫性别差异表达的消减cDNA文库的成功构建，为进一步研究性别特异基因的功能奠定了基础。

Profiling of Gender-Specific Gene Expression for Ascaris suum by Suppression Subtractive Hybridization

Attempting a new method of gender-controlling nematode infection in animal production, differentially expressed male and female cDNA libraries of the adult Ascaris suum were constructed using the technique of suppression subtractive hybridization (SSH). The gender specificity of the constructed archives was determined using Southern blots with Digoxigenin(DIG)- labeled male and female cDNA probes. Twenty-five clones from each archive were selected ad libitum for sequence determination and homology analysis by BLAST online. Twenty of the 25 ESTs from the male cDNA library have a homology to the genes from C. elegans and 11 of the 19 ESTs from the female cDNA library have a homology to the genes from C. elegans. The successful construction of differentially expressed male and female cDNA libraries have provided a foundation for further study of the gender-specific genes.