大豆再生抑制消减杂交文库的构建  

Construction of SSH library of regeneration related gene in soybean

作　　者：武小霞[1] 孙晶[1] 苏安玉[2] 李静[1] 刘明[1] 张彬彬[1] 李冬梅[1] 李文滨[1] 

机构地区：[1]东北农业大学农学院大豆生物学教育部重点实验室,哈尔滨150030 [2]东北农业大学资源与环境学院,哈尔滨150030

出　　处：《东北农业大学学报》2014年第7期38-44,共7页Journal of Northeast Agricultural University

基　　金：国家自然科学基金资助项目(31071438);黑龙江省自然科学基金项目(ZD201117);黑龙江省教育厅重点项目(12531z001)

摘　　要：大豆再生受基因型限制，高效稳定再生体系建立难、遗传转化效率低，影响生物技术的应用。为获得与大豆再生相关基因，解析大豆再生基因分子机制，从而建立高效、稳定再生体系，为生物技术在大豆分子育种中的应用奠定基础。文章利用抑制性消减杂交技术（Suppression subtractive hybridization，SSH），以东农50子叶节为材料构建消减杂交cDNA文库，从文库中随机挑取917个阳性克隆，PCR鉴定插入片段大部分集中在100~750 bp，利用BLAST在GenBank数据库进行序列相似性比对，经过同源性比对归并后得到411个高质量ESTs片段，功能分析表明，这些基因与信号转导、葡萄糖、蛋白质大分子生物合成代谢，光、叶形态发生相关；调控细胞凋亡、细胞自身防御、细胞壁分化；参与各种激素及细胞分裂素介导的信号通路。It is the world recognized chal enge to set up genetic transformation of soybean regeneration system with the problem of low efficiency. In order to establish a highly efficient and stable regeneration system, in this study, cotyledonary node of DN50 were used to construct suppression subtractive cDNA library, and 917 positive clones were screened. The length of most of the insert fragments was focused on 100-750 bp. For further ESTs screening, 411 differential y expressed functional ESTs were found and predicted by BLAST （Basic Local Alignment Search Tool） in GenBank. The ESTs were annotated by similarity to orthologs and paralogs detected with Blastn and Blastx. Function analysis showed that genes were annotated by similarity to signal transduction, macromolecules glucose, and protein synthesis and metabolism, light and leaf morphogenesis, regulation of apoptosis, the celldefense, and cellwal differentiation, participate in a variety of hormones and cytokinin-mediated signaling pathways.

关 键 词：大豆 抑制消减杂交(SSH) CDNA文库 再生 

分 类 号：S565.1[农业科学—作物学]