摘要
以玉米骨干自交系478为试验材料,从各生长时期的叶片中提取总RNA后,再利用磁珠法从中分离纯化出mRNA,然后以mRNA为模板,反转录合成cDNA第一链,再在DNA聚合酶作用下,通过长距离PCR,扩增双链cDNA(ds cDNA)。利用SMART技术,通过同源重组的方法,在酵母菌株AH109中构建了玉米叶片全长cDNA文库。经检测,文库的转化率为1.54×107转化子/3μg pGADT7-Rec,文库滴度为1.03×106pfu/mL,重组率为95%。
The maize inbred line 478 was used experiment material,and the total RNA was isolated from each periods of growth maize leaf and mRNA was separated and purified using the SA-PMPs method.Afterwords,mRNA was used as cyclostyle and synthesized the primacy cDNA.Under DNA polymerase function,DNA was expand double strand DNA by using LD-PCR approach.Using SMART technique and homologous reorganization method,the yeast two hybrid cDNA library was constructed in yeast AH109,After examination,the library conversion rate was 1.54×107 transfers beggar /3 μg pGADT7-Rec,the capacity of the library was 1.03×106 pfu/mL,and the reorganization rate was 95%.
出处
《西北农业学报》
CAS
CSCD
北大核心
2008年第5期162-165,共4页
Acta Agriculturae Boreali-occidentalis Sinica
基金
国家863计划项目(SQ2007AA10Z135024)
四川省教育厅青年基金项目(2006B010)资助
关键词
玉米叶片
酵母双杂交文库
构建
Maize leaf,Yeast two hybrid cDNA library,Construction
