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| 几个提高抑制消减杂交(SSH)文库质量的技术改进 | |
| 引用本文： | 董登峰,李杨瑞,白云彩.几个提高抑制消减杂交(SSH)文库质量的技术改进[J].西南农业学报,2011,24(5). |
| 作者姓名： | 董登峰 李杨瑞 白云彩 |
| 作者单位： | 1. 广西大学农学院,广西南宁530004;广西作物遗传改良生物技术重点实验室,广西南宁530007 2. 广西作物遗传改良生物技术重点实验室,广西南宁,530007 3. 广西大学农学院,广西南宁,530004 |
| 基金项目： | 国家自然科学基金项目，广西大学科研基金项目 |
| 摘 要： | 抑制消减杂交( SSH)高灵敏、易操作、高效富集低表达丰度基因和均一化目的基因片断,广泛用于差异表达基因的分离和cDNA文库构建.在实验操作中常出现低表达mRNA和cDNA丢失、大片断连接效率低等问题,构建的文库质量差,文章通过改进萃取操作提高目的产物(mRNA和cDNA)回收率,回收率增加30％以上；通过采用PVPP、高浓度KAc专一性结合和LiC1沉淀等措施除去RNA中的多酚和多糖、进而增加可分离mRNA种类和含量,对应的Uni-EST种类增加近3倍,通过采用胶回收并分段连接保护大片断cDNA等措施使文库中转录本丰富度增加,大片段(500 bp以上)比例增大,cDNA平均长度增大,文库质量明显提高. |
| 关 键 词： | 抑制消减杂交(SSH) 文库构建 技术改进 |
Improvement of Some Techniques for Enhancing Quality of Suppression Subtractive Hybridization cDNA Library |
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| DONG Deng-feng,LI Yang-rui,BAI Yun-cai.Improvement of Some Techniques for Enhancing Quality of Suppression Subtractive Hybridization cDNA Library[J].Southwest China Journal of Agricultural Sciences,2011,24(5). | |
| Authors: | DONG Deng-feng LI Yang-rui BAI Yun-cai |
| Institution: | DONG Deng-feng1,2,LI Yang-rui2,BAI Yun-cai1 (1.College of Agriculture,Guangxi University,Guangxi Nanning 530004,China,2.Key-Lab of Crop Genetic Improvement and Biotechnology,Guangxi Nanning 530007,China) |
| Abstract: | Suppression subtractive hybridization was widely used to isolate differentially expressed genes and construct cDNA libraries at present because of its high sensitivity,easy manipulation,high efficiency in enriching rare expressed genes and normalization of targeted fragments.In the process of experiment,the regular occurrences of loss of rare expressed genes(mRNAs and their corresponding cDNAs) and low efficiency in ligation between large cDNA fragments and plasmids usually resulted in bad library.In the pr... |
| Keywords: | Suppression subtractive hybridization Library Improvement of techniques |
| 本文献已被 CNKI 万方数据 等数据库收录！ | |