Zhang Zhongyi, Fan Huamin, Yang Yanhui, Li Mingjie, Li Juan, Xu Haixia, Chen Junying, Chen Xinjian
Centre of Engineering Research of Chinese Medicinal Materials, Henan Agriculture University, Zhengzhou 450002, China.
Zhongguo Zhong Yao Za Zhi. 2011 Feb;36(3):276-80.
To explore the molecular mechanism of continuous monoculture problem by constructing the cDNA libraries of continuous monoculture Rehmannia glutinosa.
To use the suppression subtractive hybridization (SSH) technique to construct the forward and reverse subtractive cDNA libraries of continuous monoculture R. glutinosa to adopt blue-white colony screening and PCR to detect the positive clones which would be sequenced and analyzed by bioinformatics.
The subtracted cDNA libraries of continuous monoculture R. glutinosa. were successfully constructed, and the result showed that the forward and reverse subtracted libraries obtained 300 positive clones, respectively. The forward and reverse libraries got different ESTs, and produced 232 (forward library) and 214 (reverse library) unique ESTs by sequencing. Based on homology search of BLASTX and BLASTN in NCBI, 200 and 195 of unique ESTs were homologous to known genes in the forward and reverse libraries, respectively. Categories of orthologous group (COG) showed that the forward and reverse libraries got 60 and 61 ESTs with the corresponding gene annotation, involving 21 metabolic pathways.
The information of differential expression genes in continuous monoculture R. glutinosa, and their functional annotation of differentially expressed genes indicate that continuous monoculture has a profound effect on expression of the genes in R. glutinosa. Furthermore, the research analyzed several key genes in response to replant problem, which provided a foundation for revealing the molecular mechanism of continuous monoculture R. glutinosa.
通过构建地黄连作的cDNA文库，探索地黄连作障碍的分子机制。
采用抑制性消减杂交（SSH）技术构建地黄连作的正向和反向消减cDNA文库，通过蓝白斑筛选和PCR检测阳性克隆，对阳性克隆进行测序并利用生物信息学进行分析。
成功构建了地黄连作的消减cDNA文库，结果显示正向和反向消减文库分别获得300个阳性克隆。正向和反向文库得到不同的ESTs，测序后分别产生232个（正向文库）和214个（反向文库）独特ESTs。基于NCBI中BLASTX和BLASTN的同源性搜索，正向和反向文库中分别有200个和195个独特ESTs与已知基因同源。直系同源簇（COG）分类显示，正向和反向文库分别有60个和61个ESTs具有相应的基因注释，涉及21条代谢途径。
地黄连作中差异表达基因的信息及其差异表达基因的功能注释表明，连作对地黄基因的表达有深远影响。此外，该研究分析了几个应对连作问题的关键基因，为揭示地黄连作障碍的分子机制提供了基础。
