文献类型：期刊文献

中文题名：阿维链霉菌基因表达谱芯片的初步研究

英文题名：Study on microarray gene expression profiling of Streptomyces avermitilis

作者：许迎霞[1];叶邦策[1]

机构：[1]华东理工大学生物反应器工程国家重点实验室,上海200237

年份：2009

卷号：26

期号：2

起止页码：4

中文期刊名：生物学杂志

外文期刊名：Journal of Biology

收录：CSTPCD;;CSCD:【CSCD_E2011_2012】；

基金：国家863项目(2007AA02Z331);自然科学基金(20776039);新世纪优秀人才支持计划(NCET-07-0287)

语种：中文

中文关键词：基因芯片;探针;杂交;信号

外文关键词：gene chip ; probe ; hybridization; signal

摘要：用表达谱芯片分析阿维链霉菌在发酵过程中转录组表达情况,优化微生物转录组分析条件。提取RNA,进行荧光探针的合成,杂交,扫描对数据进行分析。RNA的提取要注意破壁方法的选择以及菌体量;合成cDNA时,RNA的加入量一般在10μg左右;当cDNA用逆转录的方法得到时,用专门的纯化试剂盒要比传统的异丙醇纯化效果好;杂交温度对于合成基因来说一般在42℃,对于比较长的PCR产物来说,温度相比较要高点;对于杂交时间来说,杂交12h之上,效果要明显比6h的好,所以杂交时间一般选择12h之上。对芯片上的197个合成基因进行分析:大约有60个基因进行了表达,其中包括18个双组分或激酶基因;10个糖酵解或者三羧酸循环途径基因;3个阿维基因簇;7个调节基因;5个细胞分化或孢子形;4个运输或固定基因;两个伴侣分子;5个RNA聚合酶σ因子;3个转录基因;两个脂肪酸合成基因;1个RNA聚合酶β亚基。
Transcript profiling of Streptomyces avermitilis during fermentation using microarray was studied by extracting RNA, synthesizing fluorescent probe and hybridization result. Result showed that when cDNA synthesized, the quantity of RNA was about 10μg. When cDNA synthesized by RT reaction, the result of purification by using expert reagent was better than by isopropanol. The temperature of hybridization was 42℃ for compound gene, but for PCR product, it will become higher. The results were better after hybridized 12h. Among analyzed 197 synthetic genes on chip, approximately 60 genes have been expressed, including 18 two-component/kinase, 10 glycolysis/TCA, 3 gene clusters, 7 regulators, 5 asaption/celldivision/differentiation/sporulation, 4 transport/binding proteins, 2 chaperones, 5 RNA polymerase sigma-factors, 3 transcription/translations, 2 fatty acid synthesises and 1 RNA polymerase β subunit.