Cloning of polyadenylated mRNA fragments of Escherichia coli with restriction display-polymerase cha

• 作 者: 胡子有 马文丽 宋艳斌 张宝 吴清华 郭秋野 彭翼飞 郑文岭
• 机 构: Institute of Molecular Biology, First Military Medical University, Guangzhou 510515, China,Institute
• 出 处: Journal of Medical Colleges of PLA
• 发表日期: 2002年04期
• 关键词: oligo RNA cloned ligation digestion sequenced Cloning prior primer enriched

Objective: To investigate the polyadenylation of mRNA in E. coli. Methods: The mRNA of E. coli was enriched from the total RNA with oligo(dT)-cellulose, prior to reverse transcription using oligo(dT)18 as the primer. Double-stranded cDNA was subsequently synthesized, which was subjected to digestion with Sau3A. I to produce multiple gene fragments for ligation with the adapters. PCR was carried out in 10 groups according to 10 different pairs of the selective primers, and the PCR products were then cloned into T-vectors. Results: More than 100 gene fragments had been cloned, 30 of which were sequenced. Conclusion: Polyadenylation of E. coli mRNA may not be a biochemical curiosity but a general attribute of bacterial mRNA.

Objective: To investigate the polyadenylation of mRNA in E. coli. Methods: The mRNA of E. coli was enriched from the total RNA with oligo (dT) -cellulose, prior to reverse transcription using oligo (dT) 18 as the primer. Double -stranded cDNA was succeeded, which was subjected to digestion with Sau3A. I to produce multiple gene fragments for ligation with the adapters. PCR was carried out in 10 groups according to 10 different pairs of the selective primers, and the PCR products were then cloned into T-vectors. Results: More than 100 gene fragments had been cloned, 30 of which were sequenced. Conclusion: Polyadenylation of E. coli mRNA may not be biochemical curiosity but a general attribute of bacterial mRNA.