用PCR检测细胞培养中支原体污染

细胞培养中支原体污染已经成为严重的问题.为了扩增6种支原体(精氨酸支原体,口腔支原体,人型支原体,猪鼻支原体,发酵支原体及莱氏支原体)核糖体RNA操纵子的16s和23s DNA间区,设计了三个通用PCR引物(F1,F2及R1).当以6种支原体DNA为模板时,引物F1和R1产生340到468bp的片段,引物F2和R1产生145到211bp的片段,当用Hela细胞或E.coli DNA作为模板,用引物F1和R1时,在电泳中未观察到特定区带.此法最小能检出8.5fg精氨酸支原体DNA,相当于13个精氨酸支原体.这说明,当这些支原体污染细胞培养时,能用PCR法检测出来.

PCR Detection of Mycoplasma Contamination in Cell Culture

ycoplasma contamination of cell culture is aserious problem in biomedical reseach. Threecommon PCR primers(F1,F2 and R1) weredesigned to amplify the spacer region between16s and 23s DNA in rRNA operons of 6species of mycoplasmas (M.arginini,M.orale,M. hominis ,M .hyorhinis,M fementans and A.laidlawii). When the DNA of 6species was used as the template, primers F1and R1 produced fragments of 340 to 468 bp,and primers F2 and R1 produced fragments of145 to 211 bp. No discrete band was observedin electrophoretic gels when Hela cell or E.coli DNA was served as the template with theuse of primers F1 and R1. As little as 8.5fgDNA of M.arginini,approximately 1 3 organisms could be detected.This suggests thatmycoplasma contamination to cell cultures canbe detected by PCR.