摘要
逆转录酶 (RT)是目前发现的所有逆转录病毒的一个重要标志 ,凡是逆转录病毒均携带逆转录酶 ,逆转录酶活性的测定可以反映出制品中是否有逆转录病毒的污染 ,不受病毒基因序列的影响。采用PCR法进行逆转录酶活性检测方法 ,以整个生活周期中无DNA过程的MS2RNA为模板 ,设计一对引物 ,在逆转录系统中加入待检样品 ,经过RT PCR扩增后 ,放大检测对象物进行观察 ,从而了解制品中是否存在逆转录酶的污染 ,继而间接推测其是否污染逆转录病毒。对不同生物制品厂家生产的减毒活疫苗的逆转录酶活性进行检测 ,发现在一些疫苗里有逆转录酶活性阳性。
Reverse transcriptase (RT) assays can be used to detect the presence of retroviruses. Retroviruses had been known to infect many species, including humans, and have been associated with neoplasia, immunodeficiencies, leukaemias and lymphomas. A feature of the retrovirus life cycle is the integration of the newly reverse transcribed viral DNA into the host cell genomic DNA,forming the provirus. The presence of the provirus can activate or inactivate cellular genes,possibly with genetic consequences. In addition, retroviruses have high mutation and recombination rates which potentially can alter host range and virulence (Audrey, 1996). Safety testing for retroviruses of cell substrates used to produce biologicals is an important issue.We developed a assay using the MS2 RNA template and primers from the assay developed by Pyra et al. RNA of bacteriophage MS2 serves as the template for RT-mediated cDNA synthesis. A specific region of the cDNA product is then amplified by the polymerase chain reaction to increase the sensitivity of cDNA detection. We improved the method which significantly decrease the level of false positives.;
出处
《微生物学杂志》
CAS
CSCD
2003年第2期11-13,共3页
Journal of Microbiology
