树鼩副粘病毒(TPMV)PCR 方法的建立及初步应用

摘要

Objective To establish and preliminarily apply an effective PCR assay for detection of Tupaia(tree shrew)paramyxovirus(TPMV). Methods Using TPMV genomic DNA from NCBI GenBank, bases 8231 to 8720 were synthesized and inserted into a plasmid as a positive standard. One primer pair was designed based on this sequence. In total,60 respiratory swabs and 12 lung tissues from the tree shrews were tested in this PCR assay. Results A PCR method for detection of TPMV was successfully established,with high specificity and sensitivity of 11.5 × 10 -5μg/mL. PCR result testing 60 respiratory swabs and 12 lung tissues were negative. Conclusions PCR for detecting TPMV has good specificity and high sensitivity and can be used for conventional tree shrew paramyxovirus detection.%目的 建立树鼩副粘病毒(Tupaia paramyxovirus,TPMV)PCR检测方法,并进行初步应用.方法 根据NCBI公布的TPMV基因组序列,选择8231 bp~8720 bp区域人工合成一段DNA序列并转入质粒中,作为阳性标准质粒.根据TPMV保守区域设计一对特异性引物,考察方法的特异性和灵敏度,并应用于60份树鼩呼吸道咽拭子cDNA及12份树鼩肺组织cDNA的检测中.结果 所建立的PCR检测方法特异性好,灵敏度高,可达11.5 × 10 -5μg/mL.60份树鼩呼吸道咽拭子cDNA及12份树鼩肺组织cDNA检测均为阴性.结论 建立的树鼩副粘病毒PCR检测方法特异性强,灵敏度高,可用于树鼩副粘病毒的常规检测中.