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Development and Preliminary Application of SYBR Green I Real-Time Fluorescence Quantitative PCR Method for Detecting Porcine Parvovirus Virus
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作者 SHEN Zhi-qiang WANG Jin-liang +3 位作者 GUO Xian-po WANG Xiao-hu WANG Ming ZHAO De-ming 《Animal Husbandry and Feed Science》 CAS 2009年第11期42-46,共5页
According to VP2 gene sequence of the porcine parvovirus virus strain NADL-2 (NC001718) available in GenBank (NC_001718), a pair of specific primer was designed, and the target fragment of 431 bp was obtained by P... According to VP2 gene sequence of the porcine parvovirus virus strain NADL-2 (NC001718) available in GenBank (NC_001718), a pair of specific primer was designed, and the target fragment of 431 bp was obtained by PCR amplification. The products were ligated with pMD18- T vector and then transformed into bacteria DH5α for recombinant plasmid extraction. After PCR identification and sequencing, recombinant plasmid was used as a standard template to establish the standard curve of SYBR Green I fluorescence quantitative PCR. Sensitivity test, specificity test and repeatability test were also determined. The results indicated that there was a good linear relationship between threshold cycle of the standard curve and template concentration, R2 =0.997 6. Tm ranged from 82.3 to 82.9 ℃, while the sensitivity was 72.1 copies/μl with good specificity and repeatability. The developed SYBR Green I real-time quantitative PCR method to detect PPV VP2 gene laid the basis for further studies on patho- oenesis, early clinical diaonosis of this virus and quantitative analysis of PPV infection. 展开更多
关键词 Porcine parvovirus virus Real-time fluorescence quantitative PCR DETECTION
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