Preparation and application of PRRSV MS2 armored virus-like particles qualityas control products
10.16303/j.cnki.1005-4545.2024.11.02
- VernacularTitle:猪繁殖与呼吸综合征病毒MS2装甲病毒样颗粒质控品的制备及应用
- Author:
Jiamin HE
1
,
2
;
Xuanfei PANG
;
Lyu LUO
;
Jiazhen YANG
;
Baozhen ZHANG
;
Jianmin WU
;
Wenna LIU
;
Zhongsheng LI
;
Yiquan BAI
Author Information
1. 佛山科学技术学院生命工程与工程学院,广东佛山 528231
2. 广东海大畜牧兽医研究院有限公司,广东 广州 511400
- From:
Chinese Journal of Veterinary Science
2024;44(11):2316-2323
- CountryChina
- Language:Chinese
-
Abstract:
In order to develop a positive quality control products for the detection of porcine repro-ductive and respiratory syndrome virus(PRRSV)nucleic acid by real-time fluorescent quantitative PCR(RT-qPCR),the positive quality control products of PRRSV-1 and PRRSV-2 M genes were prepared using armored RNA technology of MS2 phage.PRRSV-1 and PRRSV-2 M genes were amplified,purified and recovered,and ligated into pET28b vector containing MS2 mature enzyme protein gene and capsid protein.After transformed into BL21(DE3),the gene products were in-duced by IPTG and purified by PEG6000 precipitation method to prepare the armored RNA virus-like particles(AR-PRRSV)containing PRRSV M gene.Following the performance evaluation,as the positive quality control products of PRRSV-1 and PRRSV-2 M genes,AR-PRRSV1M and AR-PRRSV2M were calculated using YY/T 1652-2019 standard.Results showed that it had a good u-niformity,stable storage for the armored virus-like particles at-20,4,25 ℃ for 60 d,and 37 ℃ for 30 d.The prepared armored virus-like particles AR-PRRSV1M and AR-PRRSV2M were deter-mined by digital quantitative PCR(ddPCR)after preliminary quantification by RT-qPCR.The 104 copies/μL of AR-PRRSV1M and AR-PRRSV2M ddPCR fixation was(1.33+0.50)× 104 cop-ies/μL.The above results indicates that the AR-PRRSVM can be used as the quality control of the whole detection process(nucleic acid extraction,reverse transcription and RT-qPCR).
