[1]龚双燕,李小璟,李幽幽,等.猪传染性胃肠炎病毒SYBR GreenⅡ荧光定量PCR检测方法的建立及在初乳检测上的应用[J].江苏农业学报,2017,(05):1076-1081.[doi:doi:10.3969/j.issn.1000-4440.2017.05.018]
　GONG Shuang-yan,LI Xiao-jing,LI You-you,et al.Establishment of SYBR Green Ⅱ fluorescence quantitative PCR for detection of infectious gastroenteritis virus and its application in colostrum detection[J].,2017,(05):1076-1081.[doi:doi:10.3969/j.issn.1000-4440.2017.05.018]
点击复制

猪传染性胃肠炎病毒SYBR GreenⅡ荧光定量PCR检测方法的建立及在初乳检测上的应用()
分享到：

江苏农业学报[ISSN:1006-6977/CN:61-1281/TN]

卷:
期数:
2017年05期
页码:
1076-1081
栏目:
畜牧兽医·水产养殖
出版日期:
2017-10-30

文章信息/Info

Title:
Establishment of SYBR Green Ⅱ fluorescence quantitative PCR for detection of infectious gastroenteritis virus and its application in colostrum detection
作者:
龚双燕1; 李小璟1; 李幽幽1; 陈瑛琪1; 蔡瑶1; 李雨濛1; 徐逸飞1; 徐志文1; 2; 朱玲1; 2
（1.四川农业大学动物医学院，四川成都611130；2.四川农业大学动物疫病与人类健康四川省重点实验室，四川成都611130)
Author(s):
GONG Shuang-yan1; LI Xiao-jing1; LI You-you1; CHEN Ying-qi1; CAI Yao1; LI Yu-meng1; XU Yi-fei1; XU Zhi-wen1; 2; ZHU-Ling1; 2
(1.College of Veterinary Medicine, Sichuan Agricultural University, Chengdu 611130, China;2.Key Laboratory of Animal Disease and Human Health of Sichuan Province, Sichuan Agricultural University, Chengdu 611130, China)
关键词:
猪传染性胃肠炎病毒; 荧光定量PCR检测; 初乳
Keywords:
transmissible gastroenteritis virus; fluorescence quantitative PCR; colostrum
分类号:
S858.282.65
DOI:
doi:10.3969/j.issn.1000-4440.2017.05.018
文献标志码:
A
摘要:
为了提高初乳中猪传染性胃肠炎病毒(Transmissible gastroenteritis virus，TGEV)的检出率，避免初生仔猪受到带毒初乳的危害，建立了一种SYBR GreenⅡ荧光定量PCR(qPCR)检测方法。根据GenBank已发表的TGEV的N基因序列，选择保守区域设计、合成了1对特异性引物，扩增TGEV的N基因片段（174 bp），构建含有N基因片段的重组质粒，以重组质粒作为模板建立了特异性检测TGEV的qPCR检测方法。该方法的灵敏性比普通PCR方法高，应用建立的qPCR检测方法，对2016年10月-2017年4月采集自四川省部分地区腹泻猪场的母猪初乳130份进行检测，并与普通PCR检测方法进行比较。结果显示：用qPCR方法检测，130份样本中有13份为TGEV阳性，而普通PCR方法只检测到5份TGEV阳性样本。说明qPCR检测方法的敏感性高于普通PCR方法，更适合用于猪初乳检测。
Abstract:
A SYBR Green II fluorescence quantitative PCR (qPCR) method was established to improve the detection rate of transmissible gastroenteritis virus (TGEV) in colostrum, and to avoid the damage to primary piglets by toxic colostrum. According to the N gene sequence of TGEV published in GenBank, the conservative area was selected to design a pair of specific primers, and the amplified N gene fragment was 174 bp. A recombinant plasmid containing N gene fragment was used as template to establish a qPCR method for the specific detection of TGEV. The qPCR method was used to detect 130 samples of sow colostrum collected from October 2016 to April 2017 in some parts of Sichuan province (diarrhea pig farm). The results showed that 13 of the 130 samples were positive for TGEV by qPCR, and only five samples of TGEV were detected by common PCR. This indicated that qPCR method was more sensitive than the common PCR method, and the qPCR method was more suitable for colostrum testing.

参考文献/References:

[1]殷震,刘景华. 动物病毒学[M]. 2版.北京:科学出版社, 1997.
[2]郭容利,李彬,范宝超,等. 猪传染性胃肠炎病毒分离株 JS2012 全基因组序列分析[ J]. 江苏农业学报,2016,32(6):1351-1358.
[3]JEFFREY J Z. 猪病学[M]. 赵德明,译.北京:中国农业大学出版社, 2014.
[4]张永香. 初乳中分泌型免疫球蛋白(SIgA)在防控新生仔猪腹泻方面的研究进展[J]. 中国动物保健, 2017, 19(2):72-75.
[5]VAN WETTERE W H, PAIN S J, HUGHES P E. Dietary ractopamine supplementation during the first lactation affects milk composition, piglet growth and sow reproductive performance[J]. Animal Reproduction Science, 2016, 174:87-92.
[6]BRANDTZAEG P. Induction of secretory immunity and memory at mucosal surfaces[J]. Vaccine, 2007, 25(30):5467-5484.
[7]孙瑞芹. 华南地区猪流行性腹泻病毒的分子流行病学与病原学研究[D]. 咸阳:西北农林科技大学, 2014.
[8]SUN R Q, CAI R J, CHEN Y Q, et al. Outbreak of porcine epidemic diarrhea in suckling piglets, China[J]. Emerging Infectious Diseases, 2012, 18(1):161-163.
[9]朱密. 猪传染性胃肠炎病毒分子生物学检测方法研究进展[J]. 安徽农学通报, 2016, 22(12):110-111.
[10]BRIAN D A, DENNIS D E, GUY J S. Genome of porcine transmissible gastroenteritis virus[J]. Journal of Virology, 1980, 34(2):410-415.
[11]李继良,王保有,董树仁,等. 猪传染性胃肠炎病毒荧光定量PCR检测方法的建立[J]. 北京农学院学报, 2016, 31(3):67-70.
[12]张雪,杨倩,于红欣,等. 猪传染性胃肠炎病毒实时荧光定量PCR检测方法的建立与应用[J]. 中国兽医科学, 2015(6):644-648.

备注/Memo

备注/Memo:
收稿日期：2017-06-08 基金项目：四川省科技支撑计划项目(2017NZ0038)； “十二五”农村领域国家科技计划课题(2015BAD12B04-2.3) 作者简介：龚双燕(1994-)，女，重庆开县人，硕士研究生，主要从事动物传染病病原分子生物学研究。(E-mail)gsymanas@163.com 通讯作者：朱玲，(E-mail)abtcxzw@126.com
更新日期/Last Update: 2017-11-03