家禽糖原合成酶激酶-3β基因荧光定量PCR方法的建立及应用  

Establishment and application of quantification real-time PCR assay of avian glycogen synthase kinase 3βdetection

作　　者：刘轶轩 张沈燕 崔洲昊 乔丹丹 秦爱建 钱琨[1,2,4] LIU Yixuan;ZHANG Shenyan;CUI Zhouhao;QIAO Dandan;QIN Aijian;QIAN Kun(Key Laboratory for Avian Preventive Medicine,Ministry of Education,Yangzhou University,Yangzhou 225009,China;Jiangsu Province Key Laboratory of Preventive Veterinary Medicine,Yangzhou University,Yangzhou 225009,China;Xuzhou Vocational College of Bioengineering,Xuzhou,221006,China;Jiangsu Co-innovation Center for Prevention and Control of Important Animal Infectious Diseases and Zoonoses,Yangzhou,Jiangsu 225009,China)

机构地区：[1]扬州大学教育部禽类预防医学重点实验室,江苏扬州225009 [2]扬州大学江苏省动物预防医学重点实验室,江苏扬州225009 [3]徐州生物工程职业技术学院,江苏徐州221006 [4]江苏省动物重要疫病与人兽共患病防控协同创新中心,江苏扬州225009

出　　处：《畜牧与兽医》2022年第10期129-134,共6页Animal Husbandry & Veterinary Medicine

基　　金：国家自然科学基金(32072839);江苏高校“青蓝工程”项目(苏教师函[2021]11号);江苏省优势学科项目(苏财教[2018]192号);徐州生物工程职业技术学院自然科学研究项目(XSZR202106)。

摘　　要：旨在建立家禽糖原合成酶激酶-3β(GSK-3β)基因荧光定量方法并检测分析马立克病病毒(MDV)感染雏鸡不同脏器组织中GSK-3β基因的表达水平。参考GenBank中提供的GSK-3β参考序列设计全长基因的扩增引物,构建pCAGGS-GSK-3β重组质粒作为阳性标准品,并绘制标准曲线,建立了家禽GSK-3β荧光定量PCR方法。通过家禽GSK-3β基因荧光定量PCR方法检测MDV感染雏鸡不同组织脏器中GSK-3β基因的分布和表达变化。检测表明,病毒感染雏鸡中脑组织表达GSK-3β水平最高(P<0.0001),最低的是盲肠组织(P<0.0001);而正常对照组雏鸡中GSK-3β表达水平最高的也是脑组织,较低的是肝脏组织和胰腺组织。本研究为进一步探究GSK-3β的生物学功能提供了理论依据。This study was to establish an absolute fluorescence quantitative method for gSK-3βgene expression in different organs and tissues of chickens infected with MDV.According to the reference sequence of GSK-3βuploaded in GenBank,a pair of PCR primers for GSK-3βfull-length gene amplification were designed.The recombinant plasmid was constructed and used as the standard plasmid.The specific GSK-3βprimer was designed to establish an absolute quantitative real-time PCR assay for detecting avian GSK-3βgene.The expression and distribution of GSK-3βgenes in different tissues of chickens infected with MDV were detected by this assay.Among the different tissues in the infected SPF chickens,the copy number of the GSK-3βgene was the highest in the brain and the lowest in the cecum.The highest copy number of GSK-3βwas also found in the brain,while the lowest was found in the liver and pancreas in the uninfected chickens.Taken together,the present results indicated that an absolute quantitative real-time PCR assay was successfully established,and the variations in the GSK-3βgene expression in different organs of MDV-infected chickens were detected,which provided theoretical basis for further exploration into the biological function of avian GSK-3β.

关 键 词：糖原合成酶激酶-3Β 荧光定量PCR 应用 

分 类 号：S831[农业科学—畜牧学]