不同遗传背景猪PBD-110基因多态性及表达谱分析

doi:10.16431/j.cnki.1671-7236.2021.03.021

摘要/Abstract

摘要： 为探究猪β-防御素110（PBD-110）基因多态性及其在猪组织中的表达情况，本试验以野杂猪、民猪和大白猪为研究对象，分别扩增PBD-110基因编码区（CDS）和内含子区，采用实时荧光定量PCR法探究3个群体猪肝脏和脾脏的组织表达差异。试验成功扩增出猪PBD-110基因CDS区和内含子区，测序证实3个群体猪PBD-110基因CDS区不存在突变位点，基因组PCR产物测序发现在内含子1存在C314T突变位点，3个群体多态性分析发现CC为优势基因型，基因型频率依次为0.750、0.781和0.516。Hardy-Weinberg平衡检验同时发现该位点在3个群体中均处于平衡状态（P>0.05），且处于中、低多态性状态（0.25 < PIC < 0.5；PIC < 0.25）。表达谱分析结果表明，PBD-110基因在3个群体猪肝脏和脾脏中均有表达，且存在表达差异，提示PBD-110基因可能与民猪、野杂猪抗病力较强有关，在天然免疫方面发挥重要作用。

关键词: 大白猪; 民猪; 野杂猪; PBD-110基因; 多态性; 组织表达

Abstract: In order to explore the polymorphism and tissue expression of porcine β-defensin 110 (PBD-110) gene,the CDS region and intron regions of PBD-110 gene were amplified in crossbred from wild boars,Min pigs and Large White pigs.Finally,the difference of tissue expression in liver and spleen of the three populations was explored by Real-time quantitative PCR method.The results showed that porcine PBD-110 gene CDS region and intron regions were amplified.Sequencing confirmed that there were no mutation sites in CDS region and genomic PCR product sequencing found that C314T mutation in intron 1.Polymorphism analysis showed that CC was the dominant genotype,and the genotype frequency were 0.750,0.781 and 0.516,respectively.Hardy-Weinberg equilibrium test also showed that the locus was in equilibrium and moderately or lowly polymorphic (0.25 < PIC < 0.5;PIC<0.25) in three populations.The results of expression profile analysis showed that PBD-110 gene was expressed in liver and spleen of three populations of pigs,and there were different expression pattern.PBD-110 gene might play an important role in innate immunity.

Key words: Large White pigs; Min pigs; crossbred from wild boars; PBD-110 gene; polymorphism; tissue expression

中图分类号:

S828

刘艳光, 罗新惠, 柳俭强, 贾琪, 肖成, 曹阳, 金海国, 张立春. 不同遗传背景猪PBD-110基因多态性及表达谱分析[J]. 中国畜牧兽医, 2021, 48(3): 975-981.

LIU Yanguang, LUO Xinhui, LIU Jianqiang, JIA Qi, XIAO Cheng, CAO Yang, JIN Haiguo, ZHANG Lichun. Analysis of Polymorphism and Expression Profile of PBD-110 Gene in Pigs with Different Genetic Background[J]. China Animal Husbandry and Veterinary Medicine, 2021, 48(3): 975-981.

参考文献

[1] 单安山,田昊天,邵长轩,等.抗菌肽抗细菌机理研究进展[J].东北农业大学学报.2018,49(3):84-94. SHAN A S,TIAN H T,SHAO C X,et al.Research advance on antibacterial mechanism of antimicrobial peptides[J].Journal of Northeast Agricultural University,2018,49(3):84-94.(in Chinese)
[2] 徐敏,余娟,贺会利,等.动物抗菌肽基因工程表达研究进展[J].中国兽医杂志,2013,49(9):59-62. XU M,YU J,HE H L,et al.Advances in genetic engineering expression of animal antimicrobial peptides[J].Chinese Journal of Veterinary Medicine,2013,49(9):59-62.(in Chinese)
[3] GANZ T.Defensin:Antimicrobial peptides of innate immnuity[J].Nature Reviews Immunology,2003,3(9):710-720.
[4] 王东阳,包图雅.哺乳动物雌性生殖系统β-防御素的研究进展[J].畜牧与饲料科学,2019,40(6):30-33. WANG D Y,BAO T Y.Research progress on β-defensin in female mammalian reproductive system[J].Animal Husbandry and Feed Science,2019,40(6):30-33.(in Chinese)
[5] 万遂如.养猪生产中要大力控制抗生素的使用[J].养猪,2014,2:117-120. WAN S R.Great efforts should be made to control the use of antibiotics in pig production[J].Swine Production,2014,2:117-120.(in Chinese)
[6] MIN K C,MINH T L,DINH T N,et al.Genome-level identification,gene expression,and comparative analysis of porcine β-defensin genes[J].BMC Genetics,2012,13(1):98.
[7] 张坤.猪β防御素2对大肠杆菌感染的IPEC-J2细胞NF-κB和MAPK信号通路基因表达的影响[D].郑州:河南农业大学,2017. ZHANG K.Influence of porcine beta defensin 2 on the gene expression of NF-κB and MAPK signaling pathways in E.coli infection IPCE-J2 cell[D].Zhengzhou:Henan Agricultural University,2017.(in Chinese)
[8] SANG Y,PATIL A A,ZHANG G,et al.Bioinformatic and expression analysis of novel porcine beta-defensins[J].Mammalian Genome,2006,17(4):332-339.
[9] 刘艳光,张珈溯,罗新惠,等.不同品种猪PBD-119基因的克隆与差异表达研究[J].延边大学农学学报,2020,42(2):75-82. LIU Y G,ZHANG J S,LUO X H,et al.Cloning and differential expression of PBD-119 gene in different breeds of pigs[J].Agriculture Science Journal of Yanbian University,2020,42(2):75-82.(in Chinese)
[10] TOMAS H,MARKUS W,MYLENE D,et al.RefGenes:Identification of reliable and condition specific reference genes for RT-qPCR data normaliza-tion[J].BMC Genomics,2011,12(1):156.
[11] 张立春,李兆华,孙福亮,等.长白山野杂猪、民猪和大白猪血常规及免疫学指标比较分析[J].中国兽医学报,2018,38(1):171-176. ZHAN L C,LI Z H,SUN F L,et al.The comparative analysis of blood routine and immune index among Large White,Min pig and the crossbred of wild boars from Changbai mountain[J].Chinese Journal of Veterinary Science,2018,38(1):171-176.(in Chinese)
[12] 李梦姝,孙福亮,柳俭强,等.不同遗传背景猪TNF-α基因启动子区多态性与其血清含量的关联分析[J].中国预防兽医学报,2019,41(1):69-73. LI M S,SUN F L,LIU J Q,et al.Polymorphism of TNF-α gene promoter region and correlation analysis of serum TNF-α concentration in different genetic background pig populations[J].Chinese Journal of Preventive Veterinary Medicine,2019,41(1):69-73.(in Chinese)
[13] 谭雯雯.猪β-防御素2多克隆和单克隆抗体的制备及初步应用[D].武汉:华中农业大学,2010. TAN W W.Generation and application of polyclone and monoclonal antibodies against porcine beta-defensin 2[D].Wuhan:Huazhong Agricultural University,2010.(in Chinese)
[14] 戴超辉,戴开宇,宗秋芳,等.梅山猪Mx1基因第14外显子多态性及其与繁殖性能的关联分析[J].中国畜牧兽医,2015,42(12):3282-3287. DAI C H,DAI K Y,ZONG Q F,et al.Polymorphism of Mx1 gene exon 14 and its relationship with reproductive performance in Meishan pigs[J].China Animal Husbandry & Veterinary Medicine,2015,42(12):3282-3287.(in Chinese)
[15] 陈惠娴,毛若雨,滕达,等.二硫键及其连接方式对防御素抗菌功能的影响研究进展[J].生物技术通报,2016,32(9):32-37. CHEN H X,MAO R Y,TENG D,et al.Review on effects of disulfide bond and its connection on antimicrobial activity in defensin[J].Biotechnology Bulletin,2016,32(9):32-37.(in Chinese)
[16] 刘璨颖,陈志胜,林裕锋,等.猪源β-防御素2和3的研究进展[J].中国畜牧杂志,2019,55(8):30-34. LIU C Y,CHEN Z S,LIN Y F,et al.Research progress and application prospect of porcine β-defensin 2 and β-defensin 3[J].Chinese Journal of Animal Science,2019,55(8):30-34.(in Chinese)
[17] 刘陶林,铁宇娟,艾显辉,等.哺乳动物雄性生殖系统β-防御素的研究进展[J].中国兽医科学,2018,48(11):1458-1463. LIU T L,TIE Y J,AI X H,et al.Research progress of β-defensins in mammalian male reproductive system[J].Chinese Veterinary Science,2018,48(11):1458-1463.(in Chinese)
[18] 魏薇,李海军,王鑫,等.家畜防御素体内组织表达及体外表达研究进展[J].畜牧与饲料科学,2019,40(4):52-57. WEI W,LI H J,WANG X,et al.Research advances on in vivo tissue expression and in vitro expression of defensin in livestock[J].Animal Husbandry and Feed Science,2019,40(4):52-57.(in Chinese)
[19] 魏薇.马、驴、马骡β-防御素1基因克隆、体内表达及原核表达研究[D].呼和浩特:内蒙古农业大学,2019. WEI W.Research for the expression in vivo and prokaryotic expression of β-defensin 1 in horse,ass and mule[D].Hohhot:Inner Mongolia Agricultural University,2019.(in Chinese)
[20] SU G,XIE K,CHEN D,et al.Differential expression,molecular cloning,and characterization of porcine beta defensin 114[J].Journal of Animal Science and Biotechnology,2019,10(4):884-894.