基于mtDNA Cytb基因和D-loop区序列的陕西林麝遗传多样性分析

doi:10.16431/j.cnki.1671-7236.2022.04.017

摘要/Abstract

摘要： 【目的】探究陕西林麝群体的遗传多样性,了解林麝的遗传信息。【方法】采集林麝毛发提取DNA,测定43只林麝个体的线粒体DNA(mtDNA)细胞色素b(cytochrome b,Cytb)基因和非编码序列置换环(D-loop)区序列,统计碱基组成,使用ClustalX 2.0软件对所有序列进行整合比对,以获得群体中的核苷酸多态位点(SNP);利用DNASP 5.10软件统计种群核苷酸多样性(Pi)、序列的单倍型数(H)、单倍型多样度(Hd)、平均核苷酸差异数(K)等参数;利用Mega 7.0软件计算Cytb基因和D-loop区序列不同单倍型间遗传距离,并构建Neighbor-Joining(NJ)系统进化树。【结果】 Cytb基因和D-loop区序列中AT含量均高于GC含量,显示碱基组成具有偏好性;在Cytb基因和D-loop区内分别发现241和383个SNPs位点。其中,Cytb基因和D-loop区核苷酸多样性分别为0.28343和0.07707,单倍型多样性分别为0.983和0.975,表明其群体遗传多样性丰富;Cytb基因35个单倍型之间的遗传距离为0.002~0.831,D-loop区29个单倍型之间的遗传距离为0.006~1.342。系统进化树结果显示,林麝存在2个线粒体支系,表明其具有2个线粒体母系来源;D-loop区的进化分析也支持这一结论。【结论】林麝群体的核苷酸多样性和单倍型多样较高,遗传多样性较丰富,同时进一步支持了林麝和原麝属于一个分支的观点。

关键词: 林麝; 线粒体DNA; Cytb基因; D-loop区; 遗传多样性

Abstract: 【Objective】 This study was aimed to explore the genetic diversity of Shaanxi Moschus berezovskii population,and understand the genetic information of Moschus berezovskii.【Method】 The hair of Moschus berezovskii was collected to extract DNA,the mitochondrial DNA(mtDNA) cytochrome b(Cytb) gene and D-loop sequences of 43 Moschus berezovskii individuals were determined,and the base composition was counted.All sequences were integrated and compared using ClustalX 2.0 software to obtain nucleotide polymorphic sites (SNPs) in the population.The nucleotide diversity (Pi),number of haplotype (H),haplotype diversity (Hd) and average number of nucleotide differences (K) were calculated by DNASP 5.10 software.The genetic distance among different haplotypes of Cytb gene and D-loop sequences was calculated by Mega 7.0 software,and Neighbor-Joining (NJ) phylogenetic tree was constructed.【Result】 The AT content of Cytb gene and D-loop region were higher than GC content,indicating there was bias in base composition.There were 241 and 383 SNPs of Cytb gene and D-loop region,respectively.The nucleotide diversity of Cytb gene and D-loop region were 0.28343 and 0.07707,and the haplotype diversity was 0.983 and 0.975,respectively,indicating that the population genetic diversity was rich.The genetic distances of 35 haplotypes of Cytb gene ranged from 0.002 to 0.831,and 29 haplotypes of D-loop region ranged from 0.006 to 1.342.The phylogenetic tree showed that there were two mitochondrial lineages,indicating that there were two mitochondrial maternal origins.The evolutionary analysis of D-loop region also supported this conclusion.【Conclusion】 The nucleotide diversity and haplotype diversity of Moschus berezovskii population were high,and the genetic diversity was rich.At the same time further supported the view of Moschus berezovskii and Moshus moschiferus belonged to a branch of the view.

Key words: Moschus berezovskii; mtDNA; Cytb gene; D-loop region; genetic diversity

中图分类号:

S813.3

刘晓玮, 李瑞香, 王兴龙, 李晓. 基于mtDNA Cytb基因和D-loop区序列的陕西林麝遗传多样性分析[J]. 中国畜牧兽医, 2022, 49(4): 1352-1363.

LIU Xiaowei, LI Ruixiang, WANG Xinglong, LI Xiao. Genetic Diversity Analysis of Shaanxi Moschus berezovskii Based on mtDNA Cytb Gene and D-loop Region Sequence[J]. China Animal Husbandry and Veterinary Medicine, 2022, 49(4): 1352-1363.

参考文献

[1] TRAN D,VIET D,TIEN T,et al.New records of the forest musk deer Moschus berezovskii in Vietnam revealed by camera traps[J].Oryx,2021,55(4):494-495.
[2] YANG C,HUANG W,SUN Y,et al.Effect of probiotics on diversity and function of gut microbiota in Moschus berezovskii[J].Archives of Microbiology,2021,203(6):3305-3315.
[3] FENG H,FENG C,HUANG Y,et al.Structure of mitochondrial DNA control region and genetic diversity of Moschus berezovskii populations in Shaanxi province[J].Genetics and Molecular Research,2016,15(2):15027578.
[4] CUI X J,SUN Y L,ZHANG C Q,et al.Post-marketing surveillance of Qishe Pill use for management of neck pain in a Chinese patient cohort to determine its safety,tolerability and effectiveness[J].Chinese Journal of Integrative Medicine,2021,27(6):408-416.
[5] PENG H,LIU S,ZOU F,et al.Genetic diversity of captive forest musk deer (Moschus berezovskii) inferred from the mitochondrial DNA control region[J].Animal Genetics,2009,40(1):65-72.
[6] YANG Q,MENG X,XIA L,et al.Conservation status and causes of decline of musk deer (Moschus spp.) in China[J].Biological Conservation,2003,109:333-342.
[7] JIE H,XU Z,SU Y,et al.The transcriptome analysis of males musk gland in Moschus berezovskii (Artiodactyla:Moschidae)[J].The European Zoological Journal,2019,86:402-412.
[8] ZHAO Y,WANG J,LI Y,et al.Temporal pattern of parturition in captive Alpine musk deer (Moschus sifanicus)[J].Biologia,2019,75:259-266.
[9] 杜欣颖,李东晓,张春梅,等.腹泻对圈养林麝肠道菌群的影响[J].当代畜牧,2021,6:35-40. DU X Y,LI D X,ZHANG C M,et al.Influence of diarrhoea on intestinal microflora of captive forest musk deer(Moschus berezovskii)[J].Contemporary Animal Husbandry,2021,6:35-40.(in Chinese)
[10] 杜欣颖,夏介英,李东晓,等.健康与腹泻林麝肠道菌群的多态性[J].中国微生态学杂志,2021,33(7):745-751. DU X Y,XIA J Y,LI D X,et al.Polymorphism of fetal microbiota of healthy vs diarrhea musk deer[J].Chinese Journal of Microecology,2021,33(7):745-751.(in Chinese)
[11] TIAN Q,ZHOU X,CHENG J,et al.Genome sequence of lung pathogenic Escherichia coli O78,a chimeric strain isolated from pneumonia forest musk deer[J].Genes & Genomics,2017,39:805-815.
[12] LI Y,HU X,YANG S,et al.Comparison between the fecal bacterial microbiota of healthy and diarrheic captive musk deer[J].Frontiers in Microbiology,2018,9:300.
[13] 冯慧,黄原,任轶,等.陕西省林麝mtDNA D-loop区序列结构和种群遗传多样性[J].生态学报,2014,34(20):5887-5895. FENG H,HUANG Y,REN Y,et al.Structure of the mitochondrial DNA D-loop region and genetic diversity of Moschus berezovskii in Shaanxi province[J].Acta Ecologica Sinica,2014,34(20):5887-5895.(in Chinese)
[14] 彭红元,陈伟才,岳碧松,等.麝属分子系统进化地位的再分析[J].四川动物,2011,30(4):530-536. PENG H Y,CHEN W C,YUE B S,et al.A new analysis on the molecular phylogeny of musk deer[J].Sichuan Zoology,2011,30(4):530-536.(in Chinese)
[15] SU B,WANG Y X,LAN H,et al.Phylogentic study of complete cytochromeb gene sin musk deer(gene Moschus) using museum samples[J].Molecular Phylogenetics & Evolution,1999,12(3):241-249.
[16] GUHA S,GOYAL S P,KASHYAP V K.Molecular phylogeny of musk deer:A genomic view with mitochondrial 16S rRNA and cytochrome b gene[J].Molecular Phylogenetics & Evolution,2007,42(3):585-597.
[17] 肖宇辰,伊丽,孟根达来,等.麝科动物的起源进化与遗传多样性研究进展[J].家畜生态学报,2019,40(8):8-13. XIAO Y C,YI L,MENG G D L,et al.Advances in tne origin and evolution and genetic diversity of Moschiae[J].Chinese Journal of Livestock Ecology,2019,40(8):8-13.(in Chinese)
[18] 盛和林,刘志晓.中国麝科动物[M].上海:上海科学技术出版社,2007. SHENG H L,LIU Z X.China Moschiae[M].Shanghai:Shanghai Science and Technology Press,2007.(in Chinese)
[19] 孟根达来,MYAGMARSUREN,BATKHUU,等.麝属动物的分类学研究历史沿革[J].野生动物学报,2018,39(4):966-971. MENG G D L,MYAGMARSUREN,BATKHUU,et al.Historical evolution of taxonomy research on musk deer species[J].Chinese Journal of Wildlife,2018,39(4):966-971.(in Chinese)
[20] SKURIKHINA L A,KUKHLEVSKY A D,KOVPKA N E.Relationships of osmerid fishes (Osmeridae) of Russia:Divergence of nucleotide sequences of mitochondrial and nuclear genes[J].Genes & Genomics,2013,35(4):529-539.
[21] CUI Z,LIU Y,CHU K H.Broader pattern of tandem repeats in the mitochondrial control region of Perciformes[J].Chinese Journal of Oceanology & Limnology,2010,28(4):785-794.
[22] 肖逸,王孟文,张秀英,等.基于mtDNA Cytb基因的马可·波罗盘羊遗传多样性与起源进化分析[J].中国畜牧兽医,2021,48(5):1681-1690. XIAO Y,WANG M W,ZHANG X Y,et al.Genetic diversity,origin and evolution analysis of Ovis ammon polii based on mtDNA Cytb gene[J].China Animal Husbandary & Veterinary Medicine,2021,48(5):1681-1690.(in Chinese)
[23] BROWN G G,GADALETA G,PEPE G,et al.Structural conservation and variation in the D-loop-containing region of vertebrate mitochondrial DNA[J].Journal of Molecular Biology,1986,192(3):503-511.
[24] WILKINSON G S,CHAOMAN A M.Length and sequence variation in evening bat D-loop mtDNA[J].Genetics,1991,128(3):607-617.
[25] 周月琴,朱伟,刘志萍,等.用Chelex-100快速提取微量血痕中的DNA[J].复旦学报(医学版),2003,30(4):379-380. ZHOU Y Q,ZHU W,LIU Z P,et al.A quick method of extraction of DNA by Chelex-100 from trace bloodstains[J].Fudan University Journal of Medical Sciences,2003,30(4):379-380.(in Chinese)
[26] 冯慧,冯成利,刘晓农,等.林麝毛发DNA的提取及系统发育分析[J].西北农业学报,2012,21(8):14-18. FENG H,FENG C L,LIU X N,et al.Hair DNA extraction and phylogenetic analysis of forest musk deer(Moschus berezovskii)[J].Acta Agriculturae Boreali-Occidentalis Sinica,2012,21(8):14-18.(in Chinese)
[27] GRANT W A S,BOWEN B W.Shallow population histories in deep evolutionary lineages of marine fishes:Insights from sardines and anchovies and lessons for conservation[J].Journal of Heredity,1998,89(5):415-426.
[28] HONG L,LI M S.The origin and genetic differentiation of native breeds of pigs in southwest China:An approach from mitochondrial DNA polymorphism[J].Biochemical Genetics,1993,31(1-2):51-60.
[29] 周慧,李迪强,张于光,等.藏羚羊mtDNA D-loop区遗传多样性研究[J].遗传,2006,28(3):299-305. ZHOU H,LI D Q,ZHANG Y G,et al.Study on mitochondrial DNA genetic diversity of Tibetan antelope[J].Genetics,2006,28(3):299-305.(in Chinese)
[30] HAYANO A,YOSHIOKA M,TANAKA M,et al.Population differentiation in the Pacific white-sided dolphin Lagenor hynchus obliquidens inferred from mitochondrial DNA and microsatel lite analyses[J].Science,2004,21(9):989-999.
[31] 文榕生.历史上中国麝的分布变迁[J].晋中学院学报,2016,33(5):68-74. WEN R S.The distribution and changes of musk deer(Moschus spp.) in China in the historical period[J].Journal of Jinzhong University,2016,33(5):68-74.(in Chinese)
[32] 章敬旗,周友兵,徐伟霞,等.几种麝分类地位的探讨[J].西华师范大学学报(自然科学版),2004,25(3):251-255. ZHANG J Q,ZHOU Y B,XU W X,et al.Discussion about musk-deer’s classifition[J].Journal of China West Normal University (Natural Science Edition),2004,25(3):251-255.(in Chinese)