【作者】 高晶；

【导师】 冯江；

【作者基本信息】 东北师范大学 ， 环境科学， 2006， 硕士

【摘要】 马铁菊头蝠(Rhinolophus ferrumequinum)属于旧大陆物种,在欧洲大部分地区处于濒危状态,现已被世界自然保护联盟(IUCN )受威胁物种红色名录列为低危/接近易危(LR/nt)种。我们从吉林省集安市和云南省昆明市采集了8个马铁菊头蝠个体,从其肌肉组织中提取全基因组DNA,利用PCR的方法扩增出线粒体DNA的控制区和细胞色素b基因序列并测序,应用MEGA、DnaSP等软件对其序列进行分析,探索遗传变异在吉林和云南样本中的分布及其马铁菊头蝠种内系统发育关系。主要研究结果如下:1.实验获得了8个个体mtDNA控制区部分序列,定义了6个单倍型,单倍型间的核苷酸变异频率为9.71%,变异主要发生在ETAS和CSB结构域中,中心保守结构域( CD结构域)的替换很少( ts=1, tv=3 ),表现较高的保守性。2.实验获得了6个个体mtDNA细胞色素b基因全序列。经序列比对发现吉林和云南群体各共享一个单倍型, 2个单倍型之间的核苷酸变异频率为3.68%。吉林和云南的样本在密码子使用频率上表现出较高的相似性,在使用偏好上表现出对AU的相对偏倚。3.利用MEGA3对mtDNA控制区1019bp序列进行比对,计算得到吉林和云南群体内的平均遗传距离分别是0.003和0.006,群体间的遗传距离为0.096,变异主要发生在群体之间,而群体内的遗传分化水平较低,其中马铁菊头蝠的吉林哺育群体的单倍型多样性指数和核苷酸多样性指数仅为0.800±0.172,0.00262±0.00056。4.基于细胞色素b全序列构建的系统进化树显示,吉林样本与日本马铁菊头蝠被划分在一个进化单元里,系统发育关系支持将吉林样本鉴定为马铁菊头蝠日本亚种(Rhinolophus ferrumequinum nippon)。云南样本和吉林样本之间的遗传分化程度较高,基于细胞色素b基因计算遗传距离达到0.040。更多还原

【Abstract】 The greater horseshoe bat ( Rhinolophus ferrumequinum ) distributed through most ofthe old world, and the genus includes a high incidence of species of conservation concernin all over Europe. Now it has been listed as a LR/nt species by 2004 IUCN Red List ofThreatened Species by International Union for Conservation of Nature and NaturalResource ( IUCN ). Eight individuals of greater horseshoe bats were captured from Ji’ancity in Jilin province and Kunming city of Yunnan province in China. The whole genomeDNA was extracted from their muscle tissues as templates by which the mitochondrialDNA control region and cytochrome b gene were amplified with polymerase chain reaction( PCR ). After being sequenced, the sequences of control region and cytochrome b genewere analyzed by MEGA and DnaSP to explore genetic variation and phylogeneticrelationships of the samples from Jilin and Yunnan provinces. The results were as follows:1. The control region sequences of eight individuals were obtained. The results ofalignment showed six haplotypes and nucleotide substitutions mainly occurred in extendedtermination associated sequences ( ETAS ) and conserved sequence block ( CSB ) domains,but there were only four substitutions ( ts=1, tv=3 ) in the conserved central domain ( CD ).The nucleotide variation of the eight mitochondrial DNA control regions was 9.71%.2. The cytochrome b ( cytb ) gene sequences of six individuals were obtained. Theresults of alignment showed four individuals from Jilin shared the same haplotype( TYPE-JA ) and two from Yunnan shared another same haplotype ( TYPE-YN ).Nucleotide variation of the six sequences was 3.68%. The two haplotypes were mostlysimilar in codon usage and showed a bias of adenine(A) and uracil(U).3. The six sequences of control regions of 1019bp were aligned and their geneticdistances were calculated by MEGA3. The results showed the average distances in samplesfrom Jilin and Yunnan were 0.003 and 0.006 respectively, but the distance between Jilinand Yunnan was 0.096. The results indicated that variation between the two areas is moredistinct than in each area. Additionally, the haplotype diversity and nucleotide diversitywere 0.800±0.172 and 0.00262±0.00056 in Jilin samples, respectively.4. A neighbour-joining tree was constructed for the phylogenetic analysis based onnucleotide sequences of cytochrome b gene. TYPE-JA haplotype and the sequences fromJapan were clustered in one clade, which suggested that TYPE-JA haplotype was judged asone of subspecies of greater horseshoe bats, Rhinolophus ferrumequinum nippon,. Thedegree of variation between TYPE-JA and TYPE-YN was high, and their genetic distancewas 0.040.更多还原