Toline C A, Baker A J
Department of Zoology, University of Toronto, Ontario, Canada.
Mol Ecol. 1995 Dec;4(6):745-53. doi: 10.1111/j.1365-294x.1995.tb00275.x.
Two sections of the control region and the genes coding for NADH dehydrogenase subunits 5 and 6 (ND-5/6) of mitochondrial DNA (mtDNA) were amplified from Phoxinus eos with the polymerase chain reaction. Both sections of the control region were sequenced directly while the ND-5/6 fragment was sequenced in from each end only. Additionally, the entire ND-5/6 fragment was examined for sequence variation using RFLP analysis. No sequence variation was detected in the control region among 70 individuals sampled from 18 populations across three Ontario regions (Spanish River, Madawaska R. and Cataraqui R.). To examine ND-5/6 variation, a total of 75 individuals were sampled from five populations representing two of the three regions (Madawaska River and Cataraqui R.). Six haplotypes were detected by direct sequencing and four by RFLP analysis. Estimates of population subdivision from RFLP data, sequence analysis, and the two data sets combined for the ND-5/6 fragment, suggest that gene flow is restricted within and between regions. However, estimates of sequence divergence for both sequence and RFLP analysis of this fragment suggested that populations were either founded by already differentiated populations or that populations were founded by a single stock and divergence between regions occurred prior to isolation of populations within regions. These estimates of population structure are much greater than those obtained from allozyme analysis. Additionally, high levels of heterozygosity in nuclear DNA, but low mtDNA diversity suggests that populations have experienced reductions in population size sufficient to reduce only mtDNA variation. Random lineage extinction and limited time for the accumulation of new mutations are likely responsible for low levels of mtDNA variation in ND-5/6 and the control region, while functional constraints may limit variation more than expected in the control region in dace and other fishes.
利用聚合酶链反应从溪红点鲑中扩增出线粒体DNA（mtDNA）控制区的两个片段以及编码烟酰胺腺嘌呤二核苷酸脱氢酶亚基5和6（ND-5/6）的基因。控制区的两个片段直接进行测序，而ND-5/6片段仅从两端进行测序。此外，使用限制性片段长度多态性（RFLP）分析检测整个ND-5/6片段的序列变异。在从安大略省三个地区（西班牙河、马达沃斯卡河和卡塔拉基河）的18个种群中采集的70个个体的控制区未检测到序列变异。为了检测ND-5/6的变异，从代表三个地区中的两个地区（马达沃斯卡河和卡塔拉基河）的五个种群中总共采集了75个个体。通过直接测序检测到6种单倍型，通过RFLP分析检测到4种。根据RFLP数据、序列分析以及结合ND-5/6片段的两个数据集对种群细分的估计表明，基因流在区域内和区域间受到限制。然而，对该片段的序列和RFLP分析的序列分歧估计表明，种群要么是由已经分化的种群建立的，要么是由单一群体建立的，并且区域间的分歧发生在区域内种群隔离之前。这些种群结构的估计值远大于从等位酶分析获得的估计值。此外，核DNA中的高杂合度，但线粒体DNA多样性低，表明种群经历了种群数量的减少，足以仅减少线粒体DNA变异。随机谱系灭绝和新突变积累的时间有限可能是导致ND-5/6和控制区线粒体DNA变异水平低的原因，而功能限制可能比预期更限制雅罗鱼和其他鱼类控制区的变异。
