Microsatellite markers in common carp (Cyprinus carpio L.)

Microsatellite markers of the poly (CA) type in common carp ( Cyprinus carpio L.) are described. Clones containing a (CA) repeat were isolated from a common carp genomic library and sequenced. The number of repeats found was high compared to mammals but comparable with other teleost fishes. Classification of the repeats (perfect, imperfect and compound) are compared with the Atlantic cod ( Gadus morhua L.), rainbow trout ( Oncorhynchus mykiss ), and Atlantic salmon ( Salmo salar L.). A total of 41 primer sets were designed and tested for polymorphism on a test panel of eight animals (derived from outbred lines, inbred lines and gynogenetic clones). Thirty-two markers were found to be polymorphic. The heterozygosity in the outbred animals was 60·4%, 51·1% in the inbred animals and 0% in the gynogenetic clones. The average number of alleles among the eight animals was 4·7 per marker. Six markers (18·8%) gave an additional polymorphic amplification product besides the polymorphic amplification product in the expected size range. The possibility that these loci are tetraploid is discussed. The polymorphic loci described for common carp will be valuable as genetic markers for use in population, breeding, and evolutionary studies.     

Cloning and characterization of microsatellite loci in channel catfish, Ictalurus punctatus

Short tandem repeat (microsatellite) loci were cloned from the channel catfish, Ictalurus punctatus , genome for use as molecular markers for genetic improvement of this important agricultural species. Plasmid clones containing catfish genomic DNA inserts were identified, by hybridization with tandem repeat DNA probes, and sequenced using automated laser fluorescence. A feral population of catfish displayed levels of heterozygosity greater than 0·7 for 13 of 22 loci and heterozygosity greater than 0·5 for 20 of 22 loci. Allelic polymorphism ranged from three to 17 alleles per locus in the feral population. Populations of domestic, farm-raised catfish and a research strain displayed levels of heterozygosity similar to the feral population. Non-invasive tissue sampling provided abundant material for the polymerase chain reaction-based genotype assay. The microsatellite loci will be useful in the molecular characterization and genetic improvement of channel catfish populations.     

Biochemical polymorphisms in feral and inbred mice (Mus musculus)

Examination of the frequencies of several loci controlling isozymes in three geographically distinct feral populations of mice showed the average animal to be heterozygous at 10.3% of his loci. There was no evidence for interaction between loci, nor any evidence for inbreeding in the populations. Thirty-nine inbred strains, including four newly derived ones, were also characterized for their alleles for as many as 16 polymorphic loci. Among these strains, variability is at least as great as in any single feral population, but probably less than that found among all feral populations of the species.This work was supported by NIH Research Grants GM-9966 and GM-07249 from the Division of General Medical Science, and by contracts AT(30-1)-1979 and AT(30-1)-3671 with the U.S. Atomic Energy Commission.     

PCR扩增近交系大鼠微卫星位点DNA多态性的研究

本实验选取大鼠7条染色体上的微卫星位点合成了10对引物,利用聚合酶链反应（PCR）扩增技术对国内北京和哈尔滨等4家单位提供伯6个品系（SHR、SHRSP、LEW、RCS、WKY和F344）的8个近交系大鼠群体进行了DNA多态性分析的研究。结果表明：9个微卫星位点具有显多态性;不同品系个体之间具有多态性;同一群体不同个体之间除SHR（哈）的SMST位点和WKY（哈）的AGT位点出现一定的差异,其他均没有差异;不同地区同一品系的不同个体之间也存在一定的差异。该方法能有效地对近交系与杂交系、品系与品系、品系与亚系加以区分。因此,本实验为开展近交系大鼠遗传作图、基因定位和为实验动物的遗传背景监测提供可靠的信息,为大鼠遗传基因的研究提供了一个快速简例、特异准确的方法。     

Polymorphic microsatellite markers in the outbred CFW and ICR stocks for the generation of speed congenic mice on C57BL/6 background

Reliable definition of the phenotype of particular alleles is carried out in the genetic background of inbred strains. Appearance of mutations in outbred mice therefore requires the generation of congenic mice. The aim of this study was the establishment of a list of polymorphic microsatellite markers which can be used in a polymerase chain reaction (PCR)-based marker-assisted selection protocol (MASP) to allow the use of the two common outbred stocks, CFW and ICR, as donor animals for the fast generation of congenic C57BL/6 mice. The selection of informative microsatellite markers was carried out to provide a simple evaluation of the PCR products by conventional agarose gel electrophoresis. Outbred mice from three suppliers were examined. In total, 153 microsatellite loci were analysed. Here we present 76 and 70 microsatellite markers polymorphic for the outbred ICR and CFW stocks compared to C57BL/6. At least three microsatellite loci per chromosome were chosen as informative markers for the autosomal genome, giving rise to a maximum marker distance of 58 cM. Thus, additional individual markers have to be selected for the respective outbred mouse which is chosen as a donor animal.     

Major histocompatibility complex heterozygosity reduces fitness in experimentally infected mice

It is often suggested that heterozygosity at major histocompatibility complex (MHC) loci confers enhanced resistance to infectious diseases (heterozygote advantage, HA, hypothesis), and overdominant selection should contribute to the evolution of these highly polymorphic genes. The evidence for the HA hypothesis is mixed and mainly from laboratory studies on inbred congenic mice, leaving the importance of MHC heterozygosity for natural populations unclear. We tested the HA hypothesis by infecting mice, produced by crossbreeding congenic C57BL/10 with wild ones, with different strains of Salmonella, both in laboratory and in large population enclosures. In the laboratory, we found that MHC influenced resistance, despite interacting wild-derived background loci. Surprisingly, resistance was mostly recessive rather than dominant, unlike in most inbred mouse strains, and it was never overdominant. In the enclosures, heterozygotes did not show better resistance, survival, or reproductive success compared to homozygotes. On the contrary, infected heterozygous females produced significantly fewer pups than homozygotes. Our results show that MHC effects are not masked on an outbred genetic background, and that MHC heterozygosity provides no immunological benefits when resistance is recessive, and can actually reduce fitness. These findings challenge the HA hypothesis and emphasize the need for studies on wild, genetically diverse species.     

Isozyme polymorphisms maintained by lethal loci in inbred strains of Aedes triseriatus

We analyzed the progeny of brother-sister matings of two inbred strains of the treehole mosquito, Aedes triseriatus, at several isozyme loci to determine which factors were maintaining heterozygosity at these loci after prolonged inbreeding. The triseriatus Kramer (TK) lines were polymorphic for the Odh and Hbd loci after 12 generations of full-sib mating, and the triseriatus Vero Beach (TV) lines were polymorphic for the Odh, Hk-4, Pgm, and Hbd loci after 14 generations of full-sib mating. Genetic analysis of 22 TK F13 matings and nine TV F15 matings showed that heterozygosity was enforced by lethal loci to which all the polymorphic isozyme loci were linked. The lethal loci formed a balanced lethal system. We determined the relative positions and map distances of lethals and enzyme loci.     

Gene flow in the exotic 14-spotted ladybird beetle, Propylea quatuordecimpunctata.

Propylea quatuordecimpunctata is a Palearctic ladybird beetle that has recently become established fortuitously in northeastern North America. This predator has also been cultured by the United States Department of Agriculture from geographically diverse Old World populations and propagated for release in the U.S.A. Massive releases of P. quatuordecimpunctata have not resulted in any recognized established populations, raising a number of questions about its population genetics. We sampled beetles for electrophoretic variation in three feral populations in the U.S.A., one feral population in Europe, one cultured population from Canada, and six cultured populations from the Old World. Of the 31 putative genetic loci resolved, 26 were polymorphic (84%). Mean heterozygosity was 18.32 +/- 2.84% among all loci and was 21.84 +/- 2.89% among only polymorphic loci. Cultured and feral populations showed similar levels of heterozygosity at the 11 loci scored. New World and Old World beetles showed no significant differences in heterozygosities. Wright's fixation index FST was 0.034 +/- 0.021 among feral populations and 0.331 +/- 0.101 among cultured populations. Drift was the major force driving differentiation of cultured beetle populations. Our data do not suggest that the failure of P. quatuordecimpunctata to become established in areas where it was deliberately released was related to a paucity of genie diversity. Key words : allozymes, breeding structure, biological control, gene flow, colonization.     

Genome-wide genetic diversity of 'Nici', the DNA source for the CHORI-260 turkey BAC library and candidate for whole genome sequencing

Vertebrate whole genome sequence assembly can benefit from a priori knowledge of variability in the target genome, with researchers often selecting highly inbred individuals for sequencing. However, for most species highly inbred research lines are lacking, requiring the use of an outbred individual(s). Here we examined the source DNA [Nicholas inbred (Nici)] of the CHORI-260 turkey bacterial artificial chromosome (BAC) library through analysis of microsatellites and BAC sequences. Heterozygosity of Nici was compared with that of individuals from several breeder lines. Seventy-eight microsatellites were screened for polymorphism in a total of 43 birds, identifying an average individual heterozygosity of 0.39, with Nici at 0.35. Additional loci (total of 147) were examined on a subset of individuals to obtain better genome coverage. The mean heterozygosity for this subset was 0.33 with Nici at 0.31. Examination of approximately 200 kb of genome sequence identified SNPs in the order of one per 200 bp in Nici. These data suggest that the heterozygosity of Nici is comparable to other birds of selected breeder lines and that whole genome sequencing would result in an abundant resource of genome-wide polymorphisms.     

EFFECTS OF POPULATION BOTTLENECKS ON GENETIC DIVERSITY AS MEASURED BY ALLOZYME ELECTROPHORESIS

Low levels of allozyme heterozygosity in populations are often attributed to previous population bottlenecks; however, few experiments have examined the relationship between heterozygosity and bottlenecks under natural conditions. The composition and number of founders of 55 experimental populations of the eastern mosquitofish (Gambusia holbrooki), maintained under simulated field conditions, were manipulated to examine the effects of bottlenecks on three components of allozyme diversity. Correlations between observed and expected values of allozyme heterozygosity, proportions of polymorphic loci, and numbers of alleles per locus were 0.423, 0.602, and 0.772, respectively. The numbers of polymorphic loci and of alleles per locus were more sensitive indicators of differences in genetic diversity between the pre-bottleneck and post-bottleneck populations than was multiple-locus heterozygosity. In many populations, single- and multiple-locus heterozygosity actually increased as a result of the founder event. The weak relationship between a population's heterozygosity and the number and composition of its founders resulted from an increase in the variance of heterozygosity due to drift of allele frequencies. There was little evidence that selection influenced the loss of allozyme variation. When it is not possible to estimate heterozygosity at a large number of polymorphic loci, allozyme surveys attempting to detect founder events and other types of bottlenecks should focus on levels of locus polymorphism and allelic diversity rather than on heterozygosity.     

A new method for identifying informative genetic markers in selectively bred rats

Microsatellite length polymorphisms are useful for the mapping of heritable traits in rats. Over 4000 such microsatellites have been characterized for 48 inbred rat strains and used successfully to map phenotypes that differ between strains. At present, however, it is difficult to use this microsatellite database for mapping phenotypes in selectively bred rats of unknown genotype derived from outbred populations because it is not immediately obvious which markers might differ between strains and be informative. We predicted that markers represented by many alleles among the known inbred rat strains would also be most likely to differ between selectively bred strains derived from outbred populations. Here we describe the development and successful application of a new genotyping tool (HUMMER) that assigns “heterozygosity” (Het) and “uncertainty” (Unc) scores to each microsatellite marker that corresponds to its degree of heterozygosity among the 48 genotyped inbred strains. We tested the efficiency of HUMMER on two rat strains that were selectively bred from an outbred Sprague-Dawley stock for either high or low activity in the forced swim test (SwHi rats and SwLo rats, respectively). We found that the markers with high Het and Unc scores allowed the efficient selection of markers that differed between SwHi and SwLo rats, while markers with low Het and Unc scores typically identified markers that did not differ between strains. Thus, picking markers based on Het and Unc scores is a valuable method for identifying informative microsatellite markers in selectively bred rodent strains derived from outbred populations.     

微卫星标记对WHBE兔封闭群、日本大耳白兔和新西兰兔的遗传分析

目的分析比较大耳白黑眼兔（WHBE兔）封闭群与日本大耳白兔（Jw兔）、新西兰兔（NZW兔）基因组存在的微卫星结构,研究WHBE兔封闭群的微卫星多态性。方法利用21个微卫星位点,通过微卫星分子标记技术对WHBE兔封闭群、Jw兔和NZW兔进行遗传多样性检测和对比。结果根据初步结果,在21对微卫星引物中筛选出扩增产物稳定并且具有多态性的11对引物。WHBE兔封闭群在每个位点上的等位基因数为3～8个不等,11个位点的平均有效等位基因数为2．0402个,平均杂合度为0．4810;Jw兔在每个位点上的等位基因数为2～8个不等,11个位点的平均有效等位基因数为3．6077个,平均杂合度为0．5039;NZW兔在每个位点上的等位基因数为3～9个不等,11个位点的平均有效等位基因数为2．6537个,平均杂合度为0．5334。WHBE兔封闭群在11个微卫星位点上的平均多态信息含量（PIC）为0．6005,多位点累积个体识别率达到100％,多位点累积非父排除概率（CPE）在双亲信息都是未知情况下的为0．9613,而在得知任一亲本信息的情况下,CPE值高达0．9973。在11个微卫星座位中,9个位点上出现了WHBE兔封闭群特有等位基因,其中在Sat2、Sat5、Sat7、Sat12、Sat13、Sat16、S0144和INRACCDDV0003八个位点上WHBE兔封闭群的特有等位基因为一个,在sat8位点上为两个。结论WHBE兔8个位点的平均杂合度、平均有效等位基因数均比JW兔及NZW兔低,说明WHBE兔群体的基因纯合度高于其他两个品系,具有更优的遗传稳定性。9个WHBE兔特有的等位基因可作为区分WHBE兔封闭群和其它两个品系实验兔的分子标记。     

Predictability of heterozygosity scores and polymorphism information content values for rat genetic markers

Construction of a genetic linkage map of the laboratory rat, Rattus norvegicus, establishes the rat as a genetic model. Allele sizes were reported for 432 simple sequence length polymorphisms (SSLPs) genotyped in 12 different substrains belonging to nine different inbred strains of rats. However, these nine strains represent only a fraction of the more than 140 inbred strains available. If allele sizes are not known, alternative indices of markers' polymorphism content can be used, such as heterozygosity (H) and polymorphism information content (PIC). Here, we have determined heterozygosity scores and PIC values for all markers of the rat genetic linkage map, and we evaluate the predictability of the heterozygosity and the PIC values. Correlation analysis between the nine inbred strains reported for the rat map and ten test strains yielded r=0.42 and r=0.44 for heterozygosity and PIC values, respectively. While the correlation of the indices between the two groups of animals is low, these indices do provide a means of predicting whether a genetic marker will be informative in strains where allele sizes are not known.     

Analysis of the enzyme polymorphism in a plaice, Pleuronectes platessa L., population from the north-west coast of Brittany, France

A study of enzyme polymorphism by electrophoretical techniques permitted the genetic structure and the homogeneity degree of the plaice population present in the Abers Wrac'h and Benoit along the north-west coast of Brittany to be determined. Thirteen enzymic systems encoded by 21 loci were analysed and nine of these loci were polymorphic at the 5% level. The comparison of allelic frequencies and of observed heterozygosity h _obs did not show significant differences between these two individual groups. Nei's genetic identity ( I = 0.998) and distance ( D = 0.002) as estimated from allelic frequencies showed that the plaice of the two estuaries are very similar.
Thus, the plaice of A. Wrac'h and of A. Benoit belong to the same population in which the average level of polymorphism per locus P is 47% and the observed average heterozygosity per individual H _obs is 0.163 ± 0.094.     

Isolation and characterization of 16 novel microsatellite loci in two inbred strains of the Chinese hamster (Cricetulus griseus)

The Shanyi inbred A and E strains of the Chinese hamster are widely used in biomedical research, but detailed genetic characterization has been lacking. We developed microsatellite markers that could be used for genetic diversity analysis and linkage map construction. We isolated and characterized 16 novel microsatellite loci from a microsatellite-enriched genomic DNA library. These loci were genotyped in 48 animals from the two strains, and the polymorphic information content was determined. In the Shanyi A and E populations, 14 and 15 loci were found to be polymorphic, respectively, with polymorphic information content ranging from 0.1393 to 0.8082 and from 0.1109 to 0.7397, respectively. A total of 115 alleles were found for the 16 microsatellite loci in the two populations; the mean observed heterozygosity (H(O)) was 0.5191 and 0.4333 for the A and E populations, respectively, indicating marked genetic variation within the two populations. Correspondingly, the F(ST) values ranged from 0.002 to 0.9253, with an overall mean of 0.1935, indicating significant genetic difference between the two strains. The population differentiation levels were substantiated by Nei's genetic distance and full Bayesian analyses computed with STRUCTURE. Despite the genetic diversity and differentiation within and between the two inbred populations, the 48 individuals were correctly allocated into their original populations with high statistical confidence based on these 16 microsatellite loci. These novel microsatellite loci should be useful genetic markers for these two Chinese hamster inbred strains.     

Microsatellite markers for mungbean developed from sequence database

A novel set of microsatellite markers for mungbean [Vigna radiata (L.) Wilczek] was developed from the public sequence database. Seventy-eight primers were designed and evaluated for polymorphism among 22 cultivated accessions. Eight polymorphic loci detected two to three alleles per locus with an average of 2.25. The observed heterozygosity varied from 0.00 to 0.18, while the expected heterozygosity ranged from 0.09 to 0.46. Among them, all eight loci showed significant departuring from Hardy-Weinberg equilibrium, while four pairs of loci displayed significant pairwise linkage disequilibrium values. All eight loci except DMB-SSR1 showed heterozygote deficiency.     

Genetic map of 16 polymorphic markers forming three linkage groups assigned to rat Chromosome 4

Sixteen polymorphic markers, including markers for eight new loci, forming three linkage groups, were assigned to rat Chromosome (Chr) 4 by linkage analysis of the progeny of an F₂ intercross of Fischer (F344/N) and Lewis (LEW/N) inbred rats. One gene, Igk, was mapped by restriction fragment length polymorphism (RFLP) analysis. One marker for Tcrb was identified by the polymorphic insertion of a repetitive LINE element. The remaining 14 markers contained polymorphic simple sequence repeats (SSRs). Ten were identified in genes (Tgfa, Npy, Prss1, Prss2, Aldr1, Iapp, Prp, Eno2, Cacnlla1, and Il6), one was identified in a sequence related to a gene (Egr4l1), and three were identified in anonymous DNA segments. The SSR markers were highly polymorphic in 16 inbred rat strains. These markers expand the genetic map of the rat and should be useful in future genetic studies of inbred rats.     

秦川母牛群体遗传特性的微卫星标记研究

为了从DNA分子水平揭示秦川牛群体遗传多态性和群体遗传结构,寻找可用于秦川牛的微卫星标记,本研究选择了12个普通牛（Bos taurus）微卫星标记检测了90头秦川母牛各微卫星位点的遗传变异及多态性。结果表明,在秦川母牛群体中,12个微卫星位点共检测到了247个等位基因,各位点的等位基因数在13（INRA005）～33个（HEL13）之间,平均每个微卫星位点的等位基因数为21个;总有效等位基因数和平均每个位点平均有效等位基因数（Ne）分别分为142．6229和11．8852。各位点平均基因频率取样方差（V（pij））为2．6036×10^-4。12个微卫星位点平均观察杂合度（Ho）和平均期望杂合度（He）在0．7842（INRA005）～0．9775（BM315）和0．7952（BM315）～0．9446（HEL13）之间。12个位点平均多态信息含量（PIC）在0．7653（INRA005）～0．9420（HEL13）之间,平均为0．8965．12个微卫星位点均属于高度多态位点,这表明秦川母牛群体中所检测各微卫星位点具有丰富的遗传多态性,具备较大的选择潜力。12个微卫星位点的平均固定指数（F）为-0．0076,即各位点杂合子的缺陷度不高,即偏离Hardy—Weinberg平衡的程度不大。     

薇菜SSR分子标记的开发及遗传多样性初步探讨

利用改良FIASCO法(Fast Isolation by AFLP Sequences COntaining repeats)开发出的9对多态性SSR引物评价了薇菜(Osmunda japonica Thunb.)2个野生居群(庐山和恩施)、1个栽培居群(恩施)的遗传多样性和遗传分化水平。结果显示,9个SSR标记在3个薇菜居群中共检测到47个等位基因,每个SSR位点的平均等位基因数为5.222个,观测杂合度和期望杂合度分别为0.000~0.944和0.577~0.834,香农指数为0.962~1.860,表明各SSR位点多态性较高;各居群的平均期望杂合度均大于平均观测杂合度且种内近交系数均为正值,说明3个薇菜居群中都存在非随机交配现象;对各居群的相关遗传多样性参数分析表明,恩施野生居群遗传多样性最高,而其栽培居群最低;庐山野生居群与恩施野生居群间遗传分化系数为0.092,说明两地野生薇菜居群的遗传分化程度较低,AMOVA分析也表明遗传变异主要存在于野生居群内部。     

Genic Heterozygosity and Protein Polymorphism among Local Populations of OENOTHERA BIENNIS

Twenty enzyme loci were studied in 44 Illinois populations of Oenothera biennis; four were polymorphic. Most of the variation was distributed between populations. Fifty-nine percent of the populations had one genotype, 27% two genotypes and the remaining 16% from three to five genotypes; the average was 1.50. The proportion of genetic diversity present in single populations is.38 of that present in the state. Members of single populations were uniformly heterozygous for 1 to 4 loci. The mean heterozygosity per population ranged from 0 to 20%. For Illinois populations collectively, heterozygosity averaged 4.5%. There was much gene frequency heterogeneity between populations. The true standardized genetic variance among populations for alleles at polymorphic loci varied from.40 to.78. Populations from Cook County were much more similar inter se than those downstate, had fewer genotypes and polymorphic loci, and had less heterozygosity than downstate populations. The mean normalized genetic identity among Cook County populations was.987 versus.947 for downstate populations. The mean number of genotypes per population in Cook County was 1.06 versus 2.40 in downstate populations. There was only one polymorphic locus in Cook County, VLP. The genetic structure of Oe. biennis suggests that single populations are colonized by one, or at best a few individuals. Cook County populations are judged to be less variable than downstate populations because the mean age of the populations probably is less than that of those downstate.