Genetic linkage map of the pearl oyster,Pinctada martensii (Dunker)

Genetic linkage maps were constructed with amplified fragment length polymorphism (AFLP) and microsatellite markers for the pearl oyster, Pinctada martensii (Dunker), the main bivalve used for marine pearl production in Asia. Twenty‐four AFLP and 84 microsatellite primer pairs were used for linkage analysis in a full‐sib family with two parents and 78 offspring. Of the 2357 AFLP fragments generated, 394 (16.7%) were polymorphic and segregating. Most (340 or 86.2%) of the markers segregated according to expected Mendelian ratios. Female and male linkage maps were constructed using 230 and 189 markers, including 15 and 10 microsatellites respectively. The female map consisted of 110 markers in 15 linkage groups, covering 1415.9 cM, with an average interval of 14.9 cM. The male map consisted of 98 markers in 16 linkage groups, with a total length of 1323.2 cM and an average interval of 16.1 cM. When unlinked doublets were considered, genome coverages were 78.5% for the female and 73.5% for the male map. Although preliminary, the genetic maps constructed here should be useful for future linkage and quantitative trait loci mapping efforts.     

A first genetic linkage map of bluegill sunfish (<Emphasis Type="Italic">Lepomis macrochirus</Emphasis>) using AFLP markers

Genetic linkage maps were constructed for bluegill sunfish, Lepomis macrochirus, using AFLP in a F₁ inter-population hybrid family based on a double-pseudo testcross strategy. Sixty-four primer combinations produced 4,010 loci, of which 222 maternal loci and 216 paternal loci segregated at a 1:1 Mendelian ratio, respectively. The female and male framework maps consisted of 176 and 177 markers ordered into 31 and 33 genetic linkage groups, spanning 1628.2 and 1525.3 cM, with an average marker spacing of 10.71 and 10.59 cM, respectively. Genome coverage was estimated to be 69.5 and 69.3% for the female and male framework maps, respectively. On the maternal genetic linkage map, the maximum length and marker number of the linkage groups were 122.9 cM and 14, respectively. For the paternal map, the maximum length and marker number of the linkage groups were 345.3 cM and 19, respectively, which were much greater than those on the maternal genetic linkage map. The other genetic linkage map parameters of the paternal genetic linkage map were similar to those in the maternal genetic linkage map. For both the female and male maps, the number of linkage groups was greater than the haploid chromosome number of bluegill (2n = 48), indicating some linkage groups may distribute on the same chromosome. This genetic linkage mapping is the first step toward to the QTL mapping of traits important to cultured breeding in bluegill.     

Application of amplified fragment length polymorphism markers to assess molecular polymorphisms in gynogenetic haploid embryos of turbot (Scophthalmus maximus)

Among the variety of cultured marine species, the turbot Scophthalmus maximus is a fish of growing importance in European aquaculture. In this paper, an advanced application of AFLPs to estimate the genetic diversity of haploid gynogenetic families with the aim of obtaining a preliminary genetic map is presented. Ten EcoRI/TaqI primer combinations were tested in four families comprising diploid mothers and their haploid progenies. The amplified fragment length polymorphism (AFLP) analysis revealed an average of 6.8 polymorphic bands per primer combination and a total number of 88 polymorphisms out of 579 fragments. Among various primer pairs, seven combinations were selected in relation to the quality of profiles and number of polymorphic fragments, to be used in the determination of genetic linkage relationship between AFLP markers within the largest haploid family. Co‐migration of non‐homologous fragments was also investigated in one primer combination adding a fourth selective nucleotide to the three used in the classic TaqI AFLP protocol. Surprisingly, a rate of 38.7% of non‐homologous fragments co‐migrating with monomorphic bands was identified, due to the combined effect of homoplasy and the protocol used. Additional polymorphic markers discovered by this protocol were included in the linkage map. The turbot AFLP linkage map comprises 52 AFLP markers distributed in 12 linkage groups. On the basis of this map, turbot expected total genome length sums up to 1225.6 cM. The results confirm the usefulness of AFLPs in revealing genome segregation in haploid turbot progeny.     

Genetic linkage map of bay scallop, Argopecten irradians irradians (Lamarck 1819)

The bay scallop (Argopecten irradians irradians Lamarck 1819) has become one of the most important aquaculture species in China. Genetic improvement of cultured bay scallop can benefit greatly from a better understanding of its genome. In this study, we developed amplified fragment length polymorphisms (AFLPs) and simple sequence repeat markers from expressed sequence tags (EST‐SSRs) for linkage analysis in bay scallop. Segregation of 390 AFLP and eight SSR markers was analysed in a mapping population of 97 progeny. Of the AFLP markers analysed, 326 segregated in the expected 1:1 Mendelian ratio, while the remaining 74 (or 19.0%) showed significant deviation, with 33 (44.6%) being deficient in heterozygotes (A/a). Among the eight polymorphic EST‐SSR loci, one marker (12.5%) was found skewing from its expected Mendelian ratios. Eighteen per cent of the markers segregating from female parent were distorted compared with 21% of the markers segregating from male parent. The female map included 147 markers in 17 linkage groups (LGs) and covered 1892.4 cM of the genome. In the male map, totally 146 AFLP and SSR markers were grouped in 18 LGs spanning 1937.1 cM. The average inter‐marker spacing in female and male map was 12.9 and 13.3 cM respectively. The AFLP and SSR markers were distributed evenly throughout the genome except for a few large gaps over 20 cM. Although preliminary, the genetic maps presented here provide a starting point for the mapping of the bay scallop genome.     

A first‐generation genetic map of the Japanese scallop Patinopecten yessoensis‐based AFLP and microsatellite markers

We constructed genetic linkage maps of the Japanese scallop Patinopecten yessoensis using AFLP and microsatellite markers. With 32 AFLP primer combinations, a total of 413 markers (209 from the female parent and 204 from the male parent) segregated in a 1:1 ratio, corresponding to DNA polymorphisms which were heterozygous in one parent and null in the other. Among the six microsatellite markers we used, there were four polymorphic loci. Two segregated in the female parent, and the other two segregated in both parents. In the maternal parent, 161 framework markers were mapped in 20 linkage groups, with a total coverage of 2198.8 cM. In the paternal parent, 166 framework markers established a map with 21 linkage groups, spanning a genome length of 2137.6 cM. The AFLP markers on the maps were randomly distributed with an average spacing between markers of 14.7–15.6 cM. The estimated coverage for the framework maps are 77.9% both for the female and the male. These are the first linkage maps for P. yessoensis, which constitute a basis for further genome studies and provide a useful framework for consensus map construction by adding orthologous anchor markers developed in P. yessoensis.     

AFLP-based genetic linkage map of marine shrimp Penaeus (Fenneropenaeus) chinensis

This paper presents the genetic linkage map of the Chinese shrimp Penaeus (Fenneropenaeus) chinensis constructed with 472 AFLP markers. A hundred F₁ progeny from an intercross between a female from the new variety “Yellow Sea No. 1” and wild caught male used for the mapping study. Two separate maps were constructed for each parent. The female linkage map consisted of 197 marker loci forming 35 linkage groups and spanned a total length of 2191.1 cM, with an average marker space of 13.5 cM. The male map consisted of 194 marker loci mapped to 36 linkage groups and spanned a total length of 1737.3 cM, with an average marker spacing of 11.0 cM. The level of segregation distortion observed in this study was 12.2%. The estimated genome length of P. chinensis was 3150.3 cM for the female and 2549.3 cM for the male, respectively. The observed genome coverage was 69.6% for the female and 68.1% for the male map. The linkage maps constructed in this study provide basic information for further linkage studies on Chinese shrimp, and more importantly, the construction of the maps are part of the work of the genetic breeding programs which will be used for growth discovered in the QTL analysis of P. chinensis.     

基于微卫星标记整合长牡蛎遗传图谱

为了提高长牡蛎遗传图谱上的微卫星标记密度,实验采用6个家系图谱间的共有微卫星标记作为锚定标记,构建了长牡蛎的整合图谱.该整合图谱共有161个微卫星标记,覆盖10个连锁群,图谱长度和平均间距分别为615.4和3.8 cM.各连锁群的标记数介于10 ～ 24个之间,连锁群长度为47.3～73.3 cM,是目前密度最高的长牡蛎微卫星图谱.不同作图家系连锁群上的标记分组保持一致,但标记顺序出现差异,可能与长牡蛎自然群体中存在大量的染色体重排现象有关.结果表明,该图谱可以为今后长牡蛎的遗传育种研究提供新的遗传工具.     

Identification of quantitative trait loci for growth-related traits in the Pacific abalone Haliotis discus hannai Ino

The locations and effects of quantitative trait loci (QTL) were estimated for nine characters for growth‐related traits in the Pacific abalone (Haliotis discus hannai Ino) using a randomly amplified polymorphic DNA (RAPD), amplification fragment length polymorphism (AFLP) and SSR genetic linkage map. Twenty‐eight putatively significant QTLs (LOD>2.4) were detected for nine traits (shell length, shell width, total weight, shell weight, weight of soft part, muscle weight, gonad and digestive gland weight, mantle weight and gill weight). The percentage of phenotypic variation explained by a single QTL ranged from 8.0% to 35.9%. The significant correlations (P<0.001) were found among all the growth‐related traits, and Pearson's correlation coefficients were more than 0.81. For the female map, the QTL for growth were concentrated on groups 1 and 4 linkage maps. On the male map, the QTL that influenced growth‐related traits gathered on the groups 1 and 9 linkage maps. Genetic linkage map construction and QTL analysis for growth‐related traits are the basis for the marker‐assisted selection and will eventually improve production and quality of the Pacific abalone.     

Genotyping‐by‐sequencing for construction of a new genetic linkage map and QTL analysis of growth‐related traits in Pacific bluefin tuna

Pacific bluefin tuna (Thunnus orientalis) has high market value, but its wild populations have decreased in recent years. The broodstock of Pacific bluefin tuna that were hatched artificially and reared under aquaculture conditions is beginning to be used for production. The creation of broodstock with commercially valuable traits, such as rapid growth, is therefore of great interest. Genetic linkage map‐based identification of markers associated with quantitative trait loci (QTLs) facilitates marker‐assisted selection (MAS) breeding and allows efficient genetic improvement of broodstock. Single nucleotide polymorphism (SNP)‐based genetic linkage map construction using the genotyping‐by‐sequencing method can expand the number of mapped markers and help identify growth‐related QTLs. In this study, we constructed sex‐specific maps for 24 linkage groups consisting of 677 SNP and 651 microsatellite markers. The total lengths of 93 progenies in the mapping population followed normal distribution, with an average length of 9.4 mm. We performed composite interval mapping in the mapping population. QTL analysis revealed one significant QTL in LG10 on the female linkage map. The genetic linkage map—the second such map generated for Pacific bluefin tuna—and the growth‐related QTLs detected in this study will be useful for tuna aquaculture MAS programs.     

Identification of quantitative trait loci for growth‐related traits in the swimming crab Portunus trituberculatus

The swimming crab (Portunus trituberculatus) is an economically important species in Asian aquaculture. Implementing growth‐related traits of P. trituberculatus into genetic breeding programmes is an ongoing effort. We used a previously published genetic linkage map of P. trituberculatus, containing 55 simple sequence repeat (SSR) markers and 172 amplified fragment length polymorphism (AFLP) techniques to map quantitative trait loci (QTL) for growth‐related traits in a single full sibling F₂ family. Ten growth‐related traits were measured for QTL mapping. Composite interval mapping identified 9 QTL on the female map and 16 on the male map. Individual QTL with additive effects explained 11–38% of the phenotypic variance for various traits using the female parent's map, and from 1% to 21% using the male parent's map. Two QTL explaining a large percentage of variation in body weight were detected on chromosome 17 on the female map, and on chromosome 16 on the male map, and contributed 38% and 18% of the phenotypic variance respectively. This is the first study to report the detection and positioning of major QTL affecting growth in a true crab species (Brachyura). The mapping of growth‐related QTL in this study raises the possibility of improving the growth of P. trituberculatus through marker‐assisted selection.     

A genetic linkage map of the Japanese flounder, Paralichthys olivaceus

We report the first genetic linkage map of the Japanese flounder (Paralichthys olivaceus) constructed with 111 microsatellite markers and 352 AFLP fragments. The parental male linkage map consisted of 25 linkage groups while the female map consisted of 27 groups, with an average resolution of 8 and 6.6 cM, respectively. We have identified linkage among 96% of the markers and the total map length was estimated to be around 1000–1200 cM. This study reports the first low-density linkage map for the Japanese flounder and describes differences in sex recombination. Recombination rates were higher in male flounder compared to the female (7.4:1), a rare condition among vertebrates. This map is a starting point for the mapping of single loci and quantitative traits in flatfish species.     

牙鲆遗传作图及生长性状QTL定位

采用牙鲆日本群体和韩国群体杂交的92个F₁个体作为分离群体, 利用微卫星标记和Joinmap 4.0作图软件构建了牙鲆遗传连锁图谱。共有221个SSR标记用于连锁图谱构建, 雌性图谱中, 共178个微卫星标记定位到22个连锁群上, 观测总长度为(G _oa )599.0 cM, 覆盖率(C _oa )达76.27%。雄性图谱中, 共194个微卫星标记定位到23个连锁群上, G _oa 为693.4 cM, C _oa 为78.82%。对全长、体质量、体高3组数据进行主成分分析处理, 得到可解释3个性状的89.6%特征的一组数据, 命名为牙鲆生长性状GT。用WinQTLCart 2.5软件的复合区间作图,在已构建的遗传连锁图谱上对牙鲆生长性状GT进行QTL定位, 取LOD经验值2.5为QTL存在的阈值; 对微卫星标记进行性状—标记之间的回归分析。本研究共定位3个与牙鲆生长性状GT相关的QTLs, qGT-f4 qGT-m20 qGT-f20,可解释表型变异率分别为27.60%, 13.74%, 10.27%。在性状—标记之间的回归分析中, 得到22个与生长性状GT相关(P<0.05)的微卫星标记, 单个标记可解释表型变异率介于3.70%~10.42%, 其中6个微卫星标记scaffold558_51720、scaffold558_26183、scaffold903_69232、scaffold485_47120 、scaffold1262_77386、scaffold809_65154与生长性状GT之间呈极显著相关(P><0.01), 可解释表型变异率分别为10.42%、7.31%、10.07%、10.07%、8.39%和11.26%。     

Quantitative trait locus mapping of growth‐related traits in inter‐specific F1 hybrid grouper (Epinephelus fuscoguttatus × E. lanceolatus) in a tropical climate

Growth‐related traits are the main target of genetic breeding programmes in grouper aquaculture. We constructed genetic linkage maps for tiger grouper (Epinephelus fuscoguttatus) and giant grouper (E. lanceolatus) using 399 simple sequence repeat markers and performed a quantitative trait locus (QTL) analysis to identify the genomic regions responsible for growth‐related traits in F₁ hybrid grouper (E. fuscoguttatus × E. lanceolatus). The tiger grouper (female) linkage map contained 330 markers assigned to 24 linkage groups (LGs) and spanned 1,202.0 cM. The giant grouper (male) linkage map contained 231 markers distributed in 24 LGs and spanned 953.7 cM. Six QTLs affecting growth‐related traits with 5% genome‐wide significance were detected on different LGs. Four QTLs were identified for total length and body weight on Efu_LG8, 10, 13 and 19 on the tiger grouper map, which explained 6.6%–12.0% of the phenotypic variance. An epistatic QTL with a reciprocal association was observed between Efu_LG8 and 10. Two QTLs were identified for body weight on Ela_LG3 and 10 on the giant grouper map, which explained 6.9% of the phenotypic variance. Two‐way analysis of variance indicated that the QTL on Efu_LG13 interacts with the QTLs on Ela_LG3 and 10 with large effects on body weight. Furthermore, these six QTLs showed different features among the winter, summer and rainy seasons, suggesting that environmental factors and fish age affected these QTLs. These findings will be useful to understand the genetic structure of growth and conduct genetic breeding in grouper species.     

Microsatellite–centromere mapping in bighead carp ( Aristichthys nobilis) using gynogenetic diploid families

Gene–centromere mapping using half‐tetrad analysis is a powerful tool for understanding chromosomal behaviour and determining the position of centromeres in relation to genes or markers in fish. In this study, eight gynogenetic diploid families induced by inhibiting of the second meiotic division were genotyped at 66 microsatellite loci for microsatellite–centromere (M‐C) mapping in bighead carp ( Aristichthys nobilis). The absence of paternal alleles verified the success of gynogenetic development in all gynogenetic families. All loci were consistent with Mendelian segregation in control families. The estimated recombination frequency (y) ranged from 0.057 to 0.875 with an average of 0.477 ± 0.222. Seventeen loci (25.76%) showed high M‐C recombination frequencies of over 0.667. M‐C distances ranged from 2.85 to 43.75 cM under the assumption of complete interference. Thus, these loci are distributed from the centromeres to the telomeres of their respective chromosomes, while mainly in the intermediate region. Information on centromere mapping could serve as a starting point to consolidate the genetic linkage groups in bighead carp.     

应用SSR和SRAP标记构建青虾遗传连锁图谱

采用SSR和SRAP标记结合拟测交策略构建青虾(Macrobrachium nipponense)遗传连锁图谱。共有175个标记(含27个SSR、148个SRAP标记)分布在53个连锁群上。每个连锁群含2~8个标记,其中不少于3个标记的连锁群有35个,连锁对18个,平均每个连锁群的标记数为3.3个;连锁群长度在6.7~91.2 cM之间,相邻标记间最大间隔为49.0 cM,最小为1.4 cM,平均间隔为13.1 cM。青虾框架图谱长度为997.2 cM,图谱观察总长度为2 270.5cM,根据估算,青虾遗传连锁图谱预期长度为4 380.6 cM,图谱的覆盖率为51.83%。本研究构建了青虾遗传连锁图谱,该图谱也是淡水虾蟹类第一张遗传连锁图谱,可为青虾QTL定位、基因克隆、遗传选育等提供指导,并为进一步构建高密度的青虾遗传连锁图谱奠定了基础。     

Genetic information of three pure lines of Porphyra yezoensis (Bangiales, Rhodophyta) obtained by AFLP analysis

Porphyra (Bangiales, Rhodophyta), which includes several valuable marine crops, has recently received great interest as a model plant for fundamental and applied studies in marine sciences. Amplified fragment length polymorphisms (AFLPs) are a robust and efficient means for genetic mapping, linkage analysis of genetic characters for breeding and population studies in land plant genomes. To examine whether AFLPs are applicable as genetic markers in the present study, we detected AFLP markers with three pure lines in order to promote genetic analysis in Porphyra yezoensis . The following five sets of AFLP primer pairs (E-AA, M-CAA) (E-AA, M-CAC) (E-AA, M-CAG) (E-AA, M-CAT) (E-AA, M-CTA) were tested with template DNAs from three pure lines and they showed a total of 227 bands. This suggests that AFLP markers are promising tools for genetic analysis in Porphyra .     

黄颡鱼遗传图谱构建及生长相关性状的QTL定位

以野生(♂)和人工养殖(♀)黄颡鱼杂交的100个F1个体为作图群体,用SSR、SRAP和TRAP3种DNA分子标记技术构建黄颡鱼的遗传连锁图谱。图谱整合了13个SSR标记,89个SRAP标记,26个TRAP标记。其中雌性框架图谱包括16个连锁群,图谱的长度为585.5cM;雄性框架图谱包括15个连锁群,图谱的长度为752.3cM;共享框架图谱包括5个连锁群,图谱的长度为231.3cM。用该连锁图谱对黄颡鱼的5个生长相关性状进行QTL扫描,在雌性图谱上检测到1个头宽的QTL,定位于第七连锁群上,LOD值为3.2,可解释的表型变异为13%。在雄性图谱上分别检测到1个体高和体长的QTL,均定位于第一连锁群上。体高QTL的LOD值为2.4,可解释的表型变异为12%。全长QTL的LOD值为2.1,可解释的表型变异为11%。3个QTL均可用于黄颡鱼的生长性状的标记辅助育种。     

鲤鱼的遗传连锁图谱（初报）

建立了鲤鱼的遗传连锁图谱。图谱有ＲＡＰＤ分子标记５６个，鲤鱼的ＳＳＬＰ标记２６个，鲤鱼ＳＳＬＰ标记１９个，斑马鱼的ＳＳＬＰ分子标记７０个，鲤鱼基因标记９１个，共有标记２６２个；图谱有５０个连锁组，连锁图给出鲤鱼的基因组大小在５７８９ＣＭ左右。     

A genome scan of a four-way tilapia cross supports the existence of a quantitative trait locus for cold tolerance on linkage group 23

A genome scan, searching for quantitative trait loci (QTL) for the traits cold tolerance and body weight in tilapia, was performed on a cross between a (Oreochromis niloticus×Sarotherodon galilaeus) male and a (O. mossambicus×O. aureus) female. Fifty‐four microsatellites and 23 amplified fragment length polymorphism (AFLP) primer combinations were genotyped and tested for marker–trait associations. Sex‐specific linkage maps were constructed from this data. Twenty‐three point‐wise significant marker–trait associations were found in the genome scan, and putative QTL were subsequently tested in another (On×Sg) × (Om×Oa) family. None of the putative QTL from the first experiment were significant in the second experiment. However, one microsatellite, UNH130, found to be associated to weight in the first experiment, was found to be strongly associated to cold tolerance in the second experiment. Since QTL for cold tolerance and body weight were recently found on the linkage group containing UNH130 (linkage group 23) in another study, this linkage group was investigated more closely using interval mapping. The results provide indications, but not conclusive evidence, of a QTL for cold tolerance on linkage group 23.     

基于遗传连锁图谱筛选虾夷扇贝性别相关AFLP分子标记的方法

基于近年来他人构建的遗传连锁图谱,对2011–2013年2–5月期间采集的雌性、雄性及雌雄同体虾夷扇贝进行性别相关AFLP分子标记筛选。从雄性和雌性遗传连锁图谱上选取合计18对引物组合,经3批次试验,有8对引物扩增出性别相关的条带,其中Eb Mc、Ej Mf和Ei Mk重复性好,有4对引物扩增出雌雄同体特异性条带,5对引物扩增出雌性特异性条带。结果表明,基于高密度遗传连锁图谱筛选性别相关分子标记的方法简单可行,雌雄同体的基因组DNA与雌性和雄性存在显著差异,可能是一个单独的群体。