甘蔗SSR和AFLP分子遗传连锁图谱构建

采用甘蔗商业品种Co419与野生种割手密Y75/1/2杂交,获得269个单株,组成F1群体,用F102/356与商业品种ROC25回交获得266个单株,组成BC1群体。利用筛选的多态性条带丰富的36对SSR引物和12对AFLP引物,对两个群体进行PCR扩增和分子遗传连锁分析,构建甘蔗分子遗传连锁图谱。用F1群体获得630个分离标记,经χ2检测,298个标记为单双剂量标记,占总标记数的47%;用BC1群体获得571个分离标记,有264个标记为单双剂量标记,占总标记数的46%;4个亲本获得单双剂量标记的数量依次为Co41902/356Y75/1/2ROC25。在LOD≥5.0,相邻标记遗传距离≤40cM的条件下,F1群体有134个单双剂量标记被纳入55个连锁群,其中39个连锁群归属8个同源组,16个未列入,总遗传距离为1458.3cM,标记间平均图距为10.9cM;BC1群体有133个单双剂量标记被纳入47个连锁群,其中34个连锁群归属于8个同源组,13个连锁群未列入,总遗传距离为1059.6cM,标记间平均图距为8.0cM。从4个亲本单双剂量标记进入的连锁群数来看,Co419最多,归入34个连锁群,其次为Y75/1/2,归入20个连锁群,第3为02/356和ROC25,归入19个连锁群。研究结果表明,从单双剂量标记比例、形成连锁群数量、总遗传距离来看,F1群体构图质量要优于BC1群体。     

山农01-35×藁城9411重组自交系遗传图谱构建及粒重QTL分析

为利用分子遗传图谱进行小麦产量数量性状位点定位分析,以大粒高产小麦品系山农01-35和强筋小麦藁城9411为亲本,衍生了含182个家系的重组自交系(RIL)F₈群体,用442个DArT标记、59个SSR标记和1个TaGW2-CAPS标记,构建了包括29个连锁群的分子遗传图谱,总遗传长度为4 084.5 cM,标记间平均距离为8.13 cM。定位了54个新标记位点,包括44个DArT和10个SSR标记,分布于除4D、6B、7B外的其他18条染色体上。用该分子遗传图谱和4个环境粒重表型值,共检测到7个影响粒重的加性QTL,分别位于1B、4B、5B、6A染色体,其中QGW4B-7、QGW5B-20和QGW6A-29在单环境分别定位和4个环境共同定位两种方法中均能检测到。QGW4B-7、QGW5B-12和QGW6A-29对粒重的贡献率均超过10%,为主效QTL。本研究结果可为小麦高产优质性状的QTL分析及分子标记辅助选择提供参考。     

利用2个F_2群体整合中国豌豆高密度SSR遗传连锁图谱

豌豆(Pisum sativum L.)是一种重要的食用豆类作物,在全世界范围内广泛种植,既可作为人类食物,也可作为牲畜饲料。用SSR标记构建的遗传连锁图谱在豌豆和其他作物的标记辅助育种中发挥着重要的作用。尽管对豌豆遗传连锁作图的研究已有悠久历史,但公众可获得且可转移的SSR标记以及基于遗传独特的中国豌豆种质的高密度遗传连锁图谱仍然有限。为了获得更多可转移的SSR标记和中国豌豆的高密度遗传连锁图谱,本研究首先从自主开发和文献获取的12,491个全基因组SSR标记中筛选了617个多态性SSR标记,并用于G0003973×G0005527 F_2群体遗传连锁图谱的加密。加密后的图谱全长扩展到5330.6 cM,包含603个SSR标记,标记平均间距离8.8 cM,相比之前的图谱有明显改善。基于上述结果,我们又筛选了119个具有多态性的SSR标记,用于构建大样本W6-22600×W6-15174 F_2群体的遗传连锁图谱,新图谱累积长度为1127.1 cM,包含118个SSR标记,装配在7条连锁群上。最后,将来自以上2个遗传图谱的数据进行整合,得到了一张覆盖范围6592.6 cM的整合图谱,包含668个SSR标记,由509个基因组SSR、134个EST-SSR和25个锚定标记组成,分布在7条连锁群上。这些SSR标记和遗传连锁图谱将为豌豆的遗传研究和标记辅助育种提供有力工具。     

一张含有315个SSR和40个AFLP标记的大豆分子遗传图的整合

本研究是基于“锚定SSR标记”作图策略,利用2个F2群体,选用592对SSR引物,对宛煜嵩等利用重组自交系群体Jinf构建的含有227个SSR标记的图谱的基础上进行整合。整合后的大豆分子遗传图包含315个SSR标记和40个AFLP标记,总图距为1951.7cM,相邻标记间的平均图距为5.48cM。整合后的遗传连锁图归属20个连锁群对应于大豆20条染色体,连锁群长度范围从40.8cM到184.4cM,标记数范围从11到41个。整合后的图谱新增加了87个SSR标记,其中A2、C1、C2、D1b和G连锁群有较多的标记增加。整合后的大豆分子遗传图谱中的标记顺序比原图谱与“公共图谱”有更好的线性符合度。本文还进一步对两种类型的作图群体的配合和不同作图软件的选用等问题进行了比较和深入的讨论。     

Genetic linkage map of cassava (Manihot esculenta Crantz) based on AFLP and SSR markers

To generate a genetic linkage map of cassava ( Manihot esculenta Crantz), 58 F₁ progenies from a cross between Rayong 90 (female) and Rayong 5 (male) were examined in amplification fragment length polymorphism (AFLP) and simple sequence repeat (SSR) analyses. A total of 469 polymorphic markers consisting of 378 AFLPs generated from 76 primer combinations and 91 SSRs were identified. These markers were analyzed using the joinmap ^® 3.0 package program to construct a genetic linkage map. A total of 33 linkage groups of a common map were constructed from 119 AFLPs and 18 SSRs, spanning 1095 cM with an average of 7.99 cM between markers. The genetic linkage map generated in this study will be useful for genetic studies in cassava particularly for the identification of genetic markers linked to traits of interest, although the complex cassava genome suggests that maybe a long term objective.     

A genetic linkage map of <Emphasis Type="Italic">Populus adenopoda</Emphasis> Maxim. × <Emphasis Type="Italic">P. alba</Emphasis> L. hybrid based on SSR and SRAP markers

Populus adenopoda Maxim. and P. alba L. [section Populus (aspen), genus Populus] are two tree species of ecological and economic value. To date, no high-density genetic maps are available for these two species. In this study, 1100 interspecific hybrids were obtained by controlled crossing and embryo culture. Simple sequence repeat (SSR) and sequence-related amplified polymorphisms (SRAP) were used to genotype 189 F₁ individuals. The genetic linkage map of P. adenopoda × P. alba generated from this study includes 212 markers (192 SSRs and 50 SRAPs) and consists of 26 linkage groups spanning 2178.5 cM, with an average distance of 11.7 cM between markers. This is the first SSR- and SRAP-containing genetic linkage map for aspen. The SSRs on the map will serve both as bridges for comparison with the poplar maps published to date and as a direct link to the Populus genomic sequence. Future studies focusing on the data presented here should enhance the density and precision of the map for identifying and localizing quantitative trait loci and promote genomic research on the genus.     

Development of genomic and EST-SSR markers in radish (Raphanus sativus L.)

Radish (Raphanus sativus L.) belongs to Brassicaceae family and is a close relative of Brassica. This species shows a wide morphological diversity, and is an important vegetable especially in Asia. However, molecular research of radish is behind compared to that of Brassica. For example, reports on SSR (simple sequence repeat) markers are limited. Here, we designed 417 radish SSR markers from SSR-enriched genomic libraries and the cDNA data. Of the 256 SSR markers succeeded in PCR, 130 showed clear polymorphisms between two radish lines; a rat-tail radish and a Japanese cultivar, ‘Harufuku’. As a test case for evaluation of the present SSRs, we conducted two studies. First, we selected 16 SSRs to calculate polymorphism information contents (PICs) using 16 radish cultivars and four other Brassicaceae species. These markers detected 3–15 alleles (average = 9.6). PIC values ranged from 0.54 to 0.92 (average = 0.78). Second, part of the present SSRs were tested for mapping using our previously-examined mapping population. The map spanned 672.7 cM with nine linkage groups (LGs). The 21 radish SSR markers were distributed throughout the LGs. The SSR markers developed here would be informative and useful for genetic analysis in radish and its related species.     

Construction of integrated linkage map of a recombinant inbred line population of white lupin (Lupinus albus L.)

We report the development of a Diversity Arrays Technology (DArT) marker panel and its utilisation in the development of an integrated genetic linkage map of white lupin (Lupinus albus L.) using an F₈ recombinant inbred line population derived from Kiev Mutant/{"type":"entrez-protein","attrs":{"text":"P27174","term_id":"14195583","term_text":"P27174"}}P27174. One hundred and thirty-six DArT markers were merged into the first genetic linkage map composed of 220 amplified fragment length polymorphisms (AFLPs) and 105 genic markers. The integrated map consists of 38 linkage groups of 441 markers and spans a total length of 2,169 cM, with an average interval size of 4.6 cM. The DArT markers exhibited good genome coverage and were associated with previously identified genic and AFLP markers linked with quantitative trait loci for anthracnose resistance, flowering time and alkaloid content. The improved genetic linkage map of white lupin will aid in the identification of markers for traits of interest and future syntenic studies.     

The first genetic maps of cashew (Anacardium occidentale L.)

Cashew (Anacardium occidentale) is a widespread tropical tree crop that is grown primarily for its nuts and has a global production of over 2 million Mt. In spite of its economic importance to many countries, however, no linkage map containing STS anchor sites has yet been produced for this species. This is largely attributable to a prolonged juvenile phase of the tree (limiting mapping to F₁ progenies) and difficulty in effecting sufficient hand-pollinations to create mapping populations of effective size. Here, we produce an F₁ mapping population of 85 individuals from a cross between CP 1001 (dwarf commercial clone) and CP 96 (giant genotype), and use it to generate two linkage genetic maps comprising of 205 genetic markers (194 AFLP and 11 SSR markers). The female map (CP 1001) contains 122 markers over 19 linkage groups and the male map (CP 96) comprises 120 markers assembled over 23 linkage groups. The total map distance of the female map is 1050.7 cM representing around 68% genome coverage, whereas the male map spans 944.7 cM (64% coverage). The average map distance between markers is 8.6 cM in the female map and 7.9 cM in the male map. Homology between the two maps was established between 13 linkage groups of the female map and 14 of the male map using 46 bridging markers that include 11 SSR markers. These maps represent a platform from which to identify loci controlling economically important traits in this crop.     

Genetic mapping and QTL analysis for sugar yield-related traits in sugarcane

Genetic improvement of sugar content in sugarcane would benefit from the availability of sufficient DNA markers and a genetic map. Genetic linkage maps were constructed to identify quantitative trait loci (QTLs) for seedling brix (SB), brix (B), sucrose percent in juice (SUC), stalk number (SN), stalk length (SL), stalk diameter (SD), internodes (INT), number of green leaves (NGL), at three crop cycles across seven environments in a segregating population with 207 individuals derived from a bi-parental cross of sugarcane elite cultivars. Linkage analysis led to the construction of eight linkage groups (LGs) for Co86011 and sixteen LGs for CoH70. The combined length of the two linkage maps was 2606.77 cM distributed over 24 LGs. 31 QTLs were identified: 2 for SB, 7 for B, 6 for SUC, 4 for SN, 1 for SL, 3 for SD, 6 for INT and 2 for NGL at LOD scores ranging from 2.69 to 4.75. 7 QTLs (22 %) had stable effect across crop year and locations. Markers from parents were found to be associated with both positive and negative effect on all of the traits analyzed. The most important QTLs intervals identified in this study using single-dose marker, were qB2, qSUC2, qINT2 and qB2, qSUC2, qSL2, qINT2 located between SSR markers UGSM31₅₄₈ and UGSM31₆₄₉. These QTLs could be put into use in marker assisted breeding.     

The first genetic linkage map of crape myrtle (Lagerstroemia) based on amplification fragment length polymorphisms and simple sequence repeats markers

Lagerstroemia (crape myrtle) are famous ornamental plants with large pyramidal racemes, long flower duration and diverse colours. Genetic maps provide an important genomic resource of basic and applied significance. A genetic linkage map was developed by genotyping 192 F₁ progeny from a cross between L. caudata (female) and L. indica (‘Xiang Xue Yun’) (male) with a combination of amplification fragment length polymorphisms (AFLP) and simple sequence repeats (SSR) markers in a double pseudo‐testcross mapping strategy. A total of 330 polymorphic loci consisting of 284 AFLPs and 46 SSRs showing Mendelian segregation were generated from 383 AFLP primer combinations and 150 SSR primers. The data were analysed using JoinMap 4.0 (evaluation version) to construct the linkage map. The map consisted of 20 linkage groups of 173 loci (160 AFLPs and 13 SSRs) covering 1162.1 cM with a mean distance of 10.69 cM between adjacent markers. The 20 linkage groups contained 2–49 loci and ranged in length from 7.38 to 163.57 cM. This map will serve as a framework for mapping QTLs and provide reference information for future molecular breeding work.     

Construction of a high-density reference linkage map of tea (Camellia sinensis)

A few linkage maps of tea have been constructed using pseudo-testcross theory based on dominant marker systems. However, dominant markers are not suitable as landmark markers across a wide range of materials. Therefore, we developed co-dominant SSR markers from genomic DNA and ESTs and constructed a reference map using these co-dominant markers as landmarks. A population of 54 F₁ clones derived from reciprocal crosses between ‘Sayamakaori’ and ‘Kana-Ck17’ was used for the linkage analysis. Maps of both parents were constructed from the F₁ population that was taken for BC₁ population. The order of most of the dominant markers in the parental maps was consistent. We constructed a core map by merging the linkage data for markers that detected polymorphisms in both parents. The core map contains 15 linkage groups, which corresponds to the basic chromosome number of tea. The total length of the core map is 1218 cM. Here, we present the reference map as a central core map sandwiched between the parental maps for each linkage group; the combined maps contain 441 SSRs, 7 CAPS, 2 STS and 674 RAPDs. This newly constructed linkage map can be used as a basic reference linkage map of tea.     

Construction of a linkage map for quantitative trait loci associated with economically important traits in creeping bentgrass (Agrostis stolonifera L.)

Creeping bentgrass (Agrostis stolonifera L.) is the most widely cultivated and high-value turfgrass species. Genetic linkage maps of creeping bentgrass were constructed for quantitative trait loci (QTL) analysis of gray snow mold (Typhula incarnata) resistance, recovery and leaf width. A segregating population of 188 pseudo-F₂ progeny was developed by two-way pseudo-testcross mapping strategy. Amplified fragment length polymorphism, new developed Agrostis specific expressed sequence tag-single sequence repeat (SSR), random amplified polymorphic DNA and genomic SSR markers corresponding to DNA polymorphisms heterozygous in one parent and null in the other, were scored and placed on two separate genetic linkage maps, representing each parent. In the male parent map, 100 markers were mapped to 14 linkage groups covering a total length of 793?cM with an average interval of 8.2?cM. In the female parent map, 146 markers were clustered in another 14 linkage groups spanning 805?cM with an average distance of 5.9?cM between adjacent markers. We identified three putative QTL for leaf width and one QTL for snow mold disease resistance. The construction of a linkage map and QTL analysis are expected to facilitate the development of disease resistant creeping bentgrass cultivars by using molecular marker-assisted selection.     

A high-density integrative linkage map for <Emphasis Type="Italic">Gossypium hirsutum</Emphasis>

Sequence-related amplified polymorphism (SRAP) combined with SSRs, RAPDs, and RGAPs was used to construct a high density genetic map for a F₂ population derived from the cross DH962 (G. hirsutum accession) × Jimian5 (G. hirsutum cultivar). A total of 4,096 SRAP primer combinations, 6310 SSRs, 600 RAPDs, and 10 RGAPs produced 331, 156, 17 and 2 polymorphic loci, respectively. Among the 506 loci obtained, 471 loci (309 SRAPs, 144 SSRs, 16 RAPDs and 2 RGAPs) were assigned to 51 linkage groups. Of these, 29 linkage groups were assigned to corresponding chromosomes by SSR markers with known chromosome locations. The map covered 3070.2 cM with a mean density of 6.5 cM per locus. The segregation distortion in this population was 9.49%, and these distorted loci tend to cluster at the end of linkage groups or in minor clusters on linkage groups. The majority of SRAPs in this map provided an effective tool for map construction in G. hirsutum despite of its low polymorphism. This high-density linkage map will be useful for further genetic studies in Upland cotton, including mapping of loci controlling quantitative traits, and comparative and integrative analysis with other interspecific and intraspecific linkage maps in cotton.     

小麦分子遗传图谱的加密

高密度的分子标记遗传图谱是QTL定位、图位克隆和分子标记辅助选择等研究的基础。以小麦品种“京花1号/小白冬麦”的双单倍体(DH)群体和“农大015/复壮30”的重组自交系(RIL)群体为作图群体,选用在DH群体双亲间的339个多态性标记和在RIL群体双亲间的343个多态性标记分析作图群体各个株系的基因型,对本中心近年开发的SCAR、EST-SSR标记以及他人开发的SSR、EST-SSR标记进行了染色体定位,并利用连锁分析软件Joinmap 4.0将2个作图群体的结果整合,最终构建了10个连锁群,将217个SSR、EST-SSR和SCAR位点定位在9条染色体上,进一步提高了小麦遗传图谱的密度。     

Genetic mapping of AFLP markers in Japanese bunching onion (<Emphasis Type="Italic">Allium fistulosum</Emphasis>)

Summary The first genetic linkage map of Japanese bunching onion (Allium fistulosum) based primarily on AFLP markers was constructed using reciprocally backcrossed progenies. They were 120 plants each of (P₁)BC₁ and (P₂)BC₁ populations derived from a cross between single plants of two inbred lines: D1s-15s-22 (P₁) and J1s-14s-20 (P₂). Based on the (P₂)BC₁ population, a linkage map of P₁ was constructed. It comprises 164 markers – 149 amplified fragment length polymorphisms (AFLPs), 2 cleaved amplified polymorphic sequences (CAPSs), and 12 simple sequence repeats (SSRs) from Japanese bunching onion, and 1 SSR from bulb onion (A. cepa) – on 15 linkage groups covering 947 centiMorgans (cM). The linkage map of P₂ was constructed with the (P₁)BC₁ population and composed of 120 loci – 105 AFLPs, 1 CAPS, and 13 SSRs developed from Japanese bunching onion and 1 SSR from bulb onion – on 14 linkage groups covering 775 cM. Both maps were not saturated but were considered to cover the majority of the genome. Nine linkage groups in P₂ map were connected with their counterparts in P₁ map using co-dominant anchor markers, 13 SSRs and 1 CAPS.     

Analysis of QTL for yield-related traits in cassava using an F<Subscript>1</Subscript> population from non-inbred parents

A cassava F₁ population raised from the cross SC6 × Mianbao was used to construct a genetic linkage map. The map incorporated 200 polymorphic amplified fragment length polymorphism, sequence-related amplified polymorphism, simple sequence repeat (SSR), and expressed sequence tag (EST)–SSR markers which fit a 1:1 segregation ratio. It comprised 20 linkage groups (LGs) and spanned a genetic distance of 1645.1 cM with an average marker interval of 8.2 cM. Fifty-seven repeatedly detected QTLs (rd-QTLs) for three phenotypic traits (fresh root yield, root dry matter content, and root starch content) were identified in the F₁ population in four trials of year 2003, 2004, 2005, and 2008 by inclusive composite interval mapping. Among the 57 rd-QTLs, 25 rd-QTLs were linked to SSR/EST–SSR markers, which will help to facilitate marker-assisted selective breeding in cassava, and 15 marker intervals on ten LGs showed pleiotropic effects.     

SSR作为锚定标记构建白菜&#215;芜菁分子遗传图谱

为了将已有的大白菜分子遗传图谱和已发表的A基因组参考图谱对应起来,本研究利用国际上发表的大白菜和甘蓝型油菜A基因组特异SSR标记作为锚定标记,重新构建了白菜×芜菁分子遗传图谱。利用双亲和F1对326个SSR标记进行了筛选,共获得86个多态性分子标记。在此基础上整合了已有的400个标记,最终构建了一张由10个连锁群组成,包含了347个标记的分子连锁图谱,图谱总长度为1008.7cM,标记间的平均图距为2.91cM。此图谱上包含了已在A基因组参考图谱上定位的56个SSR标记,分布于10个连锁群上,从而将各个连锁群与参考图谱的连锁群对应起来。每个连锁群上的标记数在25～58个之间,连锁群长度在60.6～177.0cM范围内,平均图距在1.33～4.92cM之间。该图谱为白菜重要性状的遗传定位奠定了良好基础。     

大豆异地衍生重组自交系群体遗传图谱的构建及比较

以Peking×7605组合分别在济南和南京衍生大豆重组自交系群体JN(RN)P7和NJ(RN)P7,利用145个在亲本之间表现多态的SSR引物和1个形态学标记BSC (黑色种皮性状)及JoinMap 4.0软件构建了2张分别含27个和25个连锁群的大豆遗传图谱,其总长度分别为1 574.80 cM和1 682.50 cM,标记间平均距离分别是13.58 cM和15.72 cM,连锁群长度范围分别为17.30~127.40 cM和20.10~137.50 cM。所构建的两张图谱均与“公共图谱”对应性较好。两图谱间整体上较为一致,但也存在诸多不同点。表明原本具有相同遗传背景的杂交后代,在不同生态环境选择压力下形成的重组自交系群体间遗传结构存在真实差异。