小麦旗叶早衰性状的QTL定位

为小麦旗叶早衰性状的精细定位和基因克隆奠定基础,该试验以普通小麦(Triticum aestivum L.)‘宁春4号’和‘宁春27号’杂交得到的128个F10代RIL群体为研究材料,利用307对多态性SSR标记对小麦旗叶早衰性状进行了QTL定位,并通过构建整合图谱的方法进行了标记加密。结果表明,共检测到1个控制旗叶早衰性状的加性QTL,位于2A染色体长臂的gwm526和gwm382标记区间内,可解释49.88%的表型变异。经遗传图谱整合后发现,gwm526和gwm382标记之间存在124个SNP标记。     

大豆叶绿素含量QTL定位及候选基因预测

叶绿素是调节光合作用的关键色素,对籽粒形成有着重要作用。本研究以美国半矮秆大豆Charleston为母本,东北地区主栽品种东农594为父本杂交衍生的147个重组自交系群体为材料,基于经SLAF测序获得的大豆高密度遗传图谱,利用复合区间作图法(CIM)、多重区间作图法(MIM)和完备区间作图法(ICIM)对大豆叶绿素含量进行QTL联合定位分析,并结合大豆基因组基因注释信息对QTL区段内的候选基因进行预测。利用CIM算法定位出2个QTL,表型遗传贡献率分别为6%和9.3%。利用MIM算法定位到了1个QTL,表型遗传贡献率为8.1%。利用ICIM算法定位到了1个QTL,表型遗传贡献率为7.76%。其中qchl-G-1被CIM和MIM两种算法同时检测到。在上述3个QTL区段内共含有151个基因,根据大豆基因组基因注释信息,筛选到了3个与叶绿素相关的候选基因,这些结果为叶绿素含量的遗传剖析和标记辅助育种提供理论基础,有利于分子辅助育种的发展。     

基于染色体片段置换系的野生稻粒宽QTL qGW8的精细定位与遗传分析

利用以栽培稻9311为受体、普通野生稻为供体的染色体单片段置换系CSSL182,检测到一个与粒宽相关的QTL。CSSL182与受体亲本9311粒型性状差异显著,且只在8号染色体有一个野生稻导入片段。构建CSSL182/9311的F2次级分离群体,将粒宽QTL初定位在8号染色体的标记RM447和RM264之间,贡献率达22.49,将该QTL命名为qGW8。随后进一步设计区间内多态性分子标记引物,检测F2群体的2000株分离个体以及F2：3群体交换单株,结合后代表型验证,最终将qGW8精细定位到8号染色体10kb区间内。该区间内含有3个候选基因,基因测序发现这3个基因在双亲之间均含有丰富的变异。对双亲籽粒颖壳细胞电镜扫描观察发现,CSSL182的颖壳细胞宽度比9311减少16.7%。这一结果表明qGW8中来自野生稻的等位基因通过改变颖壳细胞形状影响粒型。     

基于极端混合池（BSA）全基因组重测序的羽衣甘蓝白色叶基因定位

筛选羽衣甘蓝白色叶形成相关的关键基因和位点以及心叶颜色性状的遗传规律，以羽衣甘蓝红叶亲本WR、白叶亲本WB及其构建的F2分离群体为试验材料，从F2群体中挑选红叶和白叶植株各20株，分别构建2个DNA混合池，对子代混合池和亲本分别开展50×和20×覆盖深度的全基因组重测序，定位白叶性状关联区间，并根据基因注释信息预测候选基因。结果表明，重测序获得3 987 718个单核酸多态性（SNP）标记，获得位于第2、3和9染色体的8个显著关联区间，根据基因注释和功能分析，筛选到8个候选基因（Bol030253、Bol029431、Bol012077、Bol007709、Bol030318、Bol030235、Bol030286和Bol005195）。将8个基因确定为羽衣甘蓝白叶的候选基因，可能在羽衣甘蓝叶片颜色形成过程中起着重要作用。     

基于染色体片段置换系的野生稻粒宽QTL-qGW8.1的精细定位

利用以栽培稻9311为受体、普通野生稻为供体的染色体单片段置换系CSSL182,检测到一个与粒宽相关的QTL。CSSL182与受体亲本9311粒型性状差异显著,且只在8号染色体有一个野生稻导入片段。构建CSSL182/9311的F_2次级分离群体,将粒宽QTL初定位在8号染色体的标记RM447和RM264之间,贡献率达22.49%,将该QTL命名为qGW8.1。随后进一步设计区间内多态性分子标记引物,检测F_2群体的2000株分离个体以及F_(2∶3)群体交换单株,结合后代表型验证,最终将qGW8.1精细定位到8号染色体10 kb区间内。该区间内含有3个候选基因,基因测序发现这3个基因在双亲之间均含有丰富的变异。对双亲子粒颖壳细胞电镜扫描观察发现,CSSL182的颖壳细胞宽度比9311减少16.7%。这一结果表明qGW8.1中来自野生稻的等位基因通过改变颖壳细胞形状影响粒型。     

与甘蓝型油菜重要经济性状有关的DNA克隆在拟南芥遗传图谱中的整合

拟南芥与油菜同属十字花科植物芸寡族,亲缘关系很近,基因组间的同源性很高,在用拟南芥EST克隆和油菜DNA克隆作探针定位了甘蓝型油菜一系列重要性状的基础上,对25个与油菜雄性不育恢复基因,硼高效利用基因,抗菌核病QTL及油菜种间杂种营养优势相关联的克隆进行了测序,在拟南芥基因组数据库中寻找到与这25个克隆高度同源的序列,根据这些高度同源序列在拟南芥染色体上的相位位置,将油菜DNA克隆整合到了拟南芥遗传图谱上,其中油菜硼高效基因BE1两侧的标记克隆整合在拟南芥第一染色体长臂一个较小的区段内,以该目标区段内的拟南芥EST克隆PA24为探针对甘蓝型油菜基因组比较作图,将该克隆定位在油菜连锁图BE1两侧标记之间,表明了利用基因组间的相互比较作图来精细定位芸薹属作物重要基因的可能性。     

甘蓝型油菜千粒重性状的QTL定位分析

该研究利用油菜双单倍体株系(348份)群体和已构建的遗传连锁图谱,采用复合区间作图法,对2009~2013年连续5年的千粒重性状表型数据进行QTL初步定位和分析,结果共获得46个显著性千粒重QTL,主要分布在A7、C1和C6等11条染色体上;其中qTSW-09 DL11-1的表型变异最高(19.63%),qTSW-11 DL9的表型变异最小(2.73%)。通过元分析方法将所获得的46个QTL进行整合,结果显示:cqTSW-C1-2的表型变异最大(10.64%),并发现多个整合后的一致性QTL能够在连续多年试验中被检测到,其中cqTSW-C1-3连续5年被检测到,表明控制千粒重的QTL在种植环境中能够稳定表达;同时,新发现位于C1染色体上的千粒重主效QTL cqTSW-C1-2,解释表型变异达到10.64%。油菜千粒重性状的QTL分析和主效QTL的获得,为进一步实现油菜大籽粒的分子育种和高产新品种的培育提供了重要的理论指导。     

基于元分析的抗玉米丝黑穗病QTL比较定位

以玉米遗传连锁图谱IBM2 2005 Neighbors为参考图谱,通过映射整合不同试验中的抗玉米丝黑穗病QTL,构建QTL综合图谱。在国内外种质中,共发现22个抗病QTL,分布在除第7染色体外的9条玉米染色体上。采用元分析技术,获得2个“一致性”抗病QTL,图距分别为8.79 cM和18.92cM。从MaizeGDB网站下载“一致性”QTL区间内基因和标记的原始序列;采用NCBI网站在线软件BLASTx通过同源比对在2个“一致性”QTL区间内初步获得4个抗病位置候选基因。借助比较基因电子定位策略,将69个水稻和玉米抗性基因定位于玉米IBM2图谱上,在2个“一致性”QTL区间内分别发现1个水稻抗性基因,初步推断为抗病位置候选基因。本文结果为抗玉米丝黑穗病QTL精细定位和分子育种提供了基础。     

甘蓝花色性状QTL定位及候选基因变异分析

花色性状是甘蓝的一个重要性状,在吸引和指示传粉者、保护花器官、维持花组织能量平衡、测定品种异交率及纯度、检测性状转移等方面有重要作用。为了鉴定控制和影响甘蓝花色的遗传位点和候选基因,本研究利用芥蓝(白花)与野生甘蓝(黄花)构建了F2分离群体,并分别利用基于分子标记遗传连锁图谱和基于SNP芯片分析的QTL扫描技术,对甘蓝花色性状进行QTL定位。本研究结论如下:甘蓝花色性状由C03染色体上一个部分显性主效QTL位点控制,并受到C02上一个微效加性QTL的影响;主效QTL候选基因BoCCD4编码区778～780 bp处3个碱基的插入极可能导致BoCCD4功能丧失,从而呈现黄色表型。本研究确认了控制甘蓝白花性状的主效QTL位置,鉴定到了候选基因并发现了与文献报道不同的新变异位点,为进一步了解芸薹属物种花色的遗传和变异提供了新的数据。     

单核苷酸多态性及其在鸡QTL定位上的应用

单核苷酸多态性是指DNA序列上的单个碱基变异,它具有分布广、多态信息含量大、易于检测和统计分析等优点,能较好用于基因图谱构建和数量性状QTL定位研究,被称为继RFLP和微卫星标记之后的第3代基因遗传标记。本文综述了单核苷酸多态性的性质及检测技术、利用候选基因SNP进行鸡QTL定位研究的现状,并对未来SNP的应用前景进行了展望。Abstract:Single nucleotide polymorphism (SNP) refers to the change of single nucleotide in DNA sequence.Because of its high density in genomes and easy in detection and analysis statistically,SNP can be used in genetic linkage map construction and QTL mapping.Here,the characters and detecting technology of SNP,as well as the status and foreground of the use of candidate gene SNP in chicken QTL mapping are introduced.     

Characterization of metabolite quantitative trait loci and metabolic networks that control glucosinolate concentration in the seeds and leaves of Brassica napus

? Glucosinolates are a major class of secondary metabolites found in the Brassicaceae, whose degradation products are proving to be increasingly important for human health and in crop protection. ? The genetic and metabolic basis of glucosinolate accumulation was dissected through analysis of total glucosinolate concentration and its individual components in both leaves and seeds of a doubled-haploid (DH) mapping population of oilseed rape/canola (Brassica napus). ? The quantitative trait loci (QTL) that had an effect on glucosinolate concentration in either or both of the organs were integrated, resulting in 105 metabolite QTL (mQTL). Pairwise correlations between individual glucosinolates and prior knowledge of the metabolic pathways involved in the biosynthesis of different glucosinolates allowed us to predict the function of genes underlying the mQTL. Moreover, this information allowed us to construct an advanced metabolic network and associated epistatic interactions responsible for the glucosinolate composition in both leaves and seeds of B. napus. ? A number of previously unknown potential regulatory relationships involved in glucosinolate synthesis were identified and this study illustrates how genetic variation can affect a biochemical pathway.     

Quantitative trait loci for glucosinolate accumulation in Brassica rapa leaves

Glucosinolates and their breakdown products have been recognized for their effects on plant defense, human health, flavor and taste of cruciferous vegetables. Despite this importance, little is known about the regulation of the biosynthesis and degradation in Brassica rapa. Here, the identification of quantitative trait loci (QTL) for glucosinolate accumulation in B. rapa leaves in two novel segregating double haploid (DH) populations is reported: DH38, derived from a cross between yellow sarson R500 and pak choi variety HK Naibaicai; and DH30, from a cross between yellow sarson R500 and Kairyou Hakata, a Japanese vegetable turnip variety. An integrated map of 1068 cM with 10 linkage groups, assigned to the international agreed nomenclature, is developed based on the two individual DH maps with the common parent using amplified fragment length polymorphism (AFLP) and single sequence repeat (SSR) markers. Eight different glucosinolate compounds were detected in parents and F(1)s of the DH populations and found to segregate quantitatively in the DH populations. QTL analysis identified 16 loci controlling aliphatic glucosinolate accumulation, three loci controlling total indolic glucosinolate concentration and three loci regulating aromatic glucosinolate concentrations. Both comparative genomic analyses based on Arabidopsis-Brassica rapa synteny and mapping of candidate orthologous genes in B. rapa allowed the selection of genes involved in the glucosinolate biosynthesis pathway that may account for the identified QTL.     

Regional association analysis delineates a sequenced chromosome region influencing antinutritive seed meal compounds in oilseed rape

This study describes the use of regional association analyses to delineate a sequenced region of a Brassica napus chromosome with a significant effect on antinutritive seed meal compounds in oilseed rape. A major quantitative trait locus (QTL) influencing seed colour, fibre content, and phenolic compounds was mapped to the same position on B. napus chromosome A9 in biparental mapping populations from two different yellow-seeded × black-seeded B. napus crosses. Sequences of markers spanning the QTL region identified synteny to a sequence contig from the corresponding chromosome A9 in Brassica rapa. Remapping of sequence-derived markers originating from the B. rapa sequence contig confirmed their position within the QTL. One of these markers also mapped to a seed colour and fibre QTL on the same chromosome in a black-seeded × black-seeded B. napus cross. Consequently, regional association analysis was performed in a genetically diverse panel of dark-seeded, winter-type oilseed rape accessions. For this we used closely spaced simple sequence repeat (SSR) markers spanning the sequence contig covering the QTL region. Correction for population structure was performed using a set of genome-wide SSR markers. The identification of QTL-derived markers with significant associations to seed colour, fibre content, and phenolic compounds in the association panel enabled the identification of positional and functional candidate genes for B. napus seed meal quality within a small segment of the B. rapa genome sequence.     

Identification of FAD2 and FAD3 genes in Brassica napus genome and development of allele-specific markers for high oleic and low linolenic acid contents

Modification of oleic acid (C18:1) and linolenic acid (C18:3) contents in seeds is one of the major goals for quality breeding after removal of erucic acid in oilseed rape (Brassica napus). The fatty acid desaturase genes FAD2 and FAD3 have been shown as the major genes for the control of C18:1 and C18:3 contents. However, the genome structure and locus distributions of the two gene families in amphidiploid B. napus are still not completely understood to date. In the present study, all copies of FAD2 and FAD3 genes in the A- and C-genome of B. napus and its two diploid progenitor species, Brassica rapa and Brassica oleracea, were identified through bioinformatic analysis and extensive molecular cloning. Two FAD2 genes exist in B. rapa and B. oleracea, and four copies of FAD2 genes exist in B. napus. Three and six copies of FAD3 genes were identified in diploid species and amphidiploid species, respectively. The genetic control of high C18:1 and low C18:3 contents in a double haploid population was investigated through mapping of the quantitative trait loci (QTL) for the traits and the molecular cloning of the underlying genes. One major QTL of BnaA.FAD2.a located on A5 chromosome was responsible for the high C18:1 content. A deleted mutation in the BnaA.FAD2.a locus was uncovered, which represented a previously unidentified allele for the high oleic variation in B. napus species. Two major QTLs on A4 and C4 chromosomes were found to be responsible for the low C18:3 content in the DH population as well as in SW Hickory. Furthermore, several single base pair changes in BnaA.FAD3.b and BnaC.FAD3.b were identified to cause the phenotype of low C18:3 content. Based on the results of genetic mapping and identified sequences, allele-specific markers were developed for FAD2 and FAD3 genes. Particularly, single-nucleotide amplified polymorphisms markers for FAD3 alleles were demonstrated to be a reliable type of SNP markers for unambiguous identification of genotypes with different content of C18:3 in amphidiploid B. napus.     

Homoeologous loci control the accumulation of seed glucosinolates in oilseed rape (Brassica napus).

The genetic control of seed glucosinolate content in oilseed rape was investigated using two intervarietal backcross populations. Four QTLs segregating in the population derived from a Brassica napus L. 'Victor' x Brassica napus L. 'Tapidor' cross, together accounting for 76% of the phenotypic variation, were mapped. Three of these loci also appeared to control the accumulation of seed glucosinolates in a Brassica napus L. 'Bienvenu' x 'Tapidor' cross, and accounted for 86% of the phenotypic variation. The three QTLs common to both populations mapped to homoeologous regions of the B. napus genome, suggesting that seed glucosinolate accumulation is controlled by duplicate genes. It was possible to extend the comparative analysis of QTLs controlling seed glucosinolate accumulation by aligning the published genetic maps generated by several research groups. This comparative mapping demonstrated that high-glucosinolate varieties often carry low-glucosinolate alleles at one or more of the loci controlling seed glucosinolate accumulation.     

Mapping of homoeologous chromosome exchanges influencing quantitative trait variation in Brassica napus

Genomic rearrangements arising during polyploidization are an important source of genetic and phenotypic variation in the recent allopolyploid crop Brassica napus. Exchanges among homoeologous chromosomes, due to interhomoeologue pairing, and deletions without compensating homoeologous duplications are observed in both natural B. napus and synthetic B. napus. Rearrangements of large or small chromosome segments induce gene copy number variation (CNV) and can potentially cause phenotypic changes. Unfortunately, complex genome restructuring is difficult to deal with in linkage mapping studies. Here, we demonstrate how high‐density genetic mapping with codominant, physically anchored SNP markers can detect segmental homoeologous exchanges (HE) as well as deletions and accurately link these to QTL. We validated rearrangements detected in genetic mapping data by whole‐genome resequencing of parental lines along with cytogenetic analysis using fluorescence in situ hybridization with bacterial artificial chromosome probes (BAC‐FISH) coupled with PCR using primers specific to the rearranged region. Using a well‐known QTL region influencing seed quality traits as an example, we confirmed that HE underlies the trait variation in a DH population involving a synthetic B. napus trait donor, and succeeded in narrowing the QTL to a small defined interval that enables delineation of key candidate genes.     

Fine mapping of loci involved with glucosinolate biosynthesis in oilseed mustard (Brassica juncea) using genomic information from allied species

Fine mapping of six seed glucosinolate QTL (J2Gsl1, J3Gsl2, J9Gsl3, J16Gsl4, J17Gsl5 and J3Gsl6) (Ramchiary et al. in Theor Appl Genet 116:77–85, 2007a) was undertaken by the candidate gene approach. Based on the DNA sequences from Arabidopsis and Brassica oleracea for the different genes involved in the aliphatic glucosinolate biosynthesis, candidate genes were amplified and sequenced from high to low glucosinolate Brassica juncea lines Varuna and Heera, respectively. Of the 20 paralogues identified, 17 paralogues belonging to six gene families were mapped to 12 of the 18 linkage groups of B. juncea genome. Co-mapping of candidate genes with glucosinolate QTL revealed that the candidate gene BjuA.GSL-ELONG.a mapped to the QTL interval of J2Gsl1, BjuA.GSL-ELONG.c, BjuA.GSL-ELONG.d and BjuA.Myb28.a mapped to the QTL interval of J3Gsl2, BjuA.GSL-ALK.a mapped to the QTL interval of J3Gsl6 and BjuB.Myb28.a mapped to the QTL interval of J17Gsl5. The QTL J9Gsl3 and J16Gsl4 did not correspond to any of the mapped candidate genes. The functionality and contribution of different candidate genes/QTL was assessed by allelic variation study using phenotypic data of 785 BC₄DH lines. It was observed that BjuA.Myb28.a and J9Gsl3 contributed significantly to the base level glucosinolate production while J16Gsl4, probably GSL-PRO, BjuA.GSL-ELONG.a and BjuA.GSL-ELONG.c contributed to the C3, C4 and C5 elongation pathways, respectively. Three A genome QTL: J2Gsl1harbouring BjuA.GSL-ELONG.a, J3Gsl2 harbouring both BjuA.GSL-ELONG.c and BjuA.Myb28.a and J9Gsl3, possibly the ‘Bronowski genes’, were identified as most important loci for breeding low glucosinolate B. juncea. We observed two-step genetic control of seed glucosinolate in B. juncea mainly effected by these three A genome QTL. This study, therefore, provides clues to the genetic mechanism of ‘Bronowski genes’ controlling the glucosinolate trait and also provides efficient markers for marker-assisted introgression of low glucosinolate trait in B. juncea. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Mapping the mosaic of ancestral genotypes in a cultivar of oilseed rape (Brassica napus) selected via pedigree breeding.

Recent oilseed rape breeding has produced low glucosinolate cultivars that yield proteinaceous meal suitable for animal feed. The low glucosinolate character was introduced into modern cultivars from Brassica napus 'Bronowski', a cultivar that is agronomically inferior in most other respects. Residual segments of 'Bronowski' genotype in modern cultivars probably cause reduced yield, poorer winter hardiness, and lower oil content. The quantity and distribution of the 'Bronowski' genotype in the modern oilseed rape cultivar Brassica napus 'Tapidor' was investigated using a segregating population derived from a cross between 'Tapidor' and its high glucosinolate progenitor. This population was analyzed with 65 informative Brassica RFLP probes and a genetic linkage map, based on the segregation at 77 polymorphic loci, was constructed. The mapping identified 15 residual segments of donor genotype in 'Tapidor', which together occupy approximately 29% of the B. napus genome. Mapping the loci that control variation for the accumulation of total seed glucosinolates in the segregating population has identified three loci that together explain >90% of the variation for this character. All of these loci are in donor segments of the 'Tapidor' genome. This result shows the extent to which conventional breeding programmes have difficulty in eliminating residual segments of donor genotype from elite material.     

Molecular mapping of Arabidopsis thaliana lipid-related orthologous genes in Brassica napus

Quantitative Trait Loci (QTL) for oil content has been previously analyzed in a SG-DH population from a cross between a Chinese cultivar and a European cultivar of Brassica napus. Eight QTL with additive and epistatic effects, and with environmental interactions were evaluated. Here we present an integrated linkage map of this population predominantly based on informative markers derived from Brassica sequences, including 249 orthologous A. thaliana genes, where nearly half (112) are acyl lipid metabolism related genes. Comparative genomic analysis between B. napus and A. thaliana revealed 33 colinearity regions. Each of the conserved A. thaliana segments is present two to six?times in the B. napus genome. Approximately half of the mapped lipid-related orthologous gene loci (76/137) were assigned in these conserved colinearity regions. QTL analysis for seed oil content was performed using the new map and phenotypic data from 11 different field trials. Nine significant QTL were identified on linkage groups A1, A5, A7, A9, C2, C3, C6 and C8, together explaining 57.79% of the total phenotypic variation. A total of 14 lipid related candidate gene loci were located in the confidence intervals of six of these QTL, of which ten were assigned in the conserved colinearity regions and felled in the most frequently overlapped QTL intervals. The information obtained from this study demonstrates the potential role of the suggested candidate genes in rapeseed kernel oil accumulation.     

Design of New Genome- and Gene-Sourced Primers and Identification of QTL for Seed Oil Content in a Specially High-Oil Brassica napus Cultivar

Rapeseed (Brassica napus L.) is one of most important oilseed crops in the world. There are now various rapeseed cultivars in nature that differ in their seed oil content because they vary in oil-content alleles and there are high-oil alleles among the high-oil rapeseed cultivars. For these experiments, we generated doubled haploid (DH) lines derived from the cross between the specially high-oil cultivar zy036 whose seed oil content is approximately 50% and the specially low-oil cultivar 51070 whose seed oil content is approximately 36%. First, to address the deficiency in polymorphic markers, we designed 5944 pairs of newly developed genome-sourced primers and 443 pairs of newly developed primers related to oil-content genes to complement the 2244 pairs of publicly available primers. Second, we constructed a new DH genetic linkage map using 527 molecular markers, consisting of 181 publicly available markers, 298 newly developed genome-sourced markers and 48 newly developed markers related to oil-content genes. The map contained 19 linkage groups, covering a total length of 2,265.54 cM with an average distance between markers of 4.30 cM. Third, we identified quantitative trait loci (QTL) for seed oil content using field data collected at three sites over 3 years, and found a total of 12 QTL. Of the 12 QTL associated with seed oil content identified, 9 were high-oil QTL which derived from the specially high-oil cultivar zy036. Two high-oil QTL on chromosomes A2 and C9 co-localized in two out of three trials. By QTL mapping for seed oil content, we found four candidate genes for seed oil content related to four gene markers: GSNP39, GSSR161, GIFLP106 and GIFLP046. This information will be useful for cloning functional genes correlated with seed oil content in the future.