野生小豆种质资源遗传多样性的RAPD分析

利用RAPD分子标记技术对16份不同来源地的野生小豆种质进行基因组DNA多态性分析,25个随机引物共扩增出240个DNA片段,其中197条谱带表现出多态性。对基于数字化处理后的多态性谱带进行聚类,16个野生小豆种质可划分为4个类群：日本类群、中国类群、不丹类群和尼泊尔类群,材料的归组与其来源地有显著的相关性。     

利用普通六倍体小麦和西藏半野生小麦杂交衍生的重组自交系定位小麦芒长QTL

芒长是普通小麦的重要农艺性状,受多个基因控制。本研究利用长芒的普通小麦郑麦9023与无芒的西藏半野生小麦Q1028构建一个重组自交系群体(186个株系);采用SSR和DAr T分子标记,构建覆盖小麦全基因组的遗传图谱(2597 cM)。基于重组自交系群体两年芒长表型数据,采用ICIM作图法对小麦芒长性状进行QTL定位分析。共检测到2个与芒长相关的QTL,即Qwa.sau-4AS和Qwa.sau-5AL。它们分别位于4AS和5AL染色体上,可分别解释7.4%和27.3%的表型变异。这2个QTL效应可能分别来源于钩芒基因Hd与抑芒基因B1。利用连锁标记进行基因型分析,表明Qwa.sau-4AS与Qwa.sau-5AL对芒长的抑制效果具有累加效应,且Qwa.sau-5AL效应强于Qwa.sau-4AS。本研究将为精细定位及克隆这2个QTL奠定基础。     

Cross-Compatibility of Cultivated Adzuki Bean (Vigna angularis) and Rice Bean (V. umbellata) with Their Wild Relatives

The wild Ceratotropis species which occur in Japan consist of Vigna angularis var. nipponensis, V. nakashimae, V. riukiuensis and V. reflexo-pilosa. The main aim of the present study is to detect cross-compatibility of adzuki bean and rice bean with these wild relatives. The cross of V. nakasbimae with adzuki bean was successful in both directions and viable seeds were produced, while the cross with rice bean was not successful. V. riukiuensis is cross-compatible with adzuki bean and rice bean only when V. riukiuensis is used as male parent and the hybrid seeds showed normal germination. The cross between V. reflexopilosa and the two cultigens adzuki bean and rice bean did not succeed in either direction. The present results suggest that V. nakashimae and V. riukiuensis could be used as gene sources for the adzuki bean breeding programme through interspecific hybridization. Successful hybrids between adzuki bean and rice bean cannot easily be produced. However, since V. riukiuensis is cross-compatible with both adzuki bean and rice bean, V. riukiuensis could be useful as a bridge species between adzuki bean and rice bean.     

幼苗期淹水胁迫及喷施烯效唑对小豆生理和产量的影响

为探究幼苗期淹水胁迫及喷施烯效唑(S3307)对小豆(Vignaangularis)生理代谢和产量的影响,本研究于2018—2019年以龙小豆4号和天津红为试验材料,盆栽条件下,苗期进行预喷施S3307,同时连续淹水处理5 d并每天取样,测定分析相关生理指标。结果表明,幼苗期淹水胁迫引起小豆叶片H2O2和MDA含量、脯氨酸和可溶性蛋白含量显著提高, ABA、IAA和SA含量显著提高, SOD、POD和CAT活性显著提高,淹水处理5 d导致龙小豆4号单盆产量显著下降8.40%～12.61%,天津红显著下降9.91%～10.01%。S3307具有抵御淹水胁迫的作用,能有效增加小豆叶片脯氨酸和可溶性蛋白含量,显著降低H2O2和MDA含量,能够显著提高SOD和POD活性以及SOD/POD、SOD/CAT,显著增加ABA和SA含量,并显著抑制IAA含量的上升。喷施S3307使淹水4 d的龙小豆4号产量显著提高2.85%～6.18%,使天津红淹水4 d的产量显著提高2.85%～3.95%。综上,淹水胁迫下,不同品种小豆在活性氧物质积累、膜质过氧化、抗氧化酶活性以及激素水平等方面的生理应激存在显著差...     

Identification and molecular mapping of Flowering Date1 (FD1), a major photoperiod insensitivity gene in the adzuki bean (Vigna angularis)

The induction of flowering under long‐day conditions is an important adaptation by short‐day plants, such as adzuki beans (Vigna angularis), to high‐latitude environments. This study clarified the genetic control underlying the long‐day insensitivity of adzuki bean cultivar ‘Shumari’. ‘Shumari’ was found to be insensitive to a 16‐h day, whereas landrace Acc2265 was highly sensitive. When grown under natural long‐day conditions at Obihiro (42°9′N), Acc2265 initiated flowering at least 80 days after ‘Shumari’. When 86 recombinant inbred lines (RILs) derived from crosses between ‘Shumari’ and Acc2265 were grown under these conditions, their flowering dates ranged from the middle of July to the end of October. The distinct bimodal distribution in the RIL population was due to a single major gene, designated Flowering Date1 (FD1). Molecular mapping showed that FD1 was located between the SSR markers Az02‐37M3 and Az02‐40M9, at distances of 6 and 10.4 cM, respectively, on linkage group 2. RILs carrying FD1^S lacked long‐day sensitivity, whereas RILs carrying FD1^A were sensitive to long‐day conditions, confirming that FD1 controls long‐day sensitivity.     

以地理来源分组和利用表型数据构建中国小豆核心种质

中国国家种质库里保存着4 877份小豆(Vigna angularis)种质资源,但仅有部分资源在小豆的改良中被利用。构建小豆核心种质, 不但能简化管理, 而且可以提高遗传资源在小豆改良中的利用效率。根据我国小豆的地理来源进行分组, 在分组的基础上利用表型数据, 以类平均法聚类构建了中国现有小豆核心种质, 共435份资源, 占资源总量的8.92%, 涵盖98.3%的表型变异类型, 控制不同性状表型相关的相互适应的遗传复杂性在核心种质中也得到了适当的保持。不同性状的均值t测验、方差F测验、表型频率分布、χ²测验等分析表明, 所构建的小豆核心种质可以作为整体资源的代表性样本, 在小豆种质资源的充分利用中将发挥重要作用。     

快中子诱变小豆京农6号突变体筛选

为了进行小豆种质创新和获得基因定位和克隆及其基因功能分析的突变体材料,分别用30、35、40、45、50、55 Gy剂量的快中子,辐照小豆‘京农6号’干种子。M1出苗率分别为31.72%、29.06%、30.29%、36.51%、16.15%和15.17%,变异率为3.20%、4.70%、6.75%、6.61%、4.00%和2.20%。辐射剂量增加,出苗率降低、变异率呈增加趋势,M3代共获得77份叶形、株型、粒型、籽粒大小、粒色、叶色、早熟、丰产等性状变异材料。40~45 Gy为小豆快中子诱变的适宜剂量,这些突变体为小豆相关基因定位克隆、功能分析和育种提供了有价值的材料。     

Mapping of quantitative trait loci for phytic acid and phosphorus contents in seed and seedling of mungbean (Vigna radiata (L.) Wilczek)

Phytic acid (PA) is the storage form of phosphorus (P) in seeds and plays an important role in the nutritional quality of food crops. There is little information on the genetics of seed and seedling PA in mungbean [Vigna radiata (L.) Wilczek]. Quantitative trait loci (QTL) were identified for phytic acid P (PAP), total P (TP), and inorganic P (IP) in mungbean seeds and seedlings, and for flowering, maturity and seed weight, in an F₂ population developed from a cross between low PAP cultivated mungbean (V1725BG) and high PAP wild mungbean (AusTRCF321925). Seven QTLs were detected for P compounds in seed; two for PAP, four for IP and one for TP. Six QTLs were identified for P compounds in seedling; three for PAP, two for TP and one for IP. Only one QTL co-localized between P compounds in seed and seedling suggesting that low PAP seed and low PAP seedling must be selected for at different QTLs. Seed PAP and TP were positively correlated with days to flowering and maturity, indicating the importance of plant phenology to seed P content.     

Mapping QTL controlling soybean seed sucrose and oligosaccharides in a single family of soybean nested association mapping (SoyNAM) population

The meal value of Soybean for monogastric animals is determined partly by sucrose, raffinose and stachyose. Of these, sucrose is desirable, while raffinose and stachyose are indigestible, causing flatulence and abdominal discomfort. The objective of this study was to identify quantitative trait loci (QTL) controlling seed sucrose, raffinose, and stachyose in a set of 140 SoyNAM (Nested Association Mapping) recombinant inbred lines (RILs), developed from the cross between lines IA3023 and LD02‐4485. A total of 3,038 SNP markers from the Illumina SoyNAM BeadChip SNP were used to map the QTLs for sucrose and the RFOs, raffinose, and stachyose. Significant genotypic differences (p < .001) among RILs were observed for sucrose, raffinose and stachyose contents across years. A 3038 Illumina SoyNAM BeadChip SNPs identified three QTLs for sucrose, one on chromosome 1, explaining 10% variance and two on chromosome 3 each explaining 22%. Raffinose QTL was detected on chromosome 6, explaining 6% variance. The mapped QTLs were novel and spanned regions harbouring candidate genes with roles in plant growth including seed development.     

Molecular mapping of quantitative trait loci (QTLs) controlling aluminium tolerance in bread wheat

Aluminium (Al) toxicity is a major constraint to crop productivity in acidic soils. A quantitative trait locus (QTL) analysis was performed to identify the genetic basis of Al tolerance in the wheat cultivar ‘Chinese Spring’. A nutrient solution culture approach was undertaken with the root tolerance index (RTI) and hematoxylin staining method as parameters to assess the Al tolerance. Using a set of D genome introgression lines, a major Al tolerance QTL was located on chromosome arm 4DL, explaining 31% of the phenotypic variance present in the population. A doubled haploid population was used to map a second major Al tolerance QTL to chromosome arm 3BL. This major QTL (Qalt _CS .ipk-3B) in ‘Chinese Spring’ accounted for 49% of the phenotypic variation. Linkage of this latter QTL to SSR markers opens the possibility to apply marker-assisted selection (MAS) and pyramiding of this new QTL to improve the Al tolerance of wheat cultivars in breeding programmes.     

基于RAD-seq的荸荠SSR标记开发

荸荠(Eleocharis dulcis)是一种重要的特色水生蔬菜,为开发用于荸荠遗传学研究的简单重复序列(SSR)分子标记,本研究基于RAD-seq技术对荸荠进行简化基因组测序,并进行SSR标记开发与引物设计。结果显示,共检测到5039个SSR位点,其中4127个SSR位点可利用并成功设计引物。在可利用的SSR位点中,三碱基重复基序类型所占比例最高,数量为1894个,占位点总数的45.89%;其次是二碱基重复基序类型,数量为1406个,占位点总数的34.07%。随机选取了100对引物进行有效性验证,扩增成功率为93%,从扩增成功的引物中选取83对引物对两个品种进行多态性检测,83对引物共得到232条条带,其中多态性条带为128条,具有多态性的引物为60对,多态性比率为72.28%。通过RAD-seq开发的荸荠SSR标记有效性和多态性较高,为后期荸荠种质资源的高效利用、品种的遗传改良及分子育种提供了基础。     

理化诱变小豆京农6号突变体的鉴定

选用小豆品种京农6号种子,分别采用甲基磺酸乙酯(EMS)(0.5%、0.9%和1.4%处理12h和24h)、电子束(100、300、600Gy)、60Co-γ(400Gy)诱变处理,将处理后的种子种于大田,鉴定后代植株性状的变异。观察表明,EMS诱变的变异类型最丰富、60Co-γ射线次之、电子束产生的变异类型较单一。EMS处理小豆以浓度0.5%和0.9%处理24h为宜;0.5%EMS处理的粒色和荚色变异突出,有鲜红、黄白、绿白粒色和黑荚、褐荚、黑褐荚变异;0.9%处理的叶形变异突出,有鸡爪叶、剑叶、肾形叶、小密叶等突变类型;电子束诱变后,M2变异率分别为4.09%、3.64%和2.22%。400Gy60Co-γ射线处理种子,后代变异率为7.23%。通过两年的鉴定筛选,获得937个EMS诱变M3代株系,934个60Co-γ射线和电子束诱变M2代株系,已得到株高、叶形、叶色、粒形、粒色、荚色、无分枝、多分枝、叶簇生、分枝簇生、光叶、蔓生、有限结荚习性、株型松散、育性、成熟特性等突变体材料1490份。本研究为小豆基因遗传分析、基因定位与克隆及其进一步的基因功能分析奠定了基础,为小豆育种提供了重要的材料。     

大豆叶柄角的QTL定位分析

叶柄角是大豆株型的重要构成因素,影响大豆冠层结构、光合作用效率以及最终产量。解析大豆叶柄角的遗传基础对提升大豆产量具有重要意义。本研究以2个叶柄角具有显著差异的亲本BLA和SLA以及它们衍生的RIL群体为材料,构建高密度的遗传图谱,对大豆不同部位的叶柄角进行QTL分析,并利用近等基因系验证部分QTL。遗传分析结果显示,叶柄角呈连续正态分布,符合数量性状遗传特征。利用GBS技术构建了包含859个Bin标记的大豆高密度遗传图谱,总遗传长度为2326.9cM,标记间平均距离为2.763cM;共检测到14个调控叶柄角的QTL,LOD值在2.58~4.80之间,可解释遗传变异范围在6.9%~12.4%之间,其中5个QTL定位在第12染色体上且成簇存在;构建的qLA12和qLA18的近等基因系表型结果显示,叶柄角在同一对近等基因家系间差异显著,表明qLA12和qLA18是2个可信的QTL。本研究为进一步克隆调控叶柄角功能基因奠定了基础,为大豆理想株型育种提供了遗传材料。     

绿豆分子遗传图谱构建及若干农艺性状的QTL定位分析

利用花叶1号×紫茎1号杂交后代衍生的208个F2家系组建群体,构建含有95个SSR标记位点的遗传连锁图谱,该图谱包含11个连锁群,全长1457.47 c M,标记平均间距为15.34 c M。利用复合区间作图法,对株高、幼茎色、主茎色、生长习性、结荚习性、复叶叶形和成熟叶色等农艺性状进行QTL分析,分别检测到与株高、幼茎色、主茎色、复叶叶形有关的QTL各1个,贡献率在8.49%~66.64%之间;与结荚习性有关的QTL3个,贡献率在60.32%~80.36%之间;与成熟叶色有关QTL 4个,贡献率在69.06%~87.35%之间;与生长习性有关的QTL数量最多,共26个,贡献率在58.32%~99.51%之间。上述QTL主要分布在LG1、LG2、LG4、LG8和LG10连锁群,其中LG1最少,仅检测到生长习性的1个QTL,LG4最多,包含了幼茎色、主茎色、结荚习性、生长习性、复叶叶形、成熟期叶色6个农艺性状的15个QTL;这些QTL既可以应用于绿豆育种的分子标记辅助选择,也对深入研究这些性状的遗传奠定了基础。     

水稻单片段替换系群体的建立及QTL定位

水稻中许多重要的农艺性状属数量性状，由多基因(QTL)控制。研究QTL的遗传特性和遗传效应对于培育高产和稳产的水稻品种具有十分重要的意义。本研究以6个优良的品种为供体亲本，以华粳籼74为轮回亲本，通过微卫星标记辅助回交选择培育了一批单片段替换系，随后利用所培育的单片段替换系进行了QTL分析和基因定位。主要结果有：1、利用258个微卫星标记对6个供体亲本和轮回亲本间的多态性进行了筛选。6个供体亲本与轮回亲本间的多态率在32．98％至60．78％之间，平均47．81％，粳型供体亲本比籼型供体亲本的多态性要高。2、随着回交代数的增加，植株所含的替换片段数逐渐减少。在BC2F1、BC3F1、BC3F2和BC3F3代，平均每个植株携带有12．50、5．98、1．69和1．46个替换片段。替换片段的平均长度也随回交和自交代数的增加而逐渐变短。在BC2F1、BC3F1、BC3F2和BC3F3代，替换片段的平均长度分别为25．43cM、22．38cM、20．78cM和18．15cM.回交世代替换片段变短的速率(11．99％)比自交世代变短速率(7．15％)要快。在BC2F1、BC3Fl、BC3F2和BC3F3代，轮回亲本基因组的恢复率分别为82．24％、92．55％、98．04％j阳98．52％。3、在BC3F2和BC3F3代，共选育出111个单片段替换系，其中独一无二的单片段替换系共42个。BC3F2代替换系中替换片段的估算长度在2．00cM到64．80cM之间，平均为21．75cM，而BC3F3代中替换片段的估算长度在6．05cM到48．90cM之间，平均为20．95cMc，12条染色体中仅第11染色体没有选择到单片段替换系。所选育的单片段替换系中替换片段的总长为2367．50cM，基因组的覆盖长度为704．50cM，覆盖率为39．25％。4、在52个单片段替换系的22个性状中共鉴定出了234个QTL。每个性状鉴定出的QTL数在3到19个之间，平均4．50个。每个替换系鉴定出的QTL在2到15个之间，平均10．64个。QTL加性效应的大小因性状和替换系不同而不同，低至-0．02(0．79％)的加性效应(如谷粒宽度)均能检测到。在RM237．RM322、RM225和RM481标记附近的各种性状中同时检测到了增效和减效的QTL。5、通过染色体替换作图，对7个单片段替换系中16个性状的QTL进行了定位。利用携带有相同替换片段的替换系进行QTL位置的确定。6、通过分析复杂性状的加性效应，对影响植高、每穗粒数和粒型的相关性状进行了分析，分析QTL之间的相关性。7、对控制抽穗期、稃尖颜色、植株高度和粒型的基因进行了定位。抽穗期基因Hd-8表现为单基因显性遗传，定位于第8染色体上，与PSMl55紧密连锁。紫色稃尖基因Pa-6表现为单基因显性遗传，定位于第6染色体上，与RM253紧密连锁。株高基因Ph-1-3定位于第1染色体上，与PSM331紧密连锁。粒型基因Rlw-8-2为首次报道，定位于第8染色体的末端，与RM447紧密连锁，长粒表现为单基因隐性遗传。本研究通过分子标记辅助选择培育了一批单片段替换系，并成功地对这批材料进行了QTL分析和基因定位，从而证实了在水稻中通过微卫星标记辅助回交选育单片段替换系和进行QTL分析的可行性。     

De novo assembly of the complete organelle genome sequences of azuki bean (Vigna angularis) using next-generation sequencers

Since chloroplasts and mitochondria are maternally inherited and have unique features in evolution, DNA sequences of those organelle genomes have been broadly used in phylogenetic studies. Thanks to recent progress in next-generation sequencer (NGS) technology, whole-genome sequencing can be easily performed. Here, using NGS data generated by Roche GS Titanium and Illumina Hiseq 2000, we performed a hybrid assembly of organelle genome sequences of Vigna angularis (azuki bean). Both the mitochondrial genome (mtDNA) and the chloroplast genome (cpDNA) of V. angularis have very similar size and gene content to those of V. radiata (mungbean). However, in structure, mtDNA sequences have undergone many recombination events after divergence from the common ancestor of V. angularis and V. radiata, whereas cpDNAs are almost identical between the two. The stability of cpDNAs and the variability of mtDNAs was further confirmed by comparative analysis of Vigna organelles with model plants Lotus japonicus and Arabidopsis thaliana.     

Identification of QTLs controlling harvest time and fruit skin color in Japanese pear (Pyrus pyrifolia Nakai)

Using an F₁ population from a cross between Japanese pear (Pyrus pyrifolia Nakai) cultivars ‘Akiakari’ and ‘Taihaku’, we performed quantitative trait locus (QTL) analysis of seven fruit traits (harvest time, fruit skin color, flesh firmness, fruit weight, acid content, total soluble solids content, and preharvest fruit drop). The constructed simple sequence repeat-based genetic linkage map of ‘Akiakari’ consisted of 208 loci and spanned 799 cM; that of ‘Taihaku’ consisted of 275 loci and spanned 1039 cM. Out of significant QTLs, two QTLs for harvest time, one for fruit skin color, and one for flesh firmness were stably detected in two successive years. The QTLs for harvest time were located at the bottom of linkage group (LG) Tai3 (nearest marker: BGA35) and at the top of LG Tai15 (nearest markers: PPACS2 and MEST050), in good accordance with results of genome-wide association study. The PPACS2 gene, a member of the ACC synthase gene family, may control harvest time, preharvest fruit drop, and fruit storage potential. One major QTL associated with fruit skin color was identified at the top of LG 8. QTLs identified in this study would be useful for marker-assisted selection in Japanese pear breeding programs.     

Construction of high-density linkage map and identification of QTLs associated with resistance to black rot in radish (Raphanus sativus) by RAD sequencing

High-density marker-based QTL mapping can serve as an effective strategy to identify novel genomic information to facilitate crop improvement. In this study, we genotyped an F₂ population (KB12-1 × PP12-1) using a RAD-seq approach and constructed a high-density linkage map for radish. After a series of filtering procedures were performed, 17,124 SNPs and 3,336 indels with aa × bb genotyping were retained to obtain bin markers. Then, a linkage map comprising a total of 1,221 bin markers in nine linkage groups spanning 1,467.3 cM with an average marker interval of 1.2 cM was constructed. We evaluated the resistance of the F₂ mapping population to black rot using F₃ progeny, and two major QTLs related to black rot resistance were identified based on this map. Among these QTLs, qBRR2 on Chr.2 explained 26.97% of the phenotypic variation with a LOD score of 11.93, and qBRR7 on Chr.7 accounted for 27.06% of the phenotypic variation with a LOD score of 11.83. The additive effect of qBRR2 was positive (14.97); however, qBRR7 had the opposite effect (−11.99). The high-density linkage map and the major QTLs qBRR2 and qBRR7 provide new important information for disease resistance gene discovery and utilization in genetic improvement.