Assessment of molecular markers and marker-assisted selection for drought tolerance in barley(Hordeum vulgare L.)

This review updates the present status of the field of molecular markers and marker-assisted selection(MAS),using the example of drought tolerance in barley.The accuracy of selected quantitative trait loci(QTLs),candidate genes and suggested markers was assessed in the barley genome cv.Morex.Six common strategies are described for molecular marker development,candidate gene identification and verification,and their possible applications in MAS to improve the grain yield and yield components in barley under drought stress.These strategies are based on the following five principles:(1)Molecular markers are designated as genomic‘tags’,and their‘prediction’is strongly dependent on their distance from a candidate gene on genetic or physical maps;(2)plants react differently under favourable and stressful conditions or depending on their stage of development;(3)each candidate gene must be verified by confirming its expression in the relevant conditions,e.g.,drought;(4)the molecular marker identified must be validated for MAS for tolerance to drought stress and improved grain yield;and(5)the small number of molecular markers realized for MAS in breeding,from among the many studies targeting candidate genes,can be explained by the complex nature of drought stress,and multiple stress-responsive genes in each barley genotype that are expressed differentially depending on many other factors.

Mapping Quantitative Trait Loci Controlling Endosperm Traits with Molecular Marker

Based on the genetic models for triploid endosperm traits and on the methods for mapping diploid quantitative traits loci (QTLs), the genetic constitutions, components of means and genetic variances of QTL controlling endosperm traits under flanking marker genotypes of different generations were presented. From these results, a multiple linear regression method for mapping QTL underlying endosperm traits in cereals was proposed, which used the means of endosperm traits under flanking marker genotypes as a dependent variable, the coefficient of additive effect (d) and dominance effect (h1 and/or h2) of a putative QTL in a given interval as independent variables. This method can work at any position in a genome covered by markers and increase the estimation precision of QTL location and their effects by eliminating the interference of other relative QTLs. This method can also be easily used in other uneven data such as markers and quantitative traits detected or measured in plants and tissues different either in generations or at chromosomal ploidy levels, and in endosperm traits controlled by complicated genetic models considering the effects produced by genotypes of both maternal plants and seeds on them.

小麦茎基腐病抗性位点研究进展

对小麦茎基腐病(FCR)抗性位点研究进行综述。目前,在小麦所有的21条染色体上定位了140个抗性QTL,含10个抗性基因,虽然报道的抗性位点很多,但主效位点不明确,严重影响小麦FCR抗性改良进程。分析了小麦FCR抗性位点研究中的主要问题,提出了统一抗性鉴定标准、增强成株期抗性研究等建议,以明确抗FCR主效位点/基因并开发分子标记,通过分子标记辅助选择促进FCR抗性改良进程。

基于简化基因组测序宽鳍鱲的微卫星分子标记及遗传多样性分析

【目的】建立基于简化基因组测序(2b-RAD)宽鳍鱲(Zacco platypus)的微卫星分子标记,并进行遗传多样性分析,为有效开展宽鳍鱲品种间遗传多样性分析和分子育种,挖掘和利用宽鳍鱲种质资源提供参考。【方法】利用限制性内切酶EcoRⅠ打断基因组DNA,每个样本分别进行物理打碎,选取300~700 bp插入片段文库,利用Illumina HiSeq PE150测序平台进行双末端(Paired-End)测序获得海量遗传多态性标签序列。【结果】2b-RAD筛选宽鳍鱲的微卫星位点得到两端各留100 bp作为引物的SSR数量为41018个,带有引物片段的SSR数量为1522个,片段引物的设计率为37.1%,构建的文库质量较高,测序深度达高准确度下的分型标准。在筛选的64个微卫星位点中,有14个位点(21.88%)具有多态性,由于存在无效等位基因,有3个位点(Zpla08、Zpla09和Zpla10)显著偏离Hardy-Weinberg平衡(P<0.05),因此选用其中的11个微卫星位点分析宽鳍鱲群体遗传多样性。基于11个微卫星DNA位点的分析表明,宽鳍鱲7个群体的平均等位基因数(N_(A))为3.66,平均Shannon;s信息指数(I)为0.689,平均观测杂合度(H_(O))为0.315,平均期望杂合度(H_(E))为0.354,平均多态信息含量(PIC)为0.409。【结论】基于简化基因组测序(2b-RAD)开发宽鳍鱲的微卫星分子标记可用于评估野生宽鳍鱲种群的遗传多样性、遗传结构和物种鉴定。宽鳍鱲基因组DNA 14个微卫星位点中有11个微卫星位点具有中高多态性,3个位点(Zpla08、Zpla09、Zpla10)偏离Hardy-Weinberg平衡,在群体遗传研究中应慎用。

Analysis of SSR loci and development of SSR primers in Eucalyptus

In this study,28,691 genome sequences and16,566 expressed sequence tags(ESTs) of Eucalyptus were derived from the Gen Bank database.A total of 2292 SSR loci were sought out from 1785 effective sequences.Through analyses of SSR loci information,the SSR motif length was negatively correlated with the abundance of the SSRs.In the EST sequences of Eucalyptus,triplet repeat motifs were the most abundant,and dinucleotide repeats motifs had the highest frequencies.Subsequently,395 pairs of primers were designed based on the SSR loci.Using optimized SSR-PCR conditions,340 pairs of primers were successfully screened,with a success rate of 86.1%.By construction of a maximum likelihood phylogenetic tree of six eucalypt species,represented by five species of the genus Eucalyptus and one of the genus Corymbia,the genetic relationships of Eucalyptus urophylla and E.camaldulensis suggested by this tree was found to differ from that suggested by traditional morphological taxonomy.The results provide insights for evaluating geneticdiversity of Eucalyptus and analysis of Eucalyptus phylogenetics using SSR markers.

基于改进ML-KNN算法的文本分类研究

由于传统ML-KNN算法数据集中每个特征具有相同权重,与事实上的不同特征具有不同权重相违背,故提出对ML-KNN算法的改进,用ML-KNN来构建分类模型进行分类。为验证该算法的分类效果,选取算法常用的衡量标准与其他两种算法比较,结果表明由改进ML-KNN算法构造的模型要优于其他两种算法,能有效表达多领域数据集分类问题,算法效果更好。

Detecting mislabeling and identifying unique progeny in Acacia mapping population using SNP markers

Acacia hybrids offer a great potential for paper industry in Southeast Asia due to their fast growth and ability to grow on abandoned or marginal lands. Breeding Acacia hybrids with desirable traits can be achieved through marker assisted selection(MAS) breeding. To develop a MAS program requires development of linkage maps and QTL analysis. Two mapping populations were developed through interspecific hybridization for linkage mapping and QTL analysis. All seeds per pod were cultured initially to improve hybrid yield as quality and density of linkage mapping is affected by the size of the mapping population. Progenies from two mapping populations were field planted for phenotypic and genotypic evaluation at three locations in Malaysia,(1) Forest Research Institute Malaysia field station at Segamat, Johor,(2) Borneo Tree Seeds and Seedlings Supplies Sdn, Bhd.(BTS) field trial site at Bintulu, Sarawak, and(3) Asiaprima RCF field trial site at Lancang, Pahang. During field planting, mislabeling was reported at Segamat, Johor, and a similar problem was suspected for Bintulu, Sarawak. Early screening with two isozymes effectively selected hybrid progenies, and these hybrids were subsequently further confirmed by using species-specific SNPs. During field planting, clonal mislabeling was reported and later confirmed by using a small set of STMS markers. A large set of SNPs were also used to screen all ramets in both populations. A total of 65.36% mislabeled ramets were encountered in the wood density population and 60.34% in the fibre length mapping population. No interpopulation pollen contamination was detected because all ramets found their match within the same population in question.However, mislabeling was detected among ramets of the same population. Mislabeled individuals were identified and grouped as they originated from 93 pods for wood density and 53 pods for fibre length mapping populations.On average 2 meiotically unique seeds per pod(179 seeds/93 pods) for wood density and 3 meiotically unique seeds per pod(174 seeds/53 pods) for fibre length mapping population were found. A single step statistical method was used to evaluate the most informative set of SNPs that could subsequently be used for routine checks for mislabeling in multi-location field trials and for labelling superior clones to protect breeder’s rights. A preliminary set of SNPs with a high degree of informativeness was selected for the mislabeling analysis in conjunction with an assignment test. Two subsets were successfully identified,i.e., 51 SNPs for wood density and 64 SNPs for fibre length mapping populations to identify all mislabeled ramets which had been previously identified. Mislabeling seems to be a common problem due to the complexity involved in the production of mapping populations. Therefore, checking for mislabeling is imperative for breeding activities and for analyses such as linkage mapping in which a correlation between genotypic and phenotypic data is determined.

Rice molecular markers and genetic mapping:Current status and prospects

Dramatic changes in climatic conditions that supplement the biotic and abiotic stresses pose severe threat to the sustainable rice production and have made it a difficult task for rice molecular breeders to enhance production and productivity under these stress factors. The main focus of rice molecular breeders is to understand the fundamentals of molecular pathways involved in complex agronomic traits to increase the yield. The availability of complete rice genome sequence and recent improvements in rice genomics research has made it possible to detect and map accurately a large number of genes by using linkage to DNA markers. Linkage mapping is an effective approach to identify the genetic markers which are co-segregating with target traits within the family. The ideas of genetic diversity, quantitative trait locus（QTL） mapping, and marker-assisted selection（MAS） are evolving into more efficient concepts of linkage disequilibrium（LD） also called association mapping and genomic selection（GS）, respectively. The use of cost-effective DNA markers derived from the fine mapped position of the genes for important agronomic traits will provide opportunities for breeders to develop high-yielding, stress-resistant, and better quality rice cultivars. Here we focus on the progress of molecular marker technologies, their application in genetic mapping and evolution of association mapping techniques in rice.

Physio-Biochemical and Genetic Exploration for Submergence Tolerance in Rice (<i>Oryza sativa</i>L.) Landraces with Special References to <i>Sub</i>1 Loci

In the present study a group of four indigenous and less popular rice genotypes (Meghi, Panibhasha, Jabra and Sholey) reported by growers as submergence tolerant lines from flood prone areas of south Bengal were explored through study of nodal anatomy, physio-biochemical screening under submergence and genotyping with submergence tolerance linked rice microsatellite loci (RM loci). To identify the different allelic forms of different Sub1 compnents (Sub1A, Sub1B and Sub1C) among the studied lines, the genomic DNA of individual genotypes was amplified with three ethylene response factor like genes from Sub1 loci, located on rice chromosome 9. From the different physio-biochemical experiments performed in this investigation, it has been shown that Meghi and Jabra are the two probable potent genotypes which share common properties of both submergence tolerant and deep water nature whereas rest two genotypes (Sholey and Panibhasha) behave like typical deep water rice. The submergence tolerance property of Meghi was also confirmed from submergence tolerance linked SSR based genotyping by sharing with FR13A for some common alleles as reflected in fingerprint derived dendrogram. The rest of the genotypes shared a number of alleles and were included in a separate cluster. The common behaviour of Meghi and FR13A under submergence was also confirmed from genetic study of Sub1 loci through sharing of some common alleles for three Sub1 components (Sub1A, Sub1B and Sub1C loci). One SSR loci (RM 285) was identified as a potent molecular marker for submergence tolerance breeding programme involving these two selected rice lines (Meghi and Jabra) as donor plant through marker assisted selection.

Malate dehydrogenase in plants: Its genetics, structure, localization and use as a marker

This article is an overview of literature data on the structure, properties, functions and genetic control of the enzyme malate dehydrogenase (MDH) in plants. In most of the plant entities studied, this enzyme is highly polymorphic, which means that malate dehydrogenase has multiple molecular forms. It has been found that MDH polymorphism in each species is genetically determined by several loci with multiple alleles. A readily identifiable phenotypic manifestation and a high activity of malate dehydrogenase in diverse organs and tissues make this enzyme a reliable and convenient genetic marker, which can effectively be used in special, ecological and population genetics.

Identification of Co-dominant SSR Markers Associated with Genes Controlling α'- and cr-subunit-null β-conglycinin Phenotypes in Soybean (Glycine max (L.) Merr.)

Studies have shown that the three subunits of β-conglycinin are the main potential allergens of soybean sensitive patients.And β-conglycinin has adverse effects on nutrition and food processing.So solation and production of lines with lowerβ-conglycinin content has been the focus of recent soybean breeding projects.Soybean lines with deficiency in one or all subunits of β-congIycinin have been obtained.An effective and rapid system to identify such mutations will facilitate genetic manipulation of the β-conglycinin subunit composition.Here,two segregating F2 populations were developed from crosses between Cgy-1/cgy-1(CC),anα'-lacking line(△α'),and DongNong 47(DN47),a wild-type(Wt)Chinese soybean cultivar with normal globulin components,and Cgy-2/cgy-2(CB),an a-lacking line(△α),and DN47.These populations were used to estimate linkage among the egy-1(conferring α'-null)and cgy-2(α-null)loci and simple sequence repeat(SSR)markers.Seven SSR markers(Sat_038,Satt243,Sat_307,Sat_109,Sat_231,Sat_108 and Sat_190)were determined to co-scgregate with cgy-1,and six SSR markers(Satt650,Satt671,Sat_418,Sat_170,Satt292 and Sat_324)co-segregated with cgy-2.Linkage maps being composed of seven SSR markers and egy-1 locus,and six SSR markers and the cgy-2 locus were then constructed.It assigned that the egy-1 gene to chromosome 10 at a position between Sat_307 and Sat_231,and the cgy-2 gene to chromosome 20 at a position between Satt650 and Satt671.These markers should enable map-based cloning of the egy-1 and cgy-2 genes.For different subunit-deficiency types[α'-null,α-null and(α'+α)-null types],the two sets of SSR markers could also detect of polymorphism between three normal cultivars and seven related mutant lines.The identification of these markers is great significance to the molecular marker-assisted breeding of soybean/9-conglycinin subunits.

The Power of Integrative Approaches to Develop Markers for Maize Molecular Breeding

Recent advances in genomics and bioinformatics now offer real opportunities for dissecting complex traits into their component sub-traits, which will simplify the process of developing the tools necessary to

Genetic Map of Cotton with Molecular Markers

Cotton (Gossypium spp.) is the most important natural fiber in the world, and its seeds are also used as a food source. Breeding cotton for traits of interest, such as production and processing of fibers, will ensure that this natural product is as competitive as renewable synthetic fibers derived from petroleum. Thus, the mapping of the cotton genome for traits of interest may be the basis for its subsequent use in breeding programs. This work consists of a literature review, with the aim of bringing together works from different research groups working with the mapping of the cotton genome with molecular markers.

基于SSR标记的山茶属位点组合品种鉴别能力分析评价

【目的】筛选多态性高、通用性强等的SSR位点,评估筛选所得SSR位点的组合在山茶属品种中的鉴别能力;在保持最大鉴别能力的基础上,确定可用于山茶属品种鉴别所需的最少SSR位点的组合,提高品种鉴别效率。【方法】本研究以111份山茶属品种为材料,结合凝胶电泳、荧光毛细管电泳、位点多态性分析和位点组合品种鉴别力(VDP)分析等方法,筛选评价SSR位点并对SSR位点组合的品种鉴别力进行分析与评价。【结果】本研究筛选获得17个SSR位点,占筛选位点总数的30.9%,在111份山茶属品种中,共检测到166个等位基因,单位点基因型数量范围为6~35,均值为23.353个,观测等位基因数(Na)范围为3~15,均值为9.765个,多态性信息含量(PIC)的范围为0.447~0.835,均值为0.696,位点缺失率的范围为0~4.50%,均值为0.90%。当差异位点数=1时,17个SSR位点在全部品种(系)、红山茶组品种和金花茶组无性系中达到的R-VDP最大值均为1,可将SSR位点数量分别缩减到11、10、5个,分别占筛选所得位点数的64.7%,58.8%和29.4%;当差异位点数=2时,17个SSR位点在全部品种(系)、红山茶组品种和金花茶组无性系中达到的R-VDP最大值分别为0.964,0.952和1.000,可将SSR位点数量分别缩减到12、12、和7个,分别占筛选所得位点数的70.6%、70.6%和41.2%;当差异位点数=3时,17个SSR位点在全部品种(系)、红山茶组品种和金花茶组无性系中达到的R-VDP最大值分别为0.946,0.929,1.000,可将SSR位点数量分别缩减到8、7和8个,分别占筛选所得位点数的47.1%、41.2%和47.1%。当差异位点数≤6时,17个SSR位点组合对山茶属测试品种(系)群的RVDP最大值可保持相对稳定的水平。【结论】本研究筛选验证获得17个多态性高、通用性强等的SSR位点;这些位点的组合在山茶属品种(系)群中均表现出良好的品种鉴别能力;确定了山茶属不同类群品种(系)鉴别所需的最少SSR位点的组合,构建了一套山茶属SSR位点组合品种鉴别能力分析评价的体系,为山茶属品种鉴别及相关标准的制定等奠定了基础。

青海草原毛虫转录组分析及SSR位点开发

青海草原毛虫(Gynaephora qinghaiensis)是青藏高原牧区重要的害虫之一。本研究采用Illumina HiSeqTM2000高通量测序平台对青海草原毛虫成虫和4龄幼虫进行转录组测序,在此基础上筛选其微卫星(Single sequence reperts, SSR)位点并挖掘微卫星引物。本研究共获得63 335条unigenes,有12 597个微卫星位点分布于9 851条unigenes中。通过KOG,GO注释和KEGG通路数据库分析发现,unigenes注释主要集中于一般功能预测、细胞进程和碳水化合物代谢过程。SSR重复类型主要为单核苷酸和二核苷酸重复,(A/T)n是单核苷酸重复中最主要的基元类型,占总SSR位点的71.37%,(AC/GT)n为二核苷酸重复的优势基元,占总SSR位点数的6.06%,且SSR数量随重复次数增加而降低,重复次数类型随基元序列长度增长而减少。利用Primer 3软件设计出2 714对青海草原毛虫SSR引物,随机挑出25对引物进行PCR验证,有11对引物扩增出目的DNA片段。本研究基于转录组数据成功筛选出青海草原毛虫微卫星位点,为进一步研究其种群遗传学和发生动态提供数据支撑。

基于黄褐棉导入系群体定位抗黄萎病QTL

【目的】黄萎病(Verticillium wilt)是棉花生产中的最主要病害,且棉花黄萎病的致病机理尚不清楚。通过构建黄褐棉导入系群体定位棉花黄萎病抗性相关的数量性状位点(quantitative trait loci,QTL),为抗黄萎病分子标记开发和辅助育种提供参考。【方法】以陆地棉(Gossypium hirsutum)B0011为轮回亲本、黄褐棉(G.mustelinum)为供体亲本,构建有71个株系的BC_(5)S_(5)群体。利用2 839个单核苷酸多态性(single nucleotide polymorphism,SNP)标记结合表型值进行黄萎病抗性相关QTL定位。【结果】共检测到15个与黄萎病抗性相关的QTL,可解释4.21%~26.77%的表型变异。加性效应分析表明:其中6个QTL的有利等位基因来源于黄褐棉,9个QTL有利等位基因来自B0011。同时,qVW-A01-1、q VW-A02-2和qVW-A07-2在2个及以上环境中被检测到,表型变异解释率分别为15.56%~16.56%、11.95%~24.62%和13.22%~16.73%。利用BC5S5群体黄萎病抗性的最佳线性无偏预测值(best linear unbiased prediction,BLUP)进行QTL定位,共检测到5个QTL,其中qVW-A01-1B、q VW-A02-1B分别与加性效应分析稳定检测到的qVW-A01-1、qVW-A02-2物理位置一致,分别解释23.67%和17.90%的表型变异。【结论】本研究发现2个稳定的QTL即qVW-A01-1和qVW-A02-2,可为抗黄萎病分子标记辅助选择育种及候选基因功能鉴定奠定基础。

QTL mapping for pre-harvest sprouting in a recombinant inbred line population of elite wheat varieties Zhongmai 578 and Jimai 22

Pre-harvest sprouting(PHS)is one of the serious global issues in wheat production.Identification of quantitative trait loci(QTL)and closely-linked markers is greatly helpful for wheat improvement.In the present study,a recombinant inbred line(RIL)population derived from the cross of Zhongmai 578(ZM578)/Jimai 22(JM22)and parents were phenotyped in five environments and genotyped by the wheat 50 K single-nucleotide polymorphism(SNP)array.Two QTL of germination index(GI),QGI.caas-3A and QGI.caas-5A,were detected,explaining 4.33%–5.58%and 4.43%–8.02%of the phenotypic variances,respectively.The resistant effect of QGI.caas-3A was contributed by JM22,whereas that of QGI.caas.5A was from ZM578.The two QTL did not correspond to any previously identified genes or genetic loci for PHSrelated traits according to their locations in the Chinese Spring reference genome,indicating that they are likely to be new loci for PHS resistance.Four kompetitive allele-specific PCR(KASP)markers K_AX-109605367and K_AX-179559687 flanking QGI.caas-3A,and K_AX-111258240 and K_AX-109402944flanking QGI.caas-5A,were developed and validated in a natural population of 100 wheat cultivars.The distribution frequency of resistance alleles at Qphs.caas-3A and Qphs.caas-5A loci were 82.7%and57.1%,respectively,in the natural population.These findings provide new QTL and tightly linked KASP markers for improvement of PHS resistance in wheat.

运用近等基因系(NIL)、AFLP、RFLP和SCAR标记对玉米S组育性恢复基因(Rf_3)的研究

以１对近等基因系（ＮＩＬ）及其回交群体（ＢＣ１）为材料，采用ＢＳＡ法，利用ＡＦＬＰ技术，筛选与Ｒｆ３基因连锁的分子标记。在筛选的１２８个ＡＦＬＰ引物组合中，有２个能在ＮＩＬ及其可育池、不育池间扩增出多态性条带ＲＲ６和ＲＲ７。１００个ＢＣ１个体验证结果表明，ＡＦＬＰ标记ＲＲ６扩增产物中仅出现２个重组体，重组率２％，由此估测ＲＲ６距Ｒｆ６基因约２．０ｃＭ。并成功地将此标记转化为 ＳＣＡＲ标记，进行了ＮＩＬ和 ＢＣ１个体的特异性扩增。在来自综 ３ × Ｐ１３８的Ｆ２：３的群体上经ＲＦＬＰ分析后，将ＲＲ６定位于第二染色体的长臂上。不仅为辅助育种奠定了基础，而且为克隆Ｒｆ３基因提供了有益的信息。

两种供氮水平下水稻穗长QTLs的检测

在水稻 (IR6 4× Azucena) DH群体中应用分子标记连锁图 ,对不同供氮水平下的穗长性状进行 QTL区间作图分析。在高氮水平下检测到控制穗长的 QTL s 2个 ,分别位于第 1和第 4染色体上 ;在低氮水平下检测到 5个穗长 QTL s,其中 3个分别位于第 1、4和 6染色体上 ,2个位于第 3染色体的不同部位 ,说明穗长 QTL与供氮水平之间可能存在交互效应。基于这种互作效应及研究中观察到的两供试亲本对低氮胁迫的耐性差异 ,推测仅在低氮水平下检测到的穗长 QTLs可能跟水稻对低氮胁迫耐性有一定的关联。

水产动物分子育种研究进展

本文介绍了水产动物分子生物学尤其是分子标记的发展,比较了共显性标记和显性标记用于水产动物育种研究的优缺点。介绍了开展分子标记育种研究的基础研究——连锁图谱和经济性状QTL定位研究进展。同时还对国内外已开展的标记辅助的家系育种和QTL研究为基础的性状选择育种等研究进行了综述。最后,着重探讨了分子育种理论及作者建立的有关鲤的3种分子标记指导的育种技术。通过分子育种技术具有不同发展阶段的理论分析以及分子育种的实例介绍,旨在加快分子选择手段与常规表型选择的结合,从而推进分子育种技术在中国主要水产养殖动物育种研究中的应用。