Genome-Wide Analysis for Yield-Related Agronomic and Biochemical Traits of Chinese and Bangladeshi Grass Pea Genotypes Using SSR Markers

Grass pea(Lathyrus sativus L.)is an imperative food crop cultured in dryland agricultural ecology.It is a vital source of dietary protein to millions of populaces living in low-income countries in South-East Asia and Africa.This study highlights the improvement of genomic properties and their application in marker-trait relationships for 17 yield-related characters in 400 grass pea genotypes from China and Bangladesh.These characters were assessed via 56 polymorphic markers using general linear model(GLM)(P+G+Q)and mixed linear model(MLM)(P+G+Q+K)in the tassel software based on the linkage disequilibrium and population structure analysis.Population structure analysis showed two major groups and one admixed group in the populace.Statistically significant loci pairs of linkage disequilibrium(LD)mean value(D′)was 0.479.A total of 99 and 61 marker-trait associations in GLM and MLM models allied to the 17 traits were accepted at a 5%level of significance.Among these markers,21 markers were associated with more than one trait;12 marker-trait associations passed the Bonferroni correction threshold.Both models found six markers C41936,C39067,C34100,C47146,C47638,and C43047 significantly associated with days to maturity,flower color,plant height,and seed per pod were detected in the Hebei and Liaoyang location(p≤0.01),and the interpretation rate(R^(2)value)11.2%to 43.6%.Conferring to the consequences,the association analysis methodology may operative system for quantitative,qualitative,and biochemical traits related to gene position mapping and support breeders in improving novel approaches for advancing the grass pea quality.

Genetic Structure of Malus sieversii Population from Xinjiang,China,Revealed by SSR Markers

One hundred and nine Malus sieversii accessions from four geographical populations located at Kuerdening in Mohe town, Gongliu County, Jiaowutuohai, in Xinyuan County, Daxigou in Houcheng County of Ily State, and Baerluke Mountain in Yumin County of Tacheng State, Xinjiang Uygur Autonomous Region of China were studied by SSR markers. The purpose of the study was to determine the genetic structure and diversity in these eco-geographical populations with eight pair SSR primers of apple. The results indicated that： an average of 16 bands was detected in the four populations. The percentage of polymorphic bands in Gongliu population （89.06%） was the highest in the four populations. The average Nei＇s gene diversity index was 0.257 for all the loci. Totally, 128 polymorphic loci were detected and the percentage of polymorphic loci （P） was 100%, 88.28%, 84.83%, 87.50%, and 87.12%, respectively, at the species level and Gongliu, Xinyuan, Huocheng, and Yumin population levels. The Nei＇s gene diversity index （H = 0.2619） and Shannon＇s information index （1 = 0.4082） in the species level were higher than in the population level. The Nei＇s gene diversity index and Shannon＇s information index in the four populations were Gongliu 〉 Huocheng 〉 Xinyuan 〉 Yumin. Gongliu population and Xinyuan population were the highest in genetic identity and the closest in genetic distance. Gene flow between the populations was 7.265 based on genetic differentiation coefficient （GST = 0.064）. The UPGMA cluster analysis indicated that the genetic relationships between the Gongliu and Xinyuan population were the closest, and the Yumin population were the farthest with the other three populations. The UPGMA cluster analysis indicated that the four geographical populations located in Gongliu, Xinyuan, Huocheng, and Yumin were relatively independent populations. Concurrently, there was also mild gene exchange between the populations. On the basis of the study of population genetic structure and the highest genetic diversity, Gongliu population should be given a high priority consideration in Malus sieversii population＇s in situ germplasm conservation.

Assessment of Genetic Diversity of Yunnan,Tibetan,and Xinjiang Wheat Using SSR Markers

A total of 206 SSR （Simple Sequence Repeats） primer pairs were used to detect genetic diversity in 52 accessions of three unique wheat varieties of western China. A total of 488, 472, and 308 allelic variants were detected in 31 Yunnan, 15 Tibetan and 6 Xinjiang wheat accessions with an average of PIC values 0.2764, 0.3082, and 0.1944, respectively. Substantial differences in allelic polymorphisms were detected by SSR markers in all the 21 chromosomes, the 7 homoeologous groups, and the three genomes （A, B, and D） in Yunnan, Tibetan, and Xinjiang wheat. The highest and lowest allelic polymorphisms in all the 21 chromosomes were observed in 3B and 1D chromosomes, respectively. The lowest and highest allelic polymorphisms among the seven homoeologous groups was observed in 6 and 3 homoeologous groups, respectively. Among the three genomes, B genome showed the highest, A the intermediate, and D the lowest allelic polymorphism. The genetic distance （GD） indexes within Yunnan, Tibetan, and Xinjiang wheat, and between different wheat types were calculated. The GD value was found to be much higher within Yunnan and Tibetan wheat than within Xinjiang wheat, but the GD value between Yunnan and Tibetan wheat was lower than those between Yunnan and Xinjiang wheat, and between Tibetan and Xinjiang wheat. The cluster analysis indicated a closer relationship between Yunnan and Tibetan wheat than that between Yunnan and Xinjiang wheat or between Tibetan and Xinjiang wheat.

Identification of the Parentage of Corn Variety Using SSR Markers

[Objective] The study was to explore the molecular interpretation standards on parentage in the seeds of corn variety. [Method] With 16 hybrids and their parents and 202 inbred elites as materials for screening primers, the artificial groups of two standard diad and two standard triad were respectively established as the verification materials. Genomic DNA of seedlings was extracted by using CTAB method. 137 pairs of SSR primers were selected for SSR amplification and product detection, which was used for the parentage identification of maize varieties. [ Result] Twenty pairs of corn primers with high polymorphism information content （ PIC value）, clearly amplified bands and good reproducibility were screened from 137 pairs of corn SSR primers tested. The identification results of using SSR molecular were consistent with the actual situation. [ Conclusion] It is feasible to identify the parentage of maize variety using SSR markers.

QTL Analysis of Grain Storage Durability for Maize Under Controlled Deterioration Conditions Using SSR Markers

[Objective] This study aimed to analyze the quantitative trait loci （QTLs） of grain storage durability for maize under controlled deterioration conditions by using SSR markers. [Method] The recombinant inbred line population F 2 derived from the cross of shen 137/02-50 was used to do the simple sequence repeats （SSRs） and QTL analysis. [Result] The 2 parents showed good polymorphisms. Three loci were detected on chromosome 1, chromosome 6 and chromosome 9, accounting for 41.2% of total phenotypic. [Conclusion] This study provided theoretical basis for using molecular marker to assist the breeding of storage durable maize varieties.

Genetic Diversity of SSR Markers in Cultivated Hordeum vulgare L.in Qinghai Province

[Objective]The aim was to analyze genetic diversity of SSR markers in Hordeum vulgare L.in Qinghai Province and lay a foundation for screening and protecting some excellent H.vulgare cultivars.[Method]SSR markers were used to evaluate the genetic diversity of 42 cultivated H.vulgare from Qinghai Province.[Result]42 H.vulgare showed polymorphism in 7 SSR markers locus.A total of 24 alleles were identified,and the number of alleles per locus ranged from 1 to 6,with an average of 3.0.According to SSR markers polymorphism,42 H.vulgare could be divided into 4 groups,namely I,II,III and IV.[Result]The study indicated that cultivated H.vulgare from Qinghai Province is rich in genetic diversity,which will provide reference for selecting parent of H.vulgare breeding.

Purity Identification of Xinshikui 6 Using SSR Marker Technique

[Objective] SSR molecular marker technique was used to determine the purity of sunflower seed with the aim to provide accurate, convenient method for the identification of the purity of hybrid seeds in production and processing. [Method] With the DNA of Xinshikui 6 and its parents as template, about 100 pairs of SSR molecular markers were screened after DNA extraction, PCR amplification and electrophoresis production. [Results] SSR polymorphic primer marker 532 produced a specific band of 469 bp in the female parent, and a specific band of 451 bp in the male parent; primer marker 574 produced a specific band of 364 bp in the female parent, and a specific band of 384 bp in the male parent. The indoor molecular purity identification and field purity identification were consistent with each other. The primer marker 532 and 574 could be obtained from the SSR molecular marker method to distinguish the male parent, female parent and hybrid of Xinshikui 6, and both of the 2 primer markers can effectively identify the purities of the hybrid seeds of Xinshikui 6, as well as the authenticity of the seeds. [Conclusion] The proposed method was simple, fast, accurate to operate with the advantages of high reproducibility, and it had become the major method in the identification of sunflower varieties.

A Method for Rapid Identification of Ningza 11 Seeds Purity with SSR Markers

[Objective] This study aimed to establish a method for rapid identification of Ningza 11 seeds purity with SSR markers. [Method] Taking Ningza 11 hybrid seeds as experimental materials, a method for rapid identification of hybrid rape-seeds was established with SSR molecular markers; meanwhile, the test seeds were planted in the field for comparison and verification. [Result] A method for rapid identification of Ningza 11 seeds purity with SSR molecular markers was estab-lished： DNA from seeds germinated in the night was extracted by alkaline lysis method; the PCR amplification was performed for 2 h, and electrophoresis for 1.5 h, and a silver staining for 10 minutes. It took less than one day to from obtaining sampling seeds to obtaining the purity identification result, so a skil ed professional can complete the detection of at least 6 ×96 = 576 seeds per weekday. By using this set of detection system, the measured purity of seeds from nine samples was extremely significantly positively correlated to the actual purity identified in the field test, with a correlation coefficient of up to 0.984 （P〈0.01）. [Conclusion] This SSR-PCR molecular identification system can be applied for rapid and accurate identifi-cation of Ningza 11 hybrid seeds.

不同品种草莓花器与花粉特征变异及SSR遗传多样性研究

草莓花器特征和花粉形态在品种之间存在一定的差异,这些差异对草莓品种的分类具有指导意义。为研究南方地区主栽的10个草莓品种花器、花粉及其与品种分类的关系,对其花部特征、花粉形态进行比较,并对13个花粉和花器性状进行相关性和主成分分析,同时进行了品种间简单重复序列(SSR)标记遗传多样性分析。花器特征显示,甜查理的花瓣长、花瓣宽和花萼宽的特征值显著高于其他9个品种,其次是宁玉和妙香7号,红颊、红玉和越心的花瓣长、花瓣宽、花冠径则明显低于其他品种。红颊、白雪公主、妙香7号和宁玉在极轴长、花粉大小、花粉形状、萌发沟长等花粉特性上高于其他品种。主成分分析结果显示,4个主成分累积贡献率为85.96%,其中花瓣长、花瓣宽和花萼宽的大小特征在第1主成分中具有较大载荷值。SSR分析结果显示,10个草莓栽培品种间的相似系数范围为0.47~0.85,在相似系数0.65处10个草莓品种分为两大类,红颊、章姬、红玉、粉玉1号、越心、白雪公主6个品种聚为一类,甜查理、越秀、妙香7号、宁玉4个品种聚为一类。结合草莓花器特征、花粉特性及其主成分分析与SSR标记聚类结果和系谱情况,花粉形态特性在草莓品种之间的差异不明显,花器特征的花瓣长、花瓣宽、花萼宽适用于品种分类,对指导育种亲本选择有参考意义。

60份辣椒骨干亲本的SSR遗传多样性分析及指纹图谱构建

为解析江西省辣椒育种骨干亲本材料的遗传多样性并构建其DNA指纹图谱,本研究利用毛细管电泳和SSR分子标记技术对江西60份辣椒种质材料进行位点检测。结果表明,49对SSR引物共检测到202个等位基因,平均4.122个,平均有效等位基因为2.172个,平均香农信息指数为0.850,平均多态信息含量为0.414,说明60份供试材料具有较为丰富的遗传多样性;平均期望杂合度0.475大于观测杂合度0.126,表明多代自交导致材料纯合度较高。聚类分析、群体结构分析和主坐标分析结果一致,即江西辣椒育种亲本材料以南方地区和小果型尖椒材料为主,遗传背景狭窄且保持较纯的血统。此外,本研究根据多位点匹配分析确定了PM1、CAMS-855、CO911525S、PM22、Hpms E015和Hpms1214为核心引物,并利用这6对核心引物构建了60份辣椒育种亲本的指纹图谱,为江西辣椒资源的鉴定和保护提供了有力支撑,为分子辅助育种中亲本的选择提供理论依据。

白三叶杂交F1代群体的表型鉴定及SSR分析

早期快速鉴定白三叶(Trifolium repens L.)F1杂交子代,获取真杂种对白三叶优良品系的培育和遗传学研究具有重要的意义。本文针对两个白三叶亲本及55个杂交F1的6个表型性状进行分析,绘制聚类热图,再选用3对特异性SSR引物鉴定杂交F1代的真实性。结果表明,在聚类热图中57个株系可以根据亲本的表型性状划分为父本型和母本型。筛选出的19对SSR引物共扩增共得到清晰可辨条带281条,多态性条带137条,多态位点占比为48.41%,每个SSR位点的PIC在18.49%~43.58%之间,平均值为34.73%。其中3对特异性引物在55个杂交后代中共鉴定出53个真杂种,真杂种比率为96.36%,与UPGMA聚类分析结果基本一致。19对SSR分子标记引物具有较高的多态性,可直接用于白三叶遗传多样性分析、杂种鉴定等相关研究,鉴定到的白三叶F1代真杂种为后续育种工作奠定重要基础。

浙江红花油茶×广宁红花油茶杂交子代的表型性状及其SSR分子鉴定

浙江红花油茶(Camellia chekiangoleosa)种仁含油率和油酸含量高,广宁红花油茶(C.semiserrata)具有较强的生长势和抗性。为了利用浙江红花油茶和广宁红花油茶的优点,培育优良种质材料,该研究对浙江红花油茶与广宁红花油茶的45个F_(1)杂交子代进行表型性状分析,以掌握杂交子代的表型性状情况,同时利用SSR标记对其进行杂种真伪鉴定,并筛选可用于油茶杂交子代鉴定的SSR标记。结果表明:(1)浙江红花油茶×广宁红花油茶的F_(1)子代表现为树形高大、生长迅速且其叶脉、萼片、柱头均倾向于父本广宁红花油茶的性状,而花和叶片形态等性状与母本浙江红花油茶接近,叶片颜色与大小等特征介于双亲特征之间。(2)从32个SSR标记中筛选出了8个可区分双亲且能明确杂交子代来源的完全互补型标记,用于开展杂交子代鉴定,其中7个标记杂种鉴定效率高达100%,1个标记杂种鉴定效率为55.56%;8个标记相互补充鉴定出45个杂交子代全是真杂种。(3)8个SSR标记对杂交子代进行的鉴定能力验证表明,利用这些SSR标记鉴定油茶杂交子代是可行的。该研究结果为油茶物种间的杂交育种提供了参考,同时也为后续油茶物种间的杂交子代SSR标记鉴定提供了依据。

基于SSR的云南野生猕猴桃种质资源亲缘关系分析

为探讨云南猕猴桃属种质资源的亲缘关系,采用SSR分子标记方法对70份云南野生猕猴桃种质资源的亲缘关系进行分析。UPGMA聚类分析结果表明,70份猕猴桃种质资源在遗传相似系数0.17水平上可分为4个类群,类群Ⅰ共6份材料,包括滇东南麻栗坡的中越猕猴桃MLP001和MLP010、西畴的中越猕猴桃TSH-1和TSH-4,以及滇南屏边的中越猕猴桃PB001和PB005;类群Ⅱ共31份材料,主要包括滇东北的18份材料,滇西北的3份紫果猕猴桃TC-8、TC-1、TC-10,以及5份贡山猕猴桃CJ-11、CJ-13、CJ-7、CJ-3、CJ-2;类群Ⅲ共14份材料,主要包括滇东北的10份美味猕猴桃和中华猕猴桃系列材料,滇东的JZS-5、JZS-9及JZS-7,以及滇东南西畴的黄毛猕猴桃XPS-1;类群Ⅳ共19份材料,主要包括滇东北的16份材料,滇东南西畴的京梨猕猴桃XCFD-1及革叶猕猴桃XCFD-3,果实无被毛且果皮带斑点,果实较小,与其他类群具有较远的亲缘关系。SSR分子标记反映了云南猕猴桃属70份种质资源间的遗传亲缘关系,其中一些材料按种类、资源分布地及形态学性状特征形成明显有规律的聚类关系。

黄精转录组SSR分子标记开发及种质遗传多样性分析

为丰富可用于黄精属鉴定的SSR分子标记,开发可用于黄精属(Polygonatum sp.)种质资源鉴定与遗传多样性分析的简单重复序列(simple sequence repeat,SSR)分子标记,为药材黄精正品和伪品的鉴定及遗传多样性分析提供基础。通过对3种黄精属植物进行转录组测序,从60836967个Unigenes鉴定出19630个包含1~6个碱基重复类型的SSR位点,设计和筛选获得了9对SSR有效性引物,并对来源于5个省份共计31份种质资源进行鉴定与遗传多样性分析。9对SSR引物不仅能够将长梗黄精与其他3种药材黄精划分开,还能够较好地区分不同基源黄精(滇黄精、黄精、多花黄精),并且31份资源遗传相似系数范围为0.0769~0.75。此外,构建了31份黄精属种质资源的DNA指纹图谱数据库,开发了相对应的指纹图谱。本研究结果为黄精属种质资源的分类鉴定、遗传多样性分析和品种选育提供了可用的分子标记以及参考依据。

茄子果皮转录组SSR位点分析及标记开发

以茄子(Solanum melongena L.)果皮样品转录组的36568条转录本为试材,采用MISA(MicroSAtellite)软件进行SSR位点搜索,研究SSR位点的分布频率和特征,同时利用Primer 3.0软件对检索出的SSR位点设计引物,验证引物有效性,以期为茄子的种质鉴定、亲缘关系分析、分子标记辅助育种及遗传图谱构建等提供参考依据。结果表明:在转录本中共检测出9775个SSR位点,分布于6289条转录本上,平均5.49 kb存在1个SSR位点,发生频率26.73%。SSR位点特征分析显示,单核苷酸为优势重复基元类型,占SSR位点总数的47.67%;A/T基元是数量最多的重复基元,占总数的46.13%;97.91%的重复基元的重复次数在5~25,92.21%的SSR重复序列长度分布在10~30 bp。对所有SSR位点进行引物设计,共获得6619对EST-SSR引物。随机选择27对EST-SSR引物在12个茄子自交系中进行PCR扩增,引物有效性为92.59%,多态率为33.33%。

基于转录组测序的油茶SSR、SNP和InDel位点特征分析

基于油茶转录组测序数据,利用MISA和GATK3软件对SSR、SNP和InDel位点进行搜索。结果表明:在169652条unigene序列中共发现92228个SSR位点,出现频率54.37%,平均分布距离1.61 kb。油茶转录组SSR位点中,单核苷酸和二核苷酸是主要的重复类型,A/T和AG/CT是主要的重复基元类型,基元重复次数主要集中在5~11次,基序长度主要集中在12~20 bp。共得到1912501个SNP位点,转换类型SNP数量多于颠换类型SNP数量,转换类型中A/G数量最多,而颠换类型中A/T数量最多。共筛选出298984个InDel位点。油茶转录组SSR、SNP和InDel位点数量多、类型丰富,能够为油茶分子标记开发、种质资源评价、亲缘关系鉴定等研究提供支撑。

Bulked Segregant Analysis to Detect QTL Related to Heat Tolerance in Rice(Oryza sativa L.) Using SSR Markers

The study was undertaken to assess the genetic effect of quantitative trait loci （QTLs） conferring heat tolerance at flowering stage in rice. A population consisting of 279 F2 individuals from the cross between 996, a heat tolerant cultivar and 4628, a heat-sensitive cultivar, was analyzed for their segregation pattern of the difference of seed set rate under optimal temperature condition and high temperature condition. The difference of seed set rate under optimal temperature condition and high temperature condition showed normal distribution, indicating the polygenic control over the trait. To identify main effect of QTL for heat tolerance, the parents were surveyed with 200 primer pairs of simple sequence repeats （SSR）. The parental survey revealed 30% polymorphism between parents. In order to detect the main QTL association with heat tolerance, a strategy of combining the DNA pooling from selected segregants and genotyping was adopted. The association of putative markers identified based on DNA pooling from selected segregants was established by single marker analysis （SMA）. The results of SMA revealed that SSR markers, RM3735 on chromosome 4 and RM3586 on chromosome 3 showed significant association with heat tolerance respectively, accounted for 17 and 3% of the total variation respectively. The heat tolerance during flowering stage in rice was controlled by multiple gene. The SSR markers, RM3735 on chromosome 4 and RM3586 on chromosome 3 showed significant association with heat tolerance respectively, accounted for 17 and 3% of the total variation respectively. The two genetic loci, especially for RM3735 on chromosome 4, can be used in marker-assistant-selected method in heat tolerance breeding in rice.

基于SSR分子标记吉安地区的水稻亲缘关系鉴定分析

稻种亲缘关系分析是生产优质杂交水稻的必要条件。本研究选取均匀分布于水稻基因组12条染色体的SSR分子标记,对8份水稻样品进行SSR分子标记分析,通过聚类分析法将其归类并计算得出遗传相似系数后,对8份水稻样品的亲缘关系进行了鉴定。结果表明:所选取的8份水稻样品中,其相似系数为0.65~1.00,在遗传相似系数0.65处,8份水稻样品聚为两大类群,其中5号‘赣迟A04’单独为一类,其他水稻样品聚为一类。在遗传相似系数0.80处,可划分为三大类群:第一类囊括了6种样品,表明这6个品种亲缘关系较近,其中6号‘赣早A02’与7号‘吉安早A07’疑似同一样品,3号‘井冈山迟A03’与4号‘井冈山早A05’的亲缘关系最近,而6号、7号同时与8号‘吉安迟A09’保持有最近的亲缘关系;第二类只含有2号‘迟-2’样品株;第三类则仅有5号稻株‘赣迟A04’,说明其与其他7份水稻样品的亲缘关系最远。本研究结果为杂交水稻的应用提供重要信息。

Identification and Purity Test of Super Hybrid Rice with SSR Molecular Markers

Five super hybrid rice combinations, i.e. HYS-1/R105, Pei'ai 64S/E32, Liangyoupeijiu (Pei'ai 64S/9311), 88S/0293, and J23A/Q611, and their parental lines were tested by means of SSR analysis. A total of 144 SSR primer pairs distributed on 12 rice chromosomes were used, out of which 47 detected polymorphism among the tested rice lines. Among all these primers, RM337 and RM154 produced polymorphic patterns in four or more of the tested experimental materials respectively, and they could distinguish among most rice genotypes tested. Twenty-four primer pairs, two on each rice chromosome, were selected to make a reference SSR marker-based fingerprinting for the rice lines. For most of the primer pairs, F1 hybrids mainly showed complementary pattern of both parents, which could be very useful to distinguish the F1 from its parental lines. In addition, 5 primer pairs were selected as special primer pairs for five hybrid rice combinations respectively. By combining the rapid, simple method on DNA extraction, it is suggested that SSR technique has wide prospective in variety authentication and purity identification.

Assessment of genetic variation in tomato (Solanum lycopersicum L.) inbred lines using SSR molecular markers

A study was conducted to determine the genetic diversity of 39 determinate and indeterminate tomato inbred lines collected from China, Japan, S. Korea, and USA. Using 35 SSR polymorphic markers, a total of 150 alleles were found with moderate levels of diversity, and a high number of unique alleles existing in these tomato lines. The mean number of alleles per locus was 4.3 and the average polymorphism information content （PIC） was 0.31. Unweighted Pair Group Method with Arithmetic Mean （UPGMA） clustering at genetic similarity value of 0.85 grouped the inbred lines into four groups, where one USA cultivar formed a separate and more distant cluster. The most similar inbred lines are from USA, both with determinate type, whereas the most different lines are from USA （Us-16） and Japan （Ja-2） with determinate and indeterminate growth habit, respectively. Clustering was consistent with the known information regarding geographical location and growth habit. The genetic distance information reported in this study might be used by breeders when planning future crosses among these inbred lines.