Single Nucleotide Polymorphism Molecular Markers Research Articles

Authenticity Identification of F1 Hybrid Offspring and Analysis of Genetic Diversity in Pineapple

Breeding is an effective method for the varietal development of pineapple. However, due to open pollination, it is necessary to conduct authentic identification of the hybrid offspring. In this study, we identified the authenticity of offspring and analyzed the genetic diversity within the offspring F1 hybrids resulting from crosses between ‘Josapine’ and ‘MD2’ by single nucleotide polymorphism (SNP) markers. From the resequencing data, 26 homozygous loci that differentiate between the parents have been identified. Then, genotyping was performed on both the parents and 36 offspring to select SNP markers that are suitable for authentic identification. The genotyping results revealed that 2 sets of SNP primers, namely SNP4010 and SNP22550, successfully identified 395 authentic hybrids out of 451 hybrid offspring. We randomly selected two true hybrids and four pseudohybrids for sequencing validation, and the results have shown that two true hybrids had double peaks with A/G, while pseudohybrids had single peaks with base A or G. Further study showed that the identification based on SNP molecular markers remained consistent with the morphological identification results in the field, with a true hybridization rate of 87.58%. K-means clustering and UPGMA tree analysis revealed that the hybrid offspring could be categorized into two groups. Among them, 68.5% of offspring aggregated with MD2, while 31.95% were grouped with Josapine. The successful application of SNP marker to identify pineapple F1 hybrid populations provides a theoretical foundation and practical reference for the future development of rapid SNP marker-based methods for pineapple hybrid authenticity and purity testing.

Genetic Diversity Analysis and Core Germplasm Construction of Rubus chingii Hu.

Rubus chingii Hu is the only species that is used for both edible and medicinal purposes among the 194 species of the genus Rubus in China. It is well known for its sweet and sour fresh fruits that are rich in vitamins and for its dried immature fruits that are used to treat kidney-related ailments. This study aims to evaluate genetic diversity and population structure and build a core germplasm repository of 132 R. chingii accessions from the provinces of Jiangxi and Fujian, using Hyper-seq-derived single-nucleotide polymorphism (SNP) markers. This is the first genetic study of R. chingii based on SNP molecular markers, and a total of 1,303,850 SNPs and 433,159 insertions/deletions (InDels) were identified. Low values for observed heterozygosity, nucleotide diversity (Pi) and fixation indexes (Fis) indicated low genetic diversity within populations, and an analysis of molecular variance (AMOVA) showed that 37.4% and 62.6% of the variations were found between populations and within samples, respectively. Four main clusters were identified by means of neighbor-joining (NJ) trees, the ADMIXTURE program and principal component analysis (PCA). Based on the genetic diversity, we finally constructed 38 representative core collections, representing 50% of the total core germplasm samples and 95.3% of the genotypes. In summary, the results of our study can provide valuable information on the genetic structure of R. chingii germplasm resources, which is helpful for further explorations of potential high-quality genes and for formulating future breeding and conservation strategies.

Development of Single Nucleotide Polymorphism and Association Analysis with Growth Traits for Black Porgy (Acanthopagrus schlegelii).

Black porgy is an important marine aquaculture fish species whose production is at the fifth position in all kinds of marine-cultured fishes in China. In this study, Illumina high-throughput sequencing technology was used to sequence the total RNA of black porgy. Sixty-one candidate SNPs (Single Nucleotide Polymorphism) were screened out and genotyped through GATK4 (Genome Analysis ToolKit) software and MALDI-TOF MS (Matrix-Assisted Laser Desorption/ Ionization Time of Flight Mass Spectrometry). The experimental results showed that a total of sixty SNPs were successfully genotyped, with a success rate of 98.36%. The results of principal component analysis and correlation analysis of growth traits showed that body weight was the first principal component, with a cumulative contribution rate of 74%. There were significant correlations (p < 0.05) or extremely significant correlations (p < 0.01) between different growth traits. The results of genetic parameter analysis and association analysis showed that scaffold12-12716321, scaffold13-4787950, scaffold2-13687576 and scaffold290-11890 were four SNPs that met the requirement of polymorphic information content and conformed to the Hardy-Weinberg equilibrium. There were significant differences between their genotype and the phenotype of growth traits. The four SNP molecular markers developed in this research will lay a foundation for further exploration of molecular markers related to the growth traits of black porgy and will provide a scientific reference for the further study of its growth mechanisms. At the same time, these molecular markers can be applied to the production practices of black porgy, so as to realize selective breeding at the molecular level and speed up the breeding process.

Characterization of Genetic Diversity and Variation in Pathogenicity of the Rice False Smut Pathogen Ustilaginoidea virens from a Single Source.

Rice false smut, caused by Ustilaginoidea virens, is one of the most destructive fungal diseases in rice-growing countries. Studies of the genetic diversity, evolution, and pathogenicity of U. virens can provide more information for disease control and cultivar breeding. Contrary to previous studies on the genetic diversity of different geographical populations of U. virens, this study analyzed the genetic variation of U. virens from different panicles of the same rice cultivar in a field in Yunnan Province using single nucleotide polymorphism molecular markers. A total of 183 polymorphic loci and five haplotypes, hap_1 to hap_5, were identified based on the 1,350-bp combined DNA fragment of 127 isolates, showing some genetic diversity. Hap_1 and hap_3 had the highest occurrence, indicating they were the dominant haplotypes in the field. Further analysis showed that most rice panicles could be coinfected by different haplotypes, and even a few spikelets could be coinfected by multiple haplotypes. The phylogeny indicated that all isolates were divided into five genetic groups. Groups I, II, and III clustered together and were distinguished from Groups IV and V. Significant genetic variations in five pairwise comparisons of panicle populations, accounting for 72.45% of the total variation, were found according to FST values. This variation might be caused by different field microenvironments and the uneven distribution of inoculum sources. An unweighted pair-group method with arithmetic means dendrogram and the population structure revealed that the genetic composition of the isolates collected from YN1, YN2, and YN4, which were dominated by the same genetic subgroup, was different from that collected from YN3. Finally, genetic recombination was found in 11 isolates; hap_2 and hap_5, probably as genetic recombination progenies produced by sexual hybridization between hap_1 and hap_3, acquired a greater virulence than their ancestors according to population structure and pathogenicity analyses. These results will help us understand the genetic diversity, evolution, and infection process of U. virens and aid in the development of more effective management strategies for rice false smut, including new cultivars with improved resistance.

Establishment and application of an SNP molecular identification system for grape cultivars

We aimed to develop a set of single nucleotide polymorphism (SNP) markers that can be used to distinguish the main cultivated grape (Vitis L.) cultivars in China and provide technical support for domestic grape cultivar protection, cultivar registration, and market rights protection. A total of 517 high-quality loci were screened from 4 241 729 SNPs obtained by sequencing 304 grape accessions using specific locus amplified fragment sequencing, of which 442 were successfully designed as Kompetitive Allele Specific PCR (KASP) markers. A set of 27 markers that completely distinguishes 304 sequenced grape accessions was determined by using the program, and 26 effective markers were screened based on 23 representative grape cultivars. Finally, a total of 46 out of 48 KASP markers, including 22 markers selected by the research group in the early stage, were re-screened based on 348 grape accessions. Population structure, principal component, and cluster analyses all showed that the 348 grape accessions were best divided into two populations. In addition, cluster analysis subdivided them into six subpopulations. According to genetic distance, V. labrusca, V. davidii, V. heyneana, and V. amurensis were far from V. vinifera, while V. vinifera×V. labrusca and V. amurensis×V. vinifera were somewhere in between these two groups. Furthermore, a core set of 25 KASP markers could distinguish 95.69% of the 348 grape accessions, and the other 21 markers were used as extended markers. Therefore, SNP molecular markers based on KASP typing technology provide a new way for mapping DNA fingerprints in grape cultivars. With high efficiency and accuracy and low cost, this technology is more competitive than other current identification methods. It also has excellent application prospects in the grape distinctness, uniformity, and stability (DUS) test, as well as in promoting market rights protection in the near future.

Development of specific single nucleotide polymorphism molecular markers for Angelica gigas Nakai

Development of specific single nucleotide polymorphism molecular markers for <i>Angelica gigas</i> Nakai

Identification of the 'Haryejosaeng' mandarin cultivar by multiplex PCR-based SNP genotyping.

Most satsuma mandarin (Citrus unshiu Marc.) cultivars are difficult to identify in the seedling stage based only on morphological traits. Therefore, simple polymerase chain reaction (PCR)-based single-nucleotide polymorphism (SNP) markers were developed to specifically and rapidly distinguish the 'Haryejosaeng' cultivar, which is generally supplied to breeders of other satsuma mandarin cultivars. SNP markers were verified using high-resolution melt (HRM)-specific primers. PCR was performed to distinguish 'Haryejosaeng' from eight other satsuma mandarin cultivars using six SNP markers (P1-P6) specific for 'Haryejosaeng', with one negative control SNP primer pair. The best results were obtained using three SNP markers (P1, P2, and P5). In the multiplex PCR, markers P1, P2, and P5 yielded 165-, 150-, and 526-base pair amplicons, respectively, in 'Haryejosaeng', distinguishing it from other satsuma mandarin cultivars. The selected SNP markers were validated by HRM with HRM-specific primers. The multiplex PCR with P1/P5 and P2/P5 also identified 'Haryejosaeng' obtained from a farm growing 17 different cultivars of satsuma mandarin. Specific SNP molecular markers were determined for accurately identifying the 'Haryejosaeng' cultivar by multiplex PCR to save the time and costs associated with its supply to breeders of satsuma mandarin.

Genetic diversity of Iranian and some European grapes as revealed by nuclear and chloroplast microsatellite and SNP molecular markers

ABSTRACTGenetic and chloroplast haplotype variations of 35 Iranian genotypes and 10 European grape cultivars were investigated using 9 nuclear simple sequence repeats (nSSRs), 4 chloroplast simple sequence repeats (cpSSRs) and 46 single nucleotide polymorphism (SNP) markers. In total, 83 alleles were detected at nine nSSRs, giving a mean of 15.66 alleles per locus and polymorphism information content (PIC) values ≥0.75 ranged from 0.75 to 0.90. For SNP markers, PIC values varied from 0.30 to 0.39 with an average of 0.34. Analysis of molecular variance revealed 97 and 93% of partitioned genetic diversity within populations using nSSRs and SNPs markers, respectively. Un-weighted neighbour-joining (NJ) cluster analysis grouped grapes into 10 and 9 major clusters using SSR and SNP markers, respectively. Synonyms and homonyms were identified among the Iranian genotypes. Close genetic relationship among Farkhi and Bidane-Sefid genotypes may probably propose a common ancestor and mutational evolution. Most European cultivars were differentiated from Iranian genotypes, however, clustering of some Iranian genotypes with European cultivars in the same clusters suggests that clonally propagated materials have probably been exchanged from the Middle East to West or vice versa. C and D chloroplast haplotypes were the most frequent within the Iranian genotypes, while A chloroplast haplotype was exclusively observed among European cultivars.

Molecular cloning analysis of the 5' regulatory region of Ikaros gene from Oreochromis niloticus and screening of its SNP markers for Streptococcus agalactiae resistance

PDF HTML阅读 XML下载 导出引用 引用提醒 吉富尼罗罗非鱼Ikaros基因5'调控区的克隆、序列分析及抗无乳链球菌相关SNP位点筛选 DOI: 作者: 作者单位: 1. 中国水产科学研究院 珠江水产研究所, 农业部热带亚热带水产资源利用与养殖重点实验室, 广东 广州 510380;2. 上海海洋大学 水产与生命学院, 上海 201306;3. 惠州市渔业研究推广中心, 广东 惠州 516002 作者简介: 陈昆平(1991-),男,硕士研究生,研究方向为水产健康养殖及遗传育种.E-mail:colinchenkunping@163.com 通讯作者: 中图分类号: S917 基金项目: 现代农业产业技术体系专项资金项目（CARS-46）；广东省自然科学基金项目（2016A030313146）；国家自然科学基金项目（31502205）. Molecular cloning analysis of the 5' regulatory region of Ikaros gene from Oreochromis niloticus and screening of its SNP markers for Streptococcus agalactiae resistance Author: Affiliation: 1. Pearl River Fisheries Research Institute, Chinese Academy of Fishery Sciences, Guangzhou 510380, China;2. College of Fisheries and Life Science, Shanghai Ocean University, Shanghai 201306, China;3. Fisheries Research and Extension Center of Huizhou, Huizhou 516002, China Fund Project: 摘要 | 图/表 | 访问统计 | 参考文献 | 相似文献 | 引证文献 | 资源附件 | 文章评论 摘要:为获得尼罗罗非鱼（）抗链球菌病相关的单核苷酸多态性（single nucleotide polymorphism，SNP）标记，本研究通过染色体步移法克隆了尼罗罗非鱼基因的启动子和转录调控元件进行预测和分析。利用PCR产物直接测序法从亲本（P0）尼罗罗非鱼基因5'调控区中筛查到5个SNPs，分别为SNP1（g.562，G>A）、SNP2（g.217，G>T）、SNP3（g.-53，C>T）、SNP4（g.-220，T>C）和SNP5（g.-579，T>C）。利用Snapshot技术对子一代（F1）尼罗罗非鱼易感群体和抗病群体进行相应SNPs的基因分型，分析两个群体遗传多样性参数，结果显示多态信息含量（polymorphism information content，PIC）值在0.0872~0.3747之间，表明基因5'调控区SNPs与抗链球菌病性状关联分析结果表明，SNP2、SNP3、SNP4和SNP5的基因型频率和等位基因频率在易感群体和抗病群体中存在显著差异（基因5'调控区SNPs可形成1个单倍块和5种单倍型，其中GGCTT单倍型与抗病性显著相关（<0.05）。此外，还发现SNP2和SNP5处于完全连锁状态（Ikaros基因5'调控区筛选到4个抗链球菌病相关SNPs和1个单倍型（GGCTT），均可作为尼罗罗非鱼分子育种的候选分子标记。 Abstract:Tilapia () is one of the most important fishery species in the world and has been introduced and cultured widespreadly because of its fast growth rate, strong reproductive capacity, good adaptability and omnivorous feeding habit. However, disease has become the biggest threat for tilapia breeding. Recently, the epidemic and outbreak of tilapia disease caused substantial economic losses to aquaculture industry. Breeding resistant variety in tilapia was unimportant way to solve the problem of disease. The molecular marker assisted selection (MAS) has become an efficient breeding method for selecting and breeding tilapia, and will accelerate genetic improvement and increase selection intensity for disease resistance. The single nucleotide polymorphism (SNP) markers are used in many genetic and breeding studies because they are abundant in genomes, and can be genotyped easily. Ikaros is a kind of transcription factor with zinc finger structure that is essential to the development of lymphocyte and to the maintenance of normal immune function. Therefore, to obtain large amount of effective SNP molecular genetic markers and to perform MAS for disease resistance in tilapia, it is essential to study immune related candidate and to examine whether the SNPs in the gene are associated with disease resistance. In this study, the 5' regulatory region sequence of , length of 4178 bp, were obtained through Genome Walking method from . Bioinformatics software was used to analyze the 5' regulatory region sequence of gene. The predicted transcriptional start site (TSS) was in the initiation codon (ATG) upstream of 931 bp, and the core promoter regions was located at -57 bp to 48 bp when the TSS was specified as 1. The predicted promoter regions of gene included basic start of substructure components:TATA box, CCAAT box and octamer. The analysis of transcription factor binding sites (TFBS) showed that abundant of TFBS were located at -2200 bp to 1200 bp in 5' regulatory region of gene, such as GATA-1, Homeobox, CDP CR3+HD and AP-1. The analysis of CpG islands showed that two CpG islands were in 5' regulatory region sequence of gene, one of which was located in promoter regions and the other of which was located in the first exon. Five SNPs in the 5' regulatory region of gene were detected by direct sequencing method from the parents (P0), which are named SNP1 (g.562, G>A), SNP2(g.217, G>T), SNP3(g.-53, C>T), SNP4(g.-220, T>C) and SNP5(g.-579, T>C). The five SNPs were sited in various regulatory elements in promoter regions which could have major implications for exact expression of gene. Based on Snapshot method analysis, the frequencies of alleles and genotypes were calculated in susceptible groups and resistant groups of ). The polymorphisms and genetic parameter of the SNPs in resistant groups and susceptible groups were calculated by software Popgen 32 and PIC. The result showed that the polymorphism information content (PIC) of the 5' regulatory region of SNPs was 0.0872~0.3747, suggesting that all the SNPs had moderate of intermediate polymorphism. The correlation between SNPs and resistance to was analyzed by software SPSS 17.0, the results showed that four of them (SNP2, SNP3, SNP4and SNP5) were significantly associated with the resistance to <0.05). Moreover, All of the SNPs in the 5' regulatory region of could formed one haplotype block and five haplotypes from the prediction of linkage disequilibrium analysis used software Haploview 4.2. The haplotypes (GGCTT) were significantly associated with the resistance to <0.05), and two of the haplotypes (GGTCT and GTCCC) were significantly associated with <0.05). Furthermore, the SNP2 and SNP5 were completely linked with each other ('=1), which could be selected as tag SNP for research of genetic breeding in . The results suggest that the four SNPs (SNP2, SNP3, SNP4 and SNP5) and the haplotypes (GGCTT) in the 5' regulatory region of could be potential genetic markers for future molecular selection of . 参考文献 相似文献 引证文献

Population structure and genetic diversity of coffee progenies derived from Catuaí and Híbrido de Timor revealed by genome-wide SNP marker

The use of single nucleotide polymorphism (SNP) molecular markers has provided advances in selection methodologies used in breeding programs of different crops, reducing cost and time of cultivar release. Despite the great economic and social importance of Coffea arabica, studies with SNP markers are scarce and a small number of SNP are available for this species, when compared with other crops of agronomic importance. Thus, the objective of this study was to identify and validate SNP molecular markers for the species Coffea arabica and to introduce these markers to genetic breeding by means of an accurate analysis of the diversity and genetic structure of breeding populations of this species. After quality filtering, 11,187 SNP markers were selected from the coffee population obtained from crosses between the genotypes Catuai and Hibrido de Timor. A great number of markers were distributed in the 11 chromosomes, within transcribed regions, and were used to estimate the genetic dissimilarity among the individuals of the breeding population. Dendrogram analysis and a Bayesian approach demonstrated the formation of two groups and the discrimination of all genotypes evaluated. The expressive number of SNP molecular markers distributed throughout C. arabica genome was efficient to discriminate all the accessions evaluated in the experiment, clustering them according to their genealogies. This work identified mixtures within the progenies. The genotyping data also provided detailed information about the parental genotypes and led to the identification of new candidate parents to be introduced to the breeding program. The study discussed population structure and its consequence in obtaining improved varieties of C. arabica.

First interspecific genetic linkage map for Castanea sativa x Castanea crenata revealed QTLs for resistance to Phytophthora cinnamomi.

The Japanese chestnut (Castanea crenata) carries resistance to Phytophthora cinnamomi, the destructive and widespread oomycete causing ink disease. The European chestnut (Castanea sativa), carrying little to no disease resistance, is currently threatened by the presence of the oomycete pathogen in forests, orchards and nurseries. Determining the genetic basis of P. cinnamomi resistance, for further selection of molecular markers and candidate genes, is a prominent issue for implementation of marker assisted selection in the breeding programs for resistance. In this study, the first interspecific genetic linkage map of C. sativa x C. crenata allowed the detection of QTLs for P. cinnamomi resistance. The genetic map was constructed using two independent, control-cross mapping populations. Chestnut populations were genotyped using 452 microsatellite and single nucleotide polymorphism molecular markers derived from the available chestnut transcriptomes. The consensus genetic map spans 498,9 cM and contains 217 markers mapped with an average interval of 2.3 cM. For QTL analyses, the progression rate of P. cinnamomi lesions in excised shoots inoculated was used as the phenotypic metric. Using non-parametric and composite interval mapping approaches, two QTLs were identified for ink disease resistance, distributed in two linkage groups: E and K. The presence of QTLs located in linkage group E regarding P. cinnamomi resistance is consistent with a previous preliminary study developed in American x Chinese chestnut populations, suggesting the presence of common P. cinnamomi defense mechanisms across species. Results presented here extend the genomic resources of Castanea genus providing potential tools to assist the ongoing and future chestnut breeding programs.

Confirmation of quantitative trait loci for resistance to multiple-HG types of soybean cyst nematode (Heterodera glycines Ichinohe)

Genetic analysis of resistance of plant introduction (PI) 438489B to soybean cyst nematode (SCN) have shown that this PI is highly resistant to many SCN HG types. However, validation of the previously detected quantitative trait loci (QTL) has not been done. In this study, 250 F2:3 progeny of a Magellan (susceptible) × PI 438489B (resistant) cross were used for primary genetic mapping to detect putative QTL for resistance to five SCN HG types. QTL confirmation study was subsequently conducted using F6:7 recombinant inbred lines (RILs) derived from the same cross. Simple sequence repeat (SSR) and single nucleotide polymorphism (SNP) markers were employed for molecular genotyping. Interval mapping (IM), permutation tests, cofactor selection, and composite interval mapping (CIM) were performed to identify and map QTL. Results showed that five QTL intervals were associated with resistance to either multiple- or single-HG types of SCN. Among these, two major QTL for resistance to multiple-SCN HG types were mapped to chromosomes (Chr.) 8 and 18, consistent with the known rhg1 and Rhg4 locations. The other QTL were mapped to Chr. 4. The results of our study confirmed earlier reported SCN resistance QTL in this PI. Moreover, SSR and SNP molecular markers tightly linked to these QTL can be useful for the near-isogenic lines (NILs) development aiming to fine-mapping of these QTL regions and map-based cloning of SCN resistance candidate genes.