Genome-Wide Comparative Analyses of Microsatellites in Papaya

Microsatellites, or simple sequence repeats (SSRs), are highly polymorphic and universally distributed in eukaryotes. SSRs have been used extensively as sequence tagged markers in genetic studies. Recently, the functional and evolutionary importance of SSRs has received considerable attention. Here we report the mining and characterization of the SSRs in papaya genome. We analyzed SSRs from 277.4 Mb of whole genome shotgun (WGS) sequences, 51.2 Mb bacterial artificial chromosome (BAC) end sequences (BES), and 13.4 Mb expressed sequence tag (EST) sequences. The papaya SSR density was one SSR per 0.7 kb of DNA sequence in the WGS, which was higher than that in BES and EST sequences. SSR abundance was dramatically reduced as the repeat length increased. According to SSR motif length, dinucleotide repeats were the most common motif in class I, whereas hexanucleotides were the most copious in class II SSRs. The tri- and hexanucleotide repeats of both classes were greater in EST sequences compared to genomic sequences. In class I SSR, AT and AAT were the most frequent motifs in BES and WGS sequences. By contrast, AG and AAG were the most abundant in EST sequences. For SSR marker development, 9,860 primer pairs were surveyed for amplification and polymorphism. Successful amplification and polymorphic rates were 66.6% and 17.6%, respectively. The highest polymorphic rates were achieved by AT, AG, and ATG motifs. The genome wide analysis of microsatellites revealed their frequency and distribution in papaya genome, which varies among plant genomes. This complete set of SSRs markers throughout the genome will assist diverse genetic studies in papaya and related species.     

蜜蜂全基因组中微卫星的丰度及其分布

查找出蜜蜂基因组中由1～6个碱基重复单元组成的简单序列重复,分析蜜蜂基因组中微卫星的分布频率,并比较其在各染色体中的分布频率。微卫星在蜜蜂基因组中的分布频率为1/0·804kb,其中二碱基重复序列占26·86%,是最丰富的重复单元,而六、一、三、四、五碱基重复单元序列分别占24·74%,22·19%,13·65%,10·98%,2·59%。同时发现富含A和T碱基的微卫星占主导地位,富含G和C碱基的微卫星数量较少。第4,1,3条染色体微卫星分布频率较高,而第11,14,12条染色体微卫星分布频率较低。     

红原鸡全基因组中微卫星分布规律研究

本文对红原鸡Gallus gallus全基因组中微卫星数量及分布规律进行了分析,查找到l～6个碱基重复类型的微卫星序列共282728个,约占全基因组序列(1.1Gb)的0.49%,分布频率为1/3.89kb,微卫星序列的长度主要在12～70个碱基长度范围内。第1、2、3条染色体上微卫星分布频率较高,而32号染色体上无微卫星分布。不同类型微卫星中,单碱基重复类型数目最多,为184192个,占总数的65.1%;其次是四、二、三、五、六碱基重复单元序列,分别占到总数的12.8%、9.7%、7.2%、4.6%、0.8%。T、A、AT、GTTT、AAAC、G、C、ATTT、AC、GT、AAAT、ATT、AAC、AAT、GTT、AG、CT、CTTT、AAAG、GTTTT、AAACA、AAGG、CCTT是红原鸡基因组中最主要的微卫星重复类型。本研究为红原鸡微卫星标记的分离筛选、遗传多样性的研究以及不同物种微卫星的比较分析奠定了基础。     

Analysis of distribution and significance of simple sequence repeats in enteric bacteria Shigella dysenteriae SD197

We have explored the possible role of SSR density in genome to generate biological information. In our study, we have checked the SSR (simple sequence repeats) status in virulent and non virulent genes of enteric bacteria to see whether the SSRs distribution contributes to virulence. The genome, plasmid and virulent genes sequences in fasta format were downloaded from NCBI GenBank and VFDB. The sequences were subjected to SSR analysis using software tool ssr.exe. The resulting data was pasted in excel sheet and further analyzed for percentage of each type of SSR. Higher nucleotide repeats have been observed in our study. Overall high density of SSRs can enhance antigenic variance of the pathogen population in a strategy that counteracts the host immune response. Frequency of A and T repeats is higher in the chromosome, plasmid and the virulence genes. However, in dinucleotide repeats the frequencies of GC/CG repeats are higher in genome, whereas plasmid has more of AT/TA repeats. Genome has trinucleotide repeats having predominantly G and C whereas plasmid has trinucleotide repeats having predominantly A and T. The repeat number obtained and percentage of repeats is higher in virulence genes as compared to other gene families. Due to the presence of this large number of SSRs, the organism has an enormous potential for generating this genomic and phenotypic diversity.     

Genome-wide identification and characterization of simple sequence repeat loci in grape phylloxera, Daktulosphaira vitifoliae

A genome-wide sequence search was conducted to identify simple sequence repeat (SSR) loci in phylloxera, Daktulosphaira vitifoliae, a major grape pest throughout the world. Collectively, 1524 SSR loci containing mono-, di-, tri-, tetra-, penta-, and hexanucleotide motifs were identified. Among them, trinucleotide repeats were the most abundant in the phylloxera genome (34.4%), followed by hexanucleotide (20.4%) and dinucleotide (19.6%) repeats. Mono-, tetra- and pentanucleotide repeats were found at a frequency of 1.3, 11.2 and 12.9%, respectively. The abundance and inherent variations in SSRs provide valuable information for developing molecular markers. The high levels of allelic variation and codominant features of SSRs make this marker system a useful tool for genotyping, diversity assessment and population genetic studies of reproductive characteristics of phylloxera in agricultural and natural populations.     

蚊子全基因组中微卫星的丰度及其分布

微卫星是近年大力开发的一种遗传标记,为推进按蚊遗传学相关研究,对按蚊全基因组中由 1~6 个碱基重复单元组成的简单序列重复 ( 微卫星 ) 进行了分析 . 进而对其微卫星的丰度和分布进行了比较分析,也比较了染色体各个区域 ( 外显子、内含子和基因间隔区 ) 之间的分布差异 . 微卫星在按蚊基因组中的比例约占 2.14% ,其中 X 染色体拥有微卫星的密度最大 . 对按蚊基因组中微卫星丰度而言, A 碱基和 C 碱基重复在基因组中丰度相似, AC 单元的丰度是 AG 单元的两倍多,然而 AT 和 CG 单元非常稀少;对于三四碱基而言, AGC, AAAC 和 AAAT 单元最为丰富, ACG, ACT, AGG, CCG, ATGC, CCCG, ACTG, AACT, ACGT, AGAT, CCGG, ACCT 和 AGCT 单元等均很稀少,而一些五碱基重复,在某条甚至某几条染色体中均未分布 . 除两碱基重复单元在 2L 的外显子区域丰度较高外,其他重复单元均在内含子和基因间隔区丰富 . 进一步分析显示,微卫星在每条染色体两臂的丰度和分布存在着很多的相似性 .     

Genomic distribution of simple sequence repeats in Brassica rapa

Simple Sequence Repeats (SSRs) represent short tandem duplications found within all eukaryotic organisms. To examine the distribution of SSRs in the genome of Brassica rapa ssp. pekinensis, SSRs from different genomic regions representing 17.7 Mb of genomic sequence were surveyed. SSRs appear more abundant in non-coding regions (86.6%) than in coding regions (13.4%). Comparison of SSR densities in different genomic regions demonstrated that SSR density was greatest within the 5'-flanking regions of the predicted genes. The proportion of different repeat motifs varied between genomic regions, with trinucleotide SSRs more prevalent in predicted coding regions, reflecting the codon structure in these regions. SSRs were also preferentially associated with gene-rich regions, with peri-centromeric heterochromatin SSRs mostly associated with retrotransposons. These results indicate that the distribution of SSRs in the genome is non-random. Comparison of SSR abundance between B. rapa and the closely related species Arabidopsis thaliana suggests a greater abundance of SSRs in B. rapa, which may be due to the proposed genome triplication. Our results provide a comprehensive view of SSR genomic distribution and evolution in Brassica for comparison with the sequenced genomes of A. thaliana and Oryza sativa.     

Distribution and characterization of simple sequence repeats in Gossypium raimondii genome

Simple sequence repeats (SSRs) can be derived from the complete genome sequence. These markers are important for gene mapping as well as marker-assisted selection (MAS). To develop SSRs for cotton gene mapping, we selected the complete genome sequence of Gossypium raimondii, which consisted of 4447 non-redundant scaffolds. Out of 775.2 Mb sequence examined, a total of 136,345 microsatellites were identified with a density of 5.69 kb per SSR in the G. raimondii genome leading to development of 112,177 primer pairs. The distributions of SSRs in the genome were non-random. Among the different motifs ranging from 1 to 6 bp, penta-nucleotide repeats were most abundant (30.5%), followed by tetra-nucleotide repeats (18.2%) and di-nucleotide repeats (16.9%). Among all identified 457 motif types, the most frequently occurring repeat motifs were poly-AT/TA, which accounted for 79.8% of the total di-nt SSRs, followed by AAAT/TTTA with 51.5% of the total tetra-nucleotede. Further, 18,834 microsatellites were detected from the protein-coding genes, and the frequency of gene containing SSRs was 46.0% in 40,976 genes of G. raimondii. These genome-based SSRs developed in the present study will lay the groundwork for developing large numbers of SSR markers for genetic mapping, gene discovery, genetic diversity analysis, and MAS breeding in cotton.     

In silico analysis of SSRs in mitochondrial genomes of plants

Simple sequence repeats (SSRs) or microsatellites constitute a countable portion of genomes. However, the significance of SSRs in organelle genomes has not been completely understood. The availability of organelle genome sequences allows us to understand the organization of SSRs in their genic and intergenic regions. In the current study we surveyed the patterns of SSRs in mitochondrial genomes of different taxa of plants. A total of 16 mitochondrial genomes, from algae to angiosperms, have been considered to analyze the pattern of simple sequence repeats present in them. Based on study, the mononucleotide repeats of A/T were found to be more prevalent in mitochondrial genomes over other repeat types. The dinucleotides repeats, TA/AT, were the second most numerous, whereas tri-, tetra-, and pentanucleotide repeats were in less number and present in intronic or intergenic portions only. Mononucleotide repeats prevailed in protein-coding exonic portions of all organisms. These results indicates that microsatellite pattern in mitochondrial genomes is different from nuclear genomes and also focuses on organization and diversity at SSR locuses in mitochondrial genomes. This is the novel report of microsatellite polymorphism in plant mitochondrion on whole genome level.     

Analysis on frequency and density of microsatellites in coding sequences of several eukaryotic genomes

Microsatellites or simple sequence repeats (SSRs) have been found in most organisms during the last decade. Since large-scale sequences are being generated, especially those that can be used to search for microsatelUtes, the development of these markers is getting more convenient. Keeping SSRs in viewing the importance of the application, available CDS (coding sequences) or ESTs (expressed sequence tags) of some eukaryotic species were used to study the frequency and density of various types of microsatellites. On the basis of surveying CDS or EST sequences amounting to 66.6 Mb in silkworm, 37.2 Mb in fly, 20.8 Mb in mosquito, 60.0 Mb in mouse, 34.9 Mb in zebrafish and 33.5 Mb in Caenorhabditis elegans, the frequency of SSRs was 1/1.00 Kb in silkworm, 1/0.77 Kb in fly, 1/1.03 Kb in mosquito, 1/1.21 Kb in mouse, 1/1.25 Kb in zebrafish and 1/1.38 Kb in C. elegans. The overall average SSR frequency of these species is 1/1.07 Kb. Hexanucleotide repeats (64.5%-76.6%) are the most abundant class of SSR in the investigated species, followed by trimeric, dimeric, tetrameric, monomeric and pentameric repeats. Furthermore, the A-rich repeats are predominant in each type of SSRs, whereas G-rich repeats are rare in the coding regions.     

Analysis on Frequency and Density of Microsatellites in Coding Seauences of Several Eukarvotic Genomes

Microsatellites or simple sequence repeats (SSRs) have been found in most organisms during the last decade. Since large-scale sequences are being generated, especially those that can be used to search for microsatellites, the development of these markers is getting more convenient. Keeping SSRs in viewing the importance of the application, available CDS (coding sequences) or ESTs (expressed sequence tags) of some eukaryotic species were used to study the frequency and density of various types of microsatellites. On the basis of surveying CDS or EST sequences amounting to 66.6 Mb in silkworm, 37.2 Mb in fly, 20.8 Mb in mosquito, 60.0 Mb in mouse, 34.9 Mb in zebrafish and 33.5 Mb in Caenorhabditis elegans, the frequency of SSRs was 1/1.00 Kb in silkworm, 1/0.77 Kb in fly, 1/1.03 Kb in mosquito, 1/1.21 Kb in mouse, 1/1.25 Kb in zebrafish and 1/1.38 Kb in C. elegans. The overall average SSR frequency of these species is 1/1.07 Kb. Hexanucleotide repeats (64.5%-76.6%) are the most abundant class of SSR in th     

Development of microsatellite markers in autopolyploid sugarcane and comparative analysis of conserved microsatellites in sorghum and sugarcane

Sugarcane has become an increasingly important first-generation biofuel crop in tropical and subtropical regions. It has a large, complex, polyploid genome that has hindered the progress of genomic research and marker-assisted selection. Genetic mapping and ultimately genome sequence assembly require a large number of DNA markers. Simple sequence repeats (SSRs) are widely used in genetic mapping because of their abundance, high rates of polymorphism, and ease of use. The objectives of this study were to develop SSR markers for construction of a saturated genetic map and to characterize the frequency and distribution of SSRs in a polyploid genome. SSR markers were mined from expressed sequence tag (EST), reduced representation library genomic sequences, and bacterial artificial chromosome (BAC) sequences. A total of 5,675 SSR markers were surveyed in a segregating population. The overall successful amplification and polymorphic rates were 87.9 and 16.4%, respectively. The trinucleotide repeat motifs were most abundant, with tri- and hexanucleotide motifs being the most abundant for the ESTs. BAC and genomic SSRs were mostly AT-rich while the ESTs were relatively GC-rich due to codon bias. These markers were also aligned to the sorghum genome, resulting in 1,203 markers mapped in the sorghum genome. This set of SSRs conserved in sugarcane and sorghum would be the most informative for mapping quantitative trait loci in sugarcane and for comparative genomic analyses. This large collection of SSR markers is a valuable resource for sugarcane genomic research and crop improvement.     

Genome-Wide Analysis of Microsatellite Markers Based on Sequenced Database in Chinese Spring Wheat (Triticum aestivum L.)

Microsatellites or simple sequence repeats (SSRs) are distributed across both prokaryotic and eukaryotic genomes and have been widely used for genetic studies and molecular marker-assisted breeding in crops. Though an ordered draft sequence of hexaploid bread wheat have been announced, the researches about systemic analysis of SSRs for wheat still have not been reported so far. In the present study, we identified 364,347 SSRs from among 10,603,760 sequences of the Chinese spring wheat (CSW) genome, which were present at a density of 36.68 SSR/Mb. In total, we detected 488 types of motifs ranging from di- to hexanucleotides, among which dinucleotide repeats dominated, accounting for approximately 42.52% of the genome. The density of tri- to hexanucleotide repeats was 24.97%, 4.62%, 3.25% and 24.65%, respectively. AG/CT, AAG/CTT, AGAT/ATCT, AAAAG/CTTTT and AAAATT/AATTTT were the most frequent repeats among di- to hexanucleotide repeats. Among the 21 chromosomes of CSW, the density of repeats was highest on chromosome 2D and lowest on chromosome 3A. The proportions of di-, tri-, tetra-, penta- and hexanucleotide repeats on each chromosome, and even on the whole genome, were almost identical. In addition, 295,267 SSR markers were successfully developed from the 21 chromosomes of CSW, which cover the entire genome at a density of 29.73 per Mb. All of the SSR markers were validated by reverse electronic-Polymerase Chain Reaction (re-PCR); 70,564 (23.9%) were found to be monomorphic and 224,703 (76.1%) were found to be polymorphic. A total of 45 monomorphic markers were selected randomly for validation purposes; 24 (53.3%) amplified one locus, 8 (17.8%) amplified multiple identical loci, and 13 (28.9%) did not amplify any fragments from the genomic DNA of CSW. Then a dendrogram was generated based on the 24 monomorphic SSR markers among 20 wheat cultivars and three species of its diploid ancestors showing that monomorphic SSR markers represented a promising source to increase the number of genetic markers available for the wheat genome. The results of this study will be useful for investigating the genetic diversity and evolution among wheat and related species. At the same time, the results will facilitate comparative genomic studies and marker-assisted breeding (MAS) in plants.     

埃博拉病毒基因组中微卫星序列的分布分析

微卫星或简单重复序列(simple sequence repeat, SSR)在真核和原核生物以及病毒基因组中普遍存在,并被广泛用于遗传与进化研究。本研究从NCBI中下载埃博拉病毒属的四个不同种的埃博拉病毒全基因组序列,筛选36条作为实验材料,利用IMEx在线提取软件提取SSRs,用Python编程统计数据,从而分析SSRs在埃博拉病毒全基因组序列中的分布情况。分析得出,埃博拉病毒基因组序列中二型SSRs含量最为丰富,其次是一型SSRs,三型SSRs有少量,四型SSRs则更少,没有发现五型和六型SSRs。在更深入的分析中得出在埃博拉病毒属四个种中,含A/T碱基的SSRs含量远远大于含C/G碱基的SSRs。分析得出一型SSRs中(A)n/(T)n远多于(G)n/(C)n,二型SSRs中不存在(GC/CG)n,三型中也不存在(GGC/CGG/GCG/CCG/CGC/GCC) n。上述发现可能跟埃博拉病毒的致病机理有密切联系。通过对埃博拉病毒基因组序列中SSRs的分析,为研究埃博拉病毒的变异情况及致病机制提供更多参考。     

Genome-wide characterization and analysis of microsatellite sequences in camelid species

Microsatellites or simple sequence repeats (SSRs) are among the genetic markers most widely utilized in research. This includes applications in numerous fields such as genetic conservation, paternity testing, and molecular breeding. Though ordered draft genome assemblies of camels have been announced, including for the Arabian camel, systemic analysis of camel SSRs is still limited. The identification and development of informative and robust molecular SSR markers are essential for marker assisted breeding programs and paternity testing. Here we searched and compared perfect SSRs with 1–6 bp nucleotide motifs to characterize microsatellites for draft genome sequences of the Camelidae. We analyzed and compared the occurrence, relative abundance, relative density, and guanine-cytosine (GC) content in four taxonomically different camelid species: Camelus dromedarius, C. bactrianus, C. ferus, and Vicugna pacos. A total of 546762, 544494, 547974, and 437815 SSRs were mined, respectively. Mononucleotide SSRs were the most frequent in the four genomes, followed in descending order by di-, tetra-, tri-, penta-, and hexanucleotide SSRs. GC content was highest in dinucleotide SSRs and lowest in mononucleotide SSRs. Our results provide further evidence that SSRs are more abundant in noncoding regions than in coding regions. Similar distributions of microsatellites were found in all four species, which indicates that the pattern of microsatellites is conserved in family Camelidae.     

Genome-Wide Characterization and Linkage Mapping of Simple Sequence Repeats in Mei (Prunus mume Sieb. et Zucc.)

Because of its popularity as an ornamental plant in East Asia, mei (Prunus mume Sieb. et Zucc.) has received increasing attention in genetic and genomic research with the recent shotgun sequencing of its genome. Here, we performed the genome-wide characterization of simple sequence repeats (SSRs) in the mei genome and detected a total of 188,149 SSRs occurring at a frequency of 794 SSR/Mb. Mononucleotide repeats were the most common type of SSR in genomic regions, followed by di- and tetranucleotide repeats. Most of the SSRs in coding sequences (CDS) were composed of tri- or hexanucleotide repeat motifs, but mononucleotide repeats were always the most common in intergenic regions. Genome-wide comparison of SSR patterns among the mei, strawberry (Fragaria vesca), and apple (Malus×domestica) genomes showed mei to have the highest density of SSRs, slightly higher than that of strawberry (608 SSR/Mb) and almost twice as high as that of apple (398 SSR/Mb). Mononucleotide repeats were the dominant SSR motifs in the three Rosaceae species. Using 144 SSR markers, we constructed a 670 cM-long linkage map of mei delimited into eight linkage groups (LGs), with an average marker distance of 5 cM. Seventy one scaffolds covering about 27.9% of the assembled mei genome were anchored to the genetic map, depending on which the macro-colinearity between the mei genome and Prunus T×E reference map was identified. The framework map of mei constructed provides a first step into subsequent high-resolution genetic mapping and marker-assisted selection for this ornamental species.     

Sequence variations of simple sequence repeats on chromosome-4 in two subspecies of the Asian cultivated rice

Computational screening of the chromosome-4 sequence of the rice cultivar Nipponbare (Oryza sativa L. japonica) revealed 1,844 tandem simple sequence repeats (SSRs) or microsatellites with SSR motifs 20 bp and repeated unit length of 1–6 base pairs. Thus SSRs occur once in every 18.8 kb, on the average, on the chromosome with one SSR per 23.8 kb and 16 kb on the short and long arms, respectively. No SSR was detected in the core region of the centromere. Poly(AT) _n repeats represented the most abundant and length polymorphic class of SSRs on the chromosome, but it did not occur in the exons. GC-rich trinucleotide repeats were most abundant in the coding regions, representing 71.69% of the SSRs identified in the exons. Two hundred and twenty four SSRs were associated with the repetitive DNA sequences, most of them were poly(AT) _n tracts. Sequence variations of SSRs between two cultivars, representing the two subspecies of the Asian cultivated rice indica and japonica, were identified, revealing that divergence and convergence of the two subspecies could be traced by the analysis of SSRs. These results provide a great opportunity for SSR-based marker development and comparative genome analysis of the two subspecies of the Asian cultivated rice.Electronic Supplementary Material Supplementary material is available in the online version of this article at .Communicated by Q. Zhang     

烟草EST-SSR位点分析

利用MISA软件对烟草EST公共数据库中的简单重复序列(SSRs)进行了分析。结果表明,在133523条EST序列中,共获得81757条SSR序列,SSRs之间的距离约为0.92 kb。其中,六碱基重复丰度最大,占60.3%,而单碱基、三碱基、四碱基、二碱基和五碱基重复丰度分别为20.0%、11.0%、4.2%、2.8%和1.7%。在单碱基、二碱基、三碱基和四碱基重复模体中,丰度最大的分别是A/T、AG、AAG和AAAT,而CG在编码区内丰度很低。用CAP3软件进行冗余分析表明,在这6种类型的重复模体中,冗余与非冗余的烟草EST之间没有显著差异。在得到的SSR序列中随机选择10个序列设计引物,在7个烟草品种中进行PCR扩增。结果表明,10对引物全部扩增出PCR产物,其中8对引物扩增出预期片段。用这8组扩增出预期片段的PCR产物进行变性PAGE凝胶电泳检测,结果表明,其中有4对引物(EB4、EB5、EB6和EB8)扩增出多态性条带。     

Genome-wide identification of simple sequence repeats and development of polymorphic SSR markers for genetic studies in tea plant (<Emphasis Type="Italic">Camellia sinensis</Emphasis>)

The tea plant (Camellia sinensis (L.) O. Kuntze) is one of the most popular non-alcoholic beverage crops worldwide. The availability of complete genome sequences for the Camellia sinensis var. ‘Shuchazao’ has provided the opportunity to identify all types of simple sequence repeat (SSR) markers by genome-wide scan. In this study, a total of 667,980 SSRs were identified in the ~?3.08 Gb genome, with an overall density of 216.88 SSRs/Mb. Dinucleotide repeats were predominant among microsatellites (72.25%), followed by trinucleotide repeats (15.35%), while the remaining SSRs accounted for less than 13%. The motif AG/CT (49.96%) and AT/TA (40.14%) were the most and the second most abundant among all identified SSR motifs, respectively; meanwhile, AAT/ATT (41.29%) and AAAT/ATTT (67.47%) were the most common among trinucleotides and tetranucleotides, respectively. A total of 300 primer pairs were designed to screen six tea cultivars for polymorphisms of SSR markers using the five selected repeat types of microsatellite sequences. The resulting 96 SSR markers that yielded polymorphic and unambiguous bands were further deployed on 47 tea cultivars for genetic diversity assessment, demonstrating high polymorphism of these SSR markers. Remarkably, the dendrogram revealed that the phylogenetic relationships among these tea cultivars are highly consistent with their genetic backgrounds or places of origin. The identified genome-wide SSRs and newly developed SSR markers will provide a powerful means for genetic researches in tea plant, including genetic diversity and evolutionary origin analysis, fingerprinting, QTL mapping, and marker-assisted selection for breeding.