Characterization and Development of EST-SSR Markers from Transcriptome Sequences of Chrysanthemum (Chrysanthemum ×morifolium Ramat.)

Simple sequence repeat (SSR) markers are valuable for genetic and breeding applications, but SSR resources for the ornamental genus chrysanthemum (Chrysanthemum ×morifolium Ramat.) are still limited. Expressed sequence tags (ESTs) are sources of SSRs that represent an opportunity to develop SSRs to accelerate molecular breeding in chrysanthemum. In total, 4661 SSR loci were identified from 3823 SSR-containing unigenes in the chrysanthemum transcriptome with an average of one SSR per 6.98 kb. Of these SSR sequences, trinucleotide repeats (30.0%) predominated, followed by dinucleotide repeats (17.9%). In total, 1584 primer pairs were subsequently synthesized. By screening the parents and six individuals of the F1 progeny, 831 (52.5%) valid EST-SSR markers were identified, of which 361 (43.4%) were polymorphic. The annotation of unigenes containing polymorphic SSRs indicated that 330 (93.5%) demonstrated significant homology to other plant protein sequences. Twenty-five polymorphic EST-SSR markers were further selected for transferability analysis and exhibited 93% amplification in six Ajania species and six other Chrysanthemum species. Based on genotyping of the 59 samples, neighbor-joining analysis revealed six phylogenetic groupings, which was confirmed by population structure analysis and principal component analysis (PCA). Phylogenetic relationships among the 59 samples revealed by SSRs were highly consistent with the traditional taxonomic classification of Chrysanthemum and Ajania. The polymorphism information content (PIC) values ranged from 0.29 to 0.86, with a mean of 0.67, indicating high levels of informativeness. This research reveals the SSR distribution characteristics of chrysanthemum and provides a large number of new EST-SSR markers for further genetic diversity studies, genetic mapping, and molecular marker-assisted selection breeding for chrysanthemum.

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Simple Sequence Repeat and S-Locus Genotyping to Assist the Genetic Characterization and Breeding of Polyploid Prunus Species, P. spinosa and P. domestica subsp. insititia

Biochemical Genetics ◽

10.1007/s10528-021-10090-7 ◽

2021 ◽

Author(s):

Júlia Halász ◽

Noémi Makovics-Zsohár ◽

Ferenc Szőke ◽

Sezai Ercisli ◽

Attila Hegedűs

Keyword(s):

Simple Sequence Repeat ◽

Principal Component ◽

Sequence Repeat ◽

Breeding Programs ◽

Allele Number ◽

Genetic Potential ◽

Ssr Loci ◽

High Level ◽

Diversity Studies ◽

Simple Sequence

AbstractPolyploid Prunus spinosa (2n = 4 ×) and P. domestica subsp. insititia (2n = 6 ×) represent enormous genetic potential in Central Europe, which can be exploited in breeding programs. In Hungary, 16 cultivar candidates and a recognized cultivar ‘Zempléni’ were selected from wild-growing populations including ten P. spinosa, four P. domestica subsp. insititia and three P. spinosa × P. domestica hybrids (2n = 5 ×) were also created. Genotyping in eleven simple sequence repeat (SSR) loci and the multiallelic S-locus was used to characterize genetic variability and achieve a reliable identification of tested accessions. Nine SSR loci proved to be polymorphic and eight of those were highly informative (PIC values ˃ 0.7). A total of 129 SSR alleles were identified, which means 14.3 average allele number per locus and all accessions but two clones could be discriminated based on unique SSR fingerprints. A total of 23 S-RNase alleles were identified and the complete and partial S-genotype was determined for 10 and 7 accessions, respectively. The DNA sequence was determined for a total of 17 fragments representing 11 S-RNase alleles. ‘Zempléni’ was confirmed to be self-compatible carrying at least one non-functional S-RNase allele (SJ). Our results indicate that the S-allele pools of wild-growing P. spinosa and P. domestica subsp. insititia are overlapping in Hungary. Phylogenetic and principal component analyses confirmed the high level of diversity and genetic differentiation present within the analysed accessions and indicated putative ancestor–descendant relationships. Our data confirm that S-locus genotyping is suitable for diversity studies in polyploid Prunus species but non-related accessions sharing common S-alleles may distort phylogenetic inferences.

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Mining, characterization and application of transcriptome-based SSR markers in Chinese jiaotou

Plant Genetic Resources ◽

10.1017/s1479262117000338 ◽

2018 ◽

Vol 16 (4) ◽

pp. 306-314

Author(s):

Chan Liu ◽

Qing Tang ◽

Chaohua Cheng ◽

Ying Xu ◽

Zemao Yang ◽

...

Keyword(s):

Ssr Markers ◽

Large Scale ◽

Trinucleotide Repeat ◽

De Novo ◽

Local Varieties ◽

Genetic Studies ◽

Oxidation Reduction ◽

Ssr Loci ◽

Wild Accessions ◽

Expressed Sequence

AbstractChinese jiaotou is an economically important crop that is widely cultivated in East Asia. The lack of simple sequence repeat (SSR) markers has been a major obstacle for genetic studies of this crop. In the present study, SSR markers were developed for Chinese jiaotou on a large scale, based on the crop's transcriptome assembledde novoby a previous study. A search for SSR loci in the transcriptome's expressed sequence tags (ESTs) revealed 2157 SSRs, of which primer pairs could be developed for 1494. Among these resulting SSRs, trinucleotide repeat motifs were the most abundant type, with GAA/TTC motifs occurring most frequently. Analysing the annotated function of SSR-containing ESTs revealed that they enriched into the GO categories involved in transcription regulation, oxidation–reduction, transport, etc. The quality and transferability of these markers were also assessed using 100 randomly selected EST–SSRs, and the result showed that these markers were of good quality and possessed high cross-species transferability. In addition, the developed SSR markers were used to analyse the genetic diversity of 19 cultivated and four wild accessions, resulting in three distinct groups, cluster I, II and III. Interestingly, all four wild accessions were assigned to cluster III, and two local varieties from northern Hunan, China, were closely related to the wild genotypes. These results provide new insights into the origin of Chinese jiaotou. The EST–SSRs developed herein represent the first large-scale development of SSR markers in Chinese jiaotou, and they can be widely used for genetic studies of the crop.

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Development and Application of EST-SSR Markers for DNA Fingerprinting and Genetic Diversity Analysis of the Main Cultivars of Black Locust (Robinia pseudoacacia L.) in China

Forests ◽

10.3390/f10080644 ◽

2019 ◽

Vol 10 (8) ◽

pp. 644 ◽

Cited By ~ 2

Author(s):

Li Dong ◽

Yuhan Sun ◽

Keqi Zhao ◽

Jing Zhang ◽

Yuwei Zhang ◽

...

Keyword(s):

Ssr Markers ◽

Black Locust ◽

Robinia Pseudoacacia ◽

Diversity Analysis ◽

Simple Method ◽

Genetic Diversity Analysis ◽

Ssr Loci ◽

Robinia Pseudoacacia L ◽

Expressed Sequence ◽

Simple Sequence

Black locust (Robinia pseudoacacia L.) is an economically and ecologically important tree species which is used for pillar construction, honey production and soil improvement. More EST-SSR (Expressed sequence tag simple sequence repeat) markers of black locust can be used as a complement and improvement of Genomic-SSR markers for the identification of the function of gene and the construction of genetic map. Additionally, currently there is no simple method for identifying black locust cultivars. In this study, we obtained 2702 unigenes from 3095 expressed sequence tags (ESTs) from the National Center of Biotechnology Information (NCBI) database to identify simple sequence repeats (SSRs) in R. pseudoacacia samples. A total of 170 SSR loci were found to be distributed in 162 non-redundant sequences with a frequency of 6.29%. Dinucleotide repeats were the most predominant types among microsatellites (62.35%), followed by tri-nucleotide repeats (25.88%); the remaining SSRs accounted for less than 12%. The repeat motifs AG/TC (29.25%) and CT/GA (29.25%) were the most abundant among dinucleotides, and AAT/TTA (15.91%) was the most common among tri-nucleotides. A total of 62 primer pairs were designed to screen polymorphic and stable SSR loci. The resulting 25 EST-SSR markers capable of amplifying polymorphic, stable, and repeatable products. Eight newly developed EST-SSR markers and four published SSR markers were selected for DNA fingerprinting and genetic diversity analysis of the 123 main R. pseudoacacia cultivars in China. The 12 SSR loci amplified 102 alleles, with an average number of alleles per locus of 8.5 (range 4–15). The average polymorphism information content at the 12 SSR loci for the 123 cultivars was 0.670 (range 0.427–0.881). The 123 cultivars clustered into six main groups based on similarity coefficients, with most cultivars in one subgroup. Fingerprinting was performed using eight SSR markers; 110 black locust cultivars were distinguished. The results of this study increase the availability of EST-SSR markers in black locust and make it a simple method for checking the collection, the certification, and the correct attribution of clones and cultivars.

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A SSR kit to study genetic diversity in chickpea (Cicer arietinum L.)

Plant Genetic Resources ◽

10.1017/s1479262114000392 ◽

2014 ◽

Vol 12 (S1) ◽

pp. S118-S120 ◽

Cited By ~ 2

Author(s):

Rajeev Varshney ◽

Mahendar Thudi ◽

Hari Upadhyaya ◽

Sangam Dwivedi ◽

Sripada Udupa ◽

...

Keyword(s):

Genetic Diversity ◽

Information Content ◽

Ssr Markers ◽

Quality Criteria ◽

Specific Information ◽

Wide Range ◽

Ssr Loci ◽

Polymerase Chain ◽

Diversity Studies

A chickpea simple sequence repeat (SSR) marker reference kit has been developed based on the genotyping of the global chickpea composite collection (3,000 accessions) with 35 SSR markers. The kit consists of three pools of chickpea accessions along with supporting documentation on the SSR markers, polymerase chain reaction and detection conditions, and the expected allele sizes for each of the 35 SSR loci. These markers were selected based on quality criteria, genome coverage and locus-specific information content. Other important SSR selection criteria were quality of amplification products, locus complexity, polymorphism information content and well-dispersed location on a chickpea genetic map. The developed SSR kit has a wide range of applications, especially for genetic diversity studies in chickpea. Using the markers and reference accessions in the kit, scientists in other laboratories will be able to compare the genotypic data that they obtain for their germplasm with that obtained using the global composite collection.

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Evaluation of rice and sugarcane SSR markers for phylogenetic and genetic diversity analyses in bambooIHBT Publication No. 0732.

Genome ◽

10.1139/g07-101 ◽

2008 ◽

Vol 51 (2) ◽

pp. 91-103 ◽

Cited By ~ 42

Author(s):

R. K. Sharma ◽

P. Gupta ◽

V. Sharma ◽

A. Sood ◽

T. Mohapatra ◽

...

Keyword(s):

Genetic Diversity ◽

Ssr Markers ◽

Large Scale ◽

Bamboo Species ◽

Ssr Primers ◽

Taxonomical Classification ◽

High Level ◽

The Tropics ◽

Diversity Studies

Simple sequence repeat (SSR) markers are valuable tools for many purposes such as phylogenetic, fingerprinting, and molecular breeding studies. However, only a few SSR markers are known and available in bamboo species of the tropics ( Bambusa spp.). Considering that grass genomes have co-evolved and share large-scale synteny, theoretically it should be possible to use the genome sequence based SSR markers of field crops such as rice ( Oryza sativa ) and sugarcane ( Saccharum spp.) for genome analysis in bamboo. To test this, 98 mapped SSR primers representing 12 linkage groups of rice and 20 EST-derived sugarcane SSR primers were evaluated for transferability to 23 bamboo species. Of the tested markers, 44 (44.9%) rice and 15 (75%) sugarcane SSR primers showed repeatable amplification in at least one species of bamboo and thus were successfully utilized for phylogenetic and genetic diversity analyses. Transferred SSR primers revealed complex amplification patterns in bamboo, with an average of 9.62 fragments per primer, indicating a high level of polyploidy and genetic variability in bamboo. Forty-two of these primers (34 rice and 8 sugarcane SSR primers) detected an average of 2.12 unique fragments per primer and thus could be exploited for species identification. Six bamboo SSR primers exhibited cross transferability, to varying degrees, to different bamboo species. The genetic similarity coefficient indicated a high level of divergence at the species level (73%). However, a relatively low level of diversity was observed within species (25% in 20 accessions of Dendrocalamus hamiltonii ). Further, cluster analysis revealed that the major grouping was in accordance with the taxonomical classification of bamboo. Thus, the rice and sugarcane SSRs can be utilized for phylogenetic and genetic diversity studies in bamboo.

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Study of simple sequence repeat markers from coffee expressed sequences associated to leaf miner resistance

Pesquisa Agropecuária Brasileira ◽

10.1590/s0100-204x2007000300011 ◽

2007 ◽

Vol 42 (3) ◽

pp. 377-384 ◽

Cited By ~ 3

Author(s):

Fernanda de Oliveira Pinto ◽

Mirian Perez Maluf ◽

Oliveiro Guerreiro-Filho

Keyword(s):

Ssr Markers ◽

Expressed Sequence Tags ◽

Defense Mechanisms ◽

Leaf Miner ◽

Gene Transcript ◽

Est Database ◽

Hybrid Frequency ◽

Ssr Loci ◽

Expressed Sequence ◽

Simple Sequence

The objective of this work was to identify expressed simple sequence repeats (SSR) markers associated to leaf miner resistance in coffee progenies. Identification of SSR markers was accomplished by directed searches on the Brazilian Coffee Expressed Sequence Tags (EST) database. Sequence analysis of 32 selected SSR loci showed that 65% repeats are of tetra-, 21% of tri- and 14% of dinucleotides. Also, expressed SSR are localized frequently in the 5'-UTR of gene transcript. Moreover, most of the genes containing SSR are associated with defense mechanisms. Polymorphisms were analyzed in progenies segregating for resistance to the leaf miner and corresponding to advanced generations of a Coffea arabica x Coffea racemosa hybrid. Frequency of SSR alleles was 2.1 per locus. However, no polymorphism associated with leaf miner resistance was identified. These results suggest that marker-assisted selection in coffee breeding should be performed on the initial cross, in which genetic variability is still significant.

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Comparison of Phenolic Compounds and the Antioxidant Activities of Fifteen Chrysanthemum morifolium Ramat cv. ‘Hangbaiju’ in China

Antioxidants ◽

10.3390/antiox8080325 ◽

2019 ◽

Vol 8 (8) ◽

pp. 325 ◽

Cited By ~ 5

Author(s):

Jinyan Gong ◽

Bingquan Chu ◽

Lingxiao Gong ◽

Zhongxiang Fang ◽

Xiaoxu Zhang ◽

...

Keyword(s):

Principal Component Analysis ◽

High Performance Liquid Chromatography ◽

Phenolic Compounds ◽

High Performance ◽

Antioxidant Activities ◽

Principal Component ◽

Chrysanthemum Morifolium ◽

Caffeoylquinic Acids ◽

Caffeoylquinic Acid ◽

Chrysanthemum Morifolium Ramat

This study investigated the phenolic compounds of 15 Chrysanthemum morifolium Ramat cv. ‘Hangbaiju’, including 6 ‘Duoju’ and 9 ‘Taiju’, using high performance liquid chromatography (HPLC). The antioxidant activities of these ‘Hangbaiju’ were estimated by DPPH, ABTS and FRAP assays. Results show that a total of 14 phenolic compounds were detected in these flowers, including 3 mono-caffeoylquinic acids, 3 di-caffeoylquinic acids, 1 phenolic acid and 7 flavonoids. ‘Duoju’ and ‘Taiju’ possess different concentrations of phenolic compounds, and ‘Taiju’ exhibits higher caffeoylquinic acids and stronger antioxidant activities than ‘Duoju’. Caffeoylquinic acids show a strong correlation with the antioxidant activities of the samples. Principal component analysis (PCA) reveals an obvious separation between ‘Duoju’ and ‘Taiju’, using phenolic compounds as variables. Apigenin-7-O-glucoside, 3,5-di-O-caffeoylquinic acid, luteolin and acacetin were found to be the key phenolic compounds to differentiate ‘Duoju’ from ‘Taiju’.

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Diversity, characterization and evaluation in Pummelo (Citrus maxima Merr.) cultivars using SSR markers and quality parameters

Indian Journal of Genetics and Plant Breeding (The) ◽

10.31742/ijgpb.79.3.9 ◽

2019 ◽

Vol 79 (3) ◽

Author(s):

Shahnawaz . Ahmed ◽

H. S. Rattanpal ◽

Gurteg . Singh

Keyword(s):

Genetic Diversity ◽

Ssr Markers ◽

Morphological Variability ◽

Principal Component ◽

Quality Parameters ◽

Average Value ◽

Low Genetic Diversity ◽

Ssr Loci ◽

Citrus Maxima ◽

Maximum Heterozygosity

Fourteen pummelo (Citrus maxima Merr.) fruit varieties were evaluated through morphological and molecular methods to determine the genetic diversity among them. The analysis showed that maximum contribution (60%) towards diversity was due to the number of fruits per tree and rag percentage. Principal component analysis explained 80.26% of the total observed variability. Molecular characterization of pummelo varieties using 60 SSR markers revealed 26 polymorphic SSR loci having 77 amplified alleles and the number of alleles ranged from 1 to 4 with an average of 2.96 alleles per locus. The highest number of alleles per locus recorded was four as amplified by the SSR markers, CAT01, CS05, CCSM70, CIBE5156, AG14, CIBE4728 and CMS26. The PIC value ranged from 0.12 (CIBE5720) to 0.73 (CAT01) with average value of 0.53. Maximum heterozygosity was found in CAT01 (0.73) followed by CS05 (0.72) and AG14 (0.69). Pink Pummelo and White Pummelo showed the highest genetic similarity having coefficient of 89% and were closely related. The present study indicated low genetic diversity in pummelo varieties despite having high morphological variability, which could be elucidated by the fact that much of the phenotypic variation witnessed may be due to somatic mutations.

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Development and characterization of EST-derived simple sequence repeat (SSR) markers for pasture grass endophytes

Genome ◽

10.1139/g03-001 ◽

2003 ◽

Vol 46 (2) ◽

pp. 277-290 ◽

Cited By ~ 33

Author(s):

Eline van Zijll de Jong ◽

Kathryn M Guthridge ◽

German C Spangenberg ◽

John W Forster

Keyword(s):

Genetic Diversity ◽

Ssr Markers ◽

Expressed Sequence Tags ◽

Simple Sequence Repeat ◽

Fragment Length Polymorphism ◽

Sequence Repeat ◽

Pasture Grass ◽

Ssr Loci ◽

Expressed Sequence ◽

Simple Sequence

Fungal endophytes of the genus Neotyphodium are common in temperate pasture grass species and confer both beneficial and deleterious agronomic characteristics to their hosts. The aim of this study was to develop molecular markers based on simple sequence repeat (SSR) loci for the identification and assessment of genetic diversity among Neotyphodium endophytes in grasses. Expressed sequence tags (ESTs) from both Neptyphodium coenophialum and Neotyphodium lolii were examined, and unique SSR loci were identified in 9.7% of the N. coenophialum sequences and 6.3% of the N. lolii sequences. A variety of SSRs were present, although perfect trinucleotide repeat arrays were the most common. Primers were designed to 50 SSR loci from N. coenophialum and 57 SSR loci from N. lolii and were evaluated using 20 Neotyphodium and Epichloë isolates. A high proportion of the N. coenophialum and N. lolii primers produced amplification products from the majority of isolates and most of these primers detected genetic variation. SSR markers from both N. coenophialum and N. lolii detected high levels of polymorphism between Neotyphodium and Epichloë species, and low levels of polymorphism within N. coenophialum and N. lolii. SSR markers may be used in appropriate combinations to discriminate between species. Comparison with amplified fragment length polymorphism (AFLP) data demonstrated that the SSR markers were informative for the assessment of genetic variation within and between endophyte species. These markers may be used to identify endophyte taxa and to evaluate intraspecific population diversity, which may be correlated with variation for endophyte-derived agronomic traits.Key words: Neotyphodium, simple sequence repeats, expressed sequence tags, amplified fragment length polymorphism, genetic diversity.

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Genome survey of Zanthoxylum bungeanum and development of genomic-SSR markers in congeneric species

Bioscience Reports ◽

10.1042/bsr20201101 ◽

2020 ◽

Vol 40 (6) ◽

Author(s):

Jingmiao Li ◽

Siqiao Li ◽

Lijuan Kong ◽

Lihua Wang ◽

Anzhi Wei ◽

...

Keyword(s):

Ssr Markers ◽

Gc Content ◽

Repeat Sequence ◽

Pcr Analysis ◽

Minimum Length ◽

Dinucleotide Repeats ◽

Genome Survey ◽

Genomic Ssr ◽

Ssr Loci ◽

Zanthoxylum Bungeanum

Abstract Zanthoxylum bungeanum, a spice and medicinal plant, is cultivated in many parts of China and some countries in Southeast Asia; however, data on its genome are lacking. In the present study, we performed a whole-genome survey and developed novel genomic-SSR markers of Z. bungeanum. Clean data (∼197.16 Gb) were obtained and assembled into 11185221 scaffolds with an N50 of 183 bp. K-mer analysis revealed that Z. bungeanum has an estimated genome size of 3971.92 Mb, and the GC content, heterozygous rate, and repeat sequence rate are 37.21%, 1.73%, and 86.04%, respectively. These results indicate that the genome of Z. bungeanum is complex. Furthermore, 27153 simple sequence repeat (SSR) loci were identified from 57288 scaffolds with a minimum length > 1 kb. Mononucleotide repeats (19706) were the most abundant type, followed by dinucleotide repeats (5154). The most common motifs were A/T, followed by AT/AT; these SSRs accounted for 71.42% and 11.84% of all repeats, respectively. A total of 21243 non-repeating primer pairs were designed, and 100 were randomly selected and validated by PCR analysis using DNA from 10 Z. bungeanum individuals and 5 Zanthoxylum armatum individuals. Finally, 36 polymorphic SSR markers were developed with polymorphism information content (PIC) values ranging from 0.16 to 0.75. Cluster analysis revealed that Z. bungeanum and Z. armatum could be divided into two major clusters, suggesting that these newly developed SSR markers are useful for genetic diversity and germplasm resource identification in Z. bungeanum and Z. armatum.

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