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Development and Validation of EST-SSR Markers from the Transcriptome of Adzuki Bean (Vigna angularis).

Chen H, Liu L, Wang L, Wang S, Somta P, Cheng X - PLoS ONE (2015)

Bottom Line: Among the unigenes, 14,547 sequences contained a unique simple sequence repeat (SSR) and 3,350 sequences contained more than one SSR.A total of 7,947 EST-SSRs were identified as potential molecular markers, with mono-nucleotide A/T repeats (99.0%) as the most abundant motif class, followed by AG/CT (68.4%), AAG/CTT (30.0%), AAAG/CTTT (26.2%), AAAAG/CTTTT (16.1%), and AACGGG/CCCGTT (6.0%).The large number of SSR-containing sequences and EST-SSR markers will be valuable for genetic analysis of the adzuki bean and related Vigna species.

View Article: PubMed Central - PubMed

Affiliation: The National Key Facility for Crop Gene Resources and Genetic Improvement, Institute of Crop Science, Chinese Academy of Agricultural Sciences, Beijing, 100081, China.

ABSTRACT
The adzuki bean (Vigna angularis (Ohwi) Ohwi and Ohashi) is an important grain legume of Asia. It is cultivated mainly in China, Japan and Korea. Despite its importance, few genomic resources are available for molecular genetic research of adzuki bean. In this study, we developed EST-SSR markers for the adzuki bean through next-generation sequencing. More than 112 million high-quality cDNA sequence reads were obtained from adzuki bean using Illumina paired-end sequencing technology, and the sequences were de novo assembled into 65,950 unigenes. The average length of the unigenes was 1,213 bp. Among the unigenes, 14,547 sequences contained a unique simple sequence repeat (SSR) and 3,350 sequences contained more than one SSR. A total of 7,947 EST-SSRs were identified as potential molecular markers, with mono-nucleotide A/T repeats (99.0%) as the most abundant motif class, followed by AG/CT (68.4%), AAG/CTT (30.0%), AAAG/CTTT (26.2%), AAAAG/CTTTT (16.1%), and AACGGG/CCCGTT (6.0%). A total of 500 SSR markers were randomly selected for validation, of which 296 markers produced reproducible amplicons with 38 polymorphic markers among the 32 adzuki bean genotypes selected from diverse geographical locations across China. The large number of SSR-containing sequences and EST-SSR markers will be valuable for genetic analysis of the adzuki bean and related Vigna species.

No MeSH data available.


Gene ontology (GO) classification of assembled unigenes.
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pone.0131939.g002: Gene ontology (GO) classification of assembled unigenes.

Mentions: For annotation of the sequence assembly contigs and unique singletons, all unigenes were searched against the five databases (see Materials and methods). A total of 65,950 unigenes showed significant BLAST hits. Among those unigenes, 47,009 (71.3%) showed significant similarity to known proteins in the NR sequence database of which 31776 (48.2%) were similar to protein in the Swiss-Prot. A total of 51,131 unigenes were annotated in all databases. Based on annotation against NR database, 11,580 (18.4%) unigenes were assigned to gene ontology (GO) terms (Fig 2). The sequences that belonged to the biological process, cellular component and molecular function clusters were categorized into 50 terms. Under the biological process category, the highest sub-category was cellular process (23,555, 16.1%), followed by metabolic process (23,390, 16.0%), single-organism process (15,823, 10.8%) and locomotion (41, <0.1%). Under the cellular component category, cell component (26,966, 25.2%) and organelle component (21,271, 19.9%) represented the majorities, whereas only a few unigenes were assigned to virion (17, <0.1%), virion part (7, <0.1%) and extracellular matrix part (4, <0.1%). Under the molecular function category, catalytic activity (18,927, 42.1%) and catalytic binding (18,878 42.0%) were prominently represented. Furthermore, 2,637 unigenes were involved in transporter activity, whereas only a few unigenes were assigned to protein tag (3), translation regulator activity (3) and channel regulator activity (2).


Development and Validation of EST-SSR Markers from the Transcriptome of Adzuki Bean (Vigna angularis).

Chen H, Liu L, Wang L, Wang S, Somta P, Cheng X - PLoS ONE (2015)

Gene ontology (GO) classification of assembled unigenes.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4492930&req=5

pone.0131939.g002: Gene ontology (GO) classification of assembled unigenes.
Mentions: For annotation of the sequence assembly contigs and unique singletons, all unigenes were searched against the five databases (see Materials and methods). A total of 65,950 unigenes showed significant BLAST hits. Among those unigenes, 47,009 (71.3%) showed significant similarity to known proteins in the NR sequence database of which 31776 (48.2%) were similar to protein in the Swiss-Prot. A total of 51,131 unigenes were annotated in all databases. Based on annotation against NR database, 11,580 (18.4%) unigenes were assigned to gene ontology (GO) terms (Fig 2). The sequences that belonged to the biological process, cellular component and molecular function clusters were categorized into 50 terms. Under the biological process category, the highest sub-category was cellular process (23,555, 16.1%), followed by metabolic process (23,390, 16.0%), single-organism process (15,823, 10.8%) and locomotion (41, <0.1%). Under the cellular component category, cell component (26,966, 25.2%) and organelle component (21,271, 19.9%) represented the majorities, whereas only a few unigenes were assigned to virion (17, <0.1%), virion part (7, <0.1%) and extracellular matrix part (4, <0.1%). Under the molecular function category, catalytic activity (18,927, 42.1%) and catalytic binding (18,878 42.0%) were prominently represented. Furthermore, 2,637 unigenes were involved in transporter activity, whereas only a few unigenes were assigned to protein tag (3), translation regulator activity (3) and channel regulator activity (2).

Bottom Line: Among the unigenes, 14,547 sequences contained a unique simple sequence repeat (SSR) and 3,350 sequences contained more than one SSR.A total of 7,947 EST-SSRs were identified as potential molecular markers, with mono-nucleotide A/T repeats (99.0%) as the most abundant motif class, followed by AG/CT (68.4%), AAG/CTT (30.0%), AAAG/CTTT (26.2%), AAAAG/CTTTT (16.1%), and AACGGG/CCCGTT (6.0%).The large number of SSR-containing sequences and EST-SSR markers will be valuable for genetic analysis of the adzuki bean and related Vigna species.

View Article: PubMed Central - PubMed

Affiliation: The National Key Facility for Crop Gene Resources and Genetic Improvement, Institute of Crop Science, Chinese Academy of Agricultural Sciences, Beijing, 100081, China.

ABSTRACT
The adzuki bean (Vigna angularis (Ohwi) Ohwi and Ohashi) is an important grain legume of Asia. It is cultivated mainly in China, Japan and Korea. Despite its importance, few genomic resources are available for molecular genetic research of adzuki bean. In this study, we developed EST-SSR markers for the adzuki bean through next-generation sequencing. More than 112 million high-quality cDNA sequence reads were obtained from adzuki bean using Illumina paired-end sequencing technology, and the sequences were de novo assembled into 65,950 unigenes. The average length of the unigenes was 1,213 bp. Among the unigenes, 14,547 sequences contained a unique simple sequence repeat (SSR) and 3,350 sequences contained more than one SSR. A total of 7,947 EST-SSRs were identified as potential molecular markers, with mono-nucleotide A/T repeats (99.0%) as the most abundant motif class, followed by AG/CT (68.4%), AAG/CTT (30.0%), AAAG/CTTT (26.2%), AAAAG/CTTTT (16.1%), and AACGGG/CCCGTT (6.0%). A total of 500 SSR markers were randomly selected for validation, of which 296 markers produced reproducible amplicons with 38 polymorphic markers among the 32 adzuki bean genotypes selected from diverse geographical locations across China. The large number of SSR-containing sequences and EST-SSR markers will be valuable for genetic analysis of the adzuki bean and related Vigna species.

No MeSH data available.