Discovery of polymorphisms in starch-related genes in rice germplasm by amplification of pooled DNA and deeply parallel sequencing

Kharabian-Masouleh, Ardashir, Waters, Daniel L. E., Reinke, Russell F. and Henry, Robert J. (2011) Discovery of polymorphisms in starch-related genes in rice germplasm by amplification of pooled DNA and deeply parallel sequencing. Plant Biotechnology Journal, 9 9: 1074-1085.


Author Kharabian-Masouleh, Ardashir
Waters, Daniel L. E.
Reinke, Russell F.
Henry, Robert J.
Title Discovery of polymorphisms in starch-related genes in rice germplasm by amplification of pooled DNA and deeply parallel sequencing
Journal name Plant Biotechnology Journal   Check publisher's open access policy
ISSN 1467-7644
1467-7652
Publication date 2011-12
Sub-type Article (original research)
DOI 10.1111/j.1467-7652.2011.00629.x
Volume 9
Issue 9
Start page 1074
End page 1085
Total pages 12
Place of publication Oxford, United Kingdom
Publisher Wiley-Blackwell
Collection year 2012
Language eng
Abstract High-throughput sequencing of pooled DNA was applied to polymorphism discovery in candidate genes involved in starch synthesis. This approach employed semi- to long-range PCR (LR-PCR) followed by next-generation sequencing technology. A total of 17 rice starch synthesis genes encoding seven classes of enzymes, including ADP-glucose pyrophosphorylase (AGPase), granule starch synthase (GBSS), soluble starch synthase (SS), starch branching enzyme (BE), starch debranching enzyme (DBE) and starch phosphorylase (SPHOL) and phosphate translocator (GPT1) from 233 genotypes were PCR amplified using semi- to long-range PCR. The amplification products were equimolarly pooled and sequenced using massively parallel sequencing technology (MPS). By detecting single nucleotide polymorphism (SNP) ⁄ Indels in both coding and noncoding areas of the genes, we identified genetic differences and characterized the SNP ⁄ Indel variation and distribution patterns among individual starch candidate genes. Approximately, 60.9 million reads were generated, of which 54.8 million (90%) mapped to the reference sequences. The average coverage rate ranged from 12 708 to 38 300 times for SSIIa and SSIIIb, respectively. SNPs and single ⁄ multiple-base Indels were analysed in a total assembled length of 116 403 bp. In total, 501 SNPs and 113 Indels were detected across the 17 starch-related loci. The ratio of synonymous to nonsynonymous SNPs (Ka ⁄ Ks) test indicated GBSSI and isoamylase 1 (ISA1) as the least diversified (most purified) and conservative genes as the studied populations have been through cycles of selection. This report demonstrates a useful strategy for screening germplasm by MPS to discover variants in a specific target group of genes.
Keyword Starch-related genes
Starch synthase
Granule bound starch synthase
Rice
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Q-Index Code C1
Q-Index Status Confirmed Code
Institutional Status UQ

Document type: Journal Article
Sub-type: Article (original research)
Collections: Official 2012 Collection
Queensland Alliance for Agriculture and Food Innovation
 
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