Genetic Diversity Analysis and Single-nucleotide Polymorphism Marker Development in Cultivated Bulb Onion Based on Expressed Sequence Tag–Simple Sequence Repeat Markers

Abstract

<jats:p>Bulb onion (<jats:italic>Allium cepa</jats:italic> L.) is a globally significant crop, but the structure of genetic variation within and among populations is poorly understood. We broadly surveyed genetic variation in a cultivated onion germplasm using simple sequence repeat (SSR) markers and sequenced regions flanking expressed sequence tag (EST)-SSRs to develop single-nucleotide polymorphism (SNP) markers. Samples from 89 inbred and open-pollinated (OP) bulb onion populations of wide geographical adaptation and four related <jats:italic>Allium</jats:italic> L. accessions were genotyped with 56 EST-SSR and four genomic SSR markers. Multivariate analysis of genetic distances among populations resolved long-day, short-day, and Indian populations. EST-SSR markers frequently revealed two major alleles at high frequency in OP populations. The median proportion of single-locus polymorphic loci was 0.70 in OP and landrace populations compared with 0.43 in inbred lines. Resequencing of 24 marker amplicons revealed additional SNPs in 17 (68%) and five SNP assays were developed from these, suggesting that resequencing of EST markers can readily provide SNP markers for purity testing of inbreds and other applications in <jats:italic>Allium</jats:italic> genetics.</jats:p>

Microsatellite marker based analysis of genetic diversity in short day tropical Indian onion and cross amplification in related Allium spp.

“…In contrast, Araki et al (2010) reported their amplification but it was in bulb onion and closely related species of section Cepa and Phyllodon only. EST-SSR primer set revealed an average of 3.2 amplicons (range, 1-7 alleles) and is equal to the average (3.6) reported by McCallum et al (2008). Two primer sets (ACM300, ACM318) and all the three genomic SSRs were monomorphic in onion accessions but they revealed polymorphism between Fig.…”

“…Currah (2002) explained that short day onions may show greater genetic diversity because they have been maintained as landraces and open pollinated cultivars over a wide geographic area, whereas intermediate and long day cultivars are more localized. Fischer and Bachmann (2000) also revealed groupings based on geographical origins whereas McCallum et al (2008) also observed a clear resolution of a group of Indian cultivars and landraces from the main grouping of short day populations and were of the view that Indian region may have potential to provide novel germplasm resources. The present investigations support this hypothesis and this fact may be further strengthened in future by diversity assessment of short day germplasm from diverse origins.…”

“…Genomic SSRs were less polymorphic (10%) as compared to the EST derived microsatellite (53.3%). Genomic SSRs require optimization of PCR conditions and amplify rarely in onion and allied Alliums McCallum et al 2008;Santos et al 2010). In contrast, Araki et al (2010) reported their amplification but it was in bulb onion and closely related species of section Cepa and Phyllodon only.…”

“…Diversity assessment on the basis of molecular markers is preferred because of their highly polymorphic nature, codominant inheritance, frequent occurrence in genome, selective neutral behaviour, easy access, easy and fast assay, high reproducibility and easy exchange of data between laboratories (Joshi et al 1999). Molecular characterization on the basis of RAPD (Tanikawa et al 2002), RFLP (Havey 1991a;Bradeen and Havey 1995) and recently on the basis of SSR and SNP's McCallum et al 2008) has been published in long day onion. Microsatellite markers have become the markers of choice because they are locus-specific, codominant, PCR-based, highly polymorphic and useful at a range of scales from individual identification to fine-scale phylogenies.…”

“…The only disadvantage in using microsatellites is that prior information about the sequence for developing primer combinations is needed. Fischer and Bachmann (2000) were the first to develop and use genomic microsatellite primers in onion and allied species followed by development of expressed sequence tag (EST) derived microsatellites (Kuhl et al 2004;Martin et al 2005) and their utilisation for mapping (McCallum et al 2006) and cultivar discrimination McCallum et al 2008). Microsatellites have also been identified and characterised in Allium fistulosum L. (Song et al 2004;Tsukazaki et al 2007), garlic (Kim et al 2009) and tested for their applicability in onion and related wild species (Araki et al 2010).…”