Development of type I genetic markers from expressed sequence tags in highly polymorphic species

Kim, Woo-Jin, Jung, Hyungtaek, & Gaffney, Patrick (2011) Development of type I genetic markers from expressed sequence tags in highly polymorphic species. Marine Biotechnology, 13(2), pp. 127-132.

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Abstract

Expressed sequence tag (EST) databases provide a primary source of nuclear DNA sequences for genetic marker development in non-model organisms. To date, the process has been relatively inefficient for several reasons:

  • 1) priming site polymorphism in the template leads to inferior or erratic amplification;

  • 2) introns in the target amplicon are too large and/or numerous to allow effective amplification under standard screening conditions, and;

  • 3) at least occasionally, a PCR primer straddles an exon–intron junction and is unable to bind to genomic DNA template.

The first is only a minor issue for species or strains with low heterozygosity but becomes a significant problem for species with high genomic variation, such as marine organisms with extremely large effective population sizes. Problems arising from unanticipated introns are unavoidable but are most pronounced in intron-rich species, such as vertebrates and lophotrochozoans. We present an approach to marker development in the Pacific oyster Crassostrea gigas, a highly polymorphic and intron-rich species, which minimizes these problems, and should be applicable to other non-model species for which EST databases are available. Placement of PCR primers in the 3′ end of coding sequence and 3′ UTR improved PCR success rate from 51% to 97%. Almost all (37 of 39) markers developed for the Pacific oyster were polymorphic in a small test panel of wild and domesticated oysters.

Impact and interest:

16 citations in Scopus
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15 citations in Web of Science®

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ID Code: 92882
Item Type: Journal Article
Refereed: Yes
Keywords: EST, SNP, Marker, Indel, UTR
DOI: 10.1007/s10126-010-9280-4
ISSN: 1436-2236
Subjects: Australian and New Zealand Standard Research Classification > BIOLOGICAL SCIENCES (060000)
Australian and New Zealand Standard Research Classification > TECHNOLOGY (100000) > AGRICULTURAL BIOTECHNOLOGY (100100) > Agricultural Marine Biotechnology (100102)
Divisions: Current > Research Centres > Centre for Tropical Crops and Biocommodities
Current > Schools > School of Earth, Environmental & Biological Sciences
Current > Institutes > Institute for Future Environments
Current > QUT Faculties and Divisions > Science & Engineering Faculty
Deposited On: 18 Feb 2016 23:01
Last Modified: 03 Mar 2016 05:41

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