Construction of random small-insert genomic libraries highly enriched for simple sequence repeats

Abstract

Dinucleotide (CA)n repeat sequences are highly abundant and interspersed in eukaryotic genomes. Individual sites or loci can be identified by PCR-based assays using unique sequence oligonucleotides that flank specific CA-repeats. The number of CA-repeats at a given locus is variable making these markers highly informative for genetic analysis in humans (1) and other species (2). Unique sequences flanking specific (CA)n loci are usually identified by analyzing genomic libraries containing small size inserts, suitable for sequencing, generated by restriction enzymes. However the construction and screening of these type

Genetic evaluation of the self-sustaining status of a population of the endangered Danube salmon, Hucho hucho

“…We cloned new tetranucleotide microsatellite loci using a selective hybridization approach (Karagyozov et al, 1993) and streptavidin-coated magnetic beads (Kandpal et al, 1994;Kijas et al, 1994;Paetkau, 1999). This approach was improved by Diniz et al (2007) and McPherson et al (2001), and finally our protocol followed the double-hybridization method as outlined in Winkler & Weiss (2008), including all details of adaptor and probe design.…”

Development and polymorphism of simple sequence repeat DNA markers for Shorea curtisii and other Dipterocarpaceae species

“…Several improvements in methodology have been reported recently for efficient developments of SSR markers (Ostrander et al, 1992;Karagyozov et al, 1993;Lyall et al, 1993;Cifarelli et al, 1995;Kirkpatrick et al, 1995;Takahashi et al, 1996), one of which is the vectorette PCR strategy (Lench et al, 1996). The procedure relies on PCR amplification using a vectorette-specific primer in combination with anchored dinucleotide repeat primers.…”

Methods for Development of Microsatellite Markers: An Overview

“…These protocols generally involve the fragmentation of DNA either by sonication (Karagyozov et al, 1993;Kandpal et al, 1994;Geng et al, 2010) or restriction enzymes (Brown et al, 1995;Chen et al, 1995;Edwards et al, 1996;Prochazka., 1996;Refseth et al, 1997;Fischer and Bachmann, 1998;Hamilton et al, 1999;Glenn and Schable, 2005;Nunome et al, 2006) or nebulisation (Kumpatla et al, 2004;Connell et al, 1998) and its subsequent ligation to a known sequence (linker or adaptors) or directly to a vector. DNA is then denatured and subjected to enrichment by hybridization with a) biotinylated oligos followed by capture of biotinylated hybrids (oligo bound DNA fragments) in vectrex-avidin matrix (Kandpal et al, 1994) or b) oligonucleotides bound to nylon membrane (Karagyozov et al, 1993;Chen et al, 1995;Edwards et al, 1996) or c) 5′ biotinylated repeat oligos and subsequent capture of biotinylated hybrids by streptavidin coated magnetic beads (Brown et al, 1995;Refseth et al, 1997;Fischer and Bachmann, 1998;Connell et al, 1998;Hamilton et al, 1999;Kumpatla et al, 2004;Dixit et al, 2005;Glenn and Schable, 2005;Nunome et al, 2006;Geng et al, 2010) or d) 'biotinylated SSR probe-streptavidin coated magnetic bead complex' ('Triplex affinity capture' protocol; White and Powell, 1997). The enriched DNA fragments were then amplified, either cloned and sequenced or sequenced directly and searched for the presence of SSR motifs.…”