Effects of methylation-sensitive enzymes on the enrichment of genic SNPs and the degree of genome complexity reduction in a two-enzyme genotyping-by-sequencing GBS approach: a case study in oil palm Elaeis guineensisReportar como inadecuado




Effects of methylation-sensitive enzymes on the enrichment of genic SNPs and the degree of genome complexity reduction in a two-enzyme genotyping-by-sequencing GBS approach: a case study in oil palm Elaeis guineensis - Descarga este documento en PDF. Documentación en PDF para descargar gratis. Disponible también para leer online.

Molecular Breeding

, 36:154

First Online: 10 November 2016Received: 20 April 2016Accepted: 20 October 2016

Abstract

Advances in next generation sequencing have facilitated a large-scale single nucleotide polymorphism SNP discovery in many crop species. Genotyping-by-sequencing GBS approach couples next generation sequencing with genome complexity reduction techniques to simultaneously identify and genotype SNPs. Choice of enzymes used in GBS library preparation depends on several factors including the number of markers required, the desired level of multiplexing, and whether the enrichment of genic SNP is preferred. We evaluated various combinations of methylation-sensitive AatII, PstI, MspI and methylation-insensitive SphI, MseI enzymes for their effectiveness in genome complexity reduction and enrichment of genic SNPs. We discovered that the use of two methylation-sensitive enzymes effectively reduced genome complexity and did not require a size selection step. On the contrary, the genome coverage of libraries constructed with methylation-insensitive enzymes was quite high, and the additional size selection step may be required to increase the overall read depth. We also demonstrated the effectiveness of methylation-sensitive enzymes in enriching for SNPs located in genic regions. When two methylation-insensitive enzymes were used, only 16% of SNPs identified were located in genes and 18% in the vicinity ± 5 kb of the genic regions, while most SNPs resided in the intergenic regions. In contrast, a remarkable degree of enrichment was observed when two methylation-sensitive enzymes were employed. Almost two thirds of the SNPs were located either inside 32–36% or in the vicinity 28–31% of the genic regions. These results provide useful information to help researchers choose appropriate GBS enzymes in oil palm and other crop species.

KeywordsSNP discovery SNP genotyping Genotyping-by-sequencing GBS Methylation-sensitive enzyme Reduced representation library Genome complexity reduction Genic SNP enrichment Oil palm Elaeis guineensis Electronic supplementary materialThe online version of this article doi:10.1007-s11032-016-0572-x contains supplementary material, which is available to authorized users.

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Autor: Wirulda Pootakham - Chutima Sonthirod - Chaiwat Naktang - Nukoon Jomchai - Duangjai Sangsrakru - Sithichoke Tangphatsornruan

Fuente: https://link.springer.com/







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