Use of targeted amplicon sequencing in peanut to generate allele information on allotetraploid sub-genomes

Roshan Kulkarni; Ratan Chopra; Jennifer Chagoya; Charles E. Simpson; Michael R. Baring; Andrew Hillhouse; Naveen Puppala; Kelly Chamberlin; Mark D. Burow

doi:10.3390/genes11101220

Use of targeted amplicon sequencing in peanut to generate allele information on allotetraploid sub-genomes

Roshan Kulkarni, Ratan Chopra, Jennifer Chagoya, Charles E. Simpson, Michael R. Baring, Andrew Hillhouse, Naveen Puppala, Kelly Chamberlin, Mark D. Burow

Agronomy and Plant Genetics

Research output: Contribution to journal › Article › peer-review

2 Scopus citations

Abstract

The use of molecular markers in plant breeding has become a routine practice, but the cost per accession can be a hindrance to the routine use of Quantitative Trait Loci (QTL) identification in breeding programs. In this study, we demonstrate the use of targeted re-sequencing as a proof of concept of a cost-effective approach to retrieve highly informative allele information, as well as develop a bioinformatics strategy to capture the genome-specific information of a polyploid species. SNPs were identified from alignment of raw transcriptome reads (2 × 50 bp) to a synthetic tetraploid genome using BWA followed by a GATK pipeline. Regions containing high polymorphic SNPs in both A genome and B genomes were selected as targets for the resequencing study. Targets were amplified using multiplex PCR followed by sequencing on an Illumina HiSeq. Eighty-one percent of the SNP calls in diploids and 68% of the SNP calls in tetraploids were confirmed. These results were also confirmed by KASP validation. Based on this study, we find that targeted resequencing technologies have potential for obtaining maximum allele information in allopolyploids at reduced cost.

Original language	English (US)
Article number	1220
Pages (from-to)	1-18
Number of pages	18
Journal	Genes
Volume	11
Issue number	10
DOIs	https://doi.org/10.3390/genes11101220
State	Published - 2020

Bibliographical note

Funding Information:
This research was supported by the Peanut Foundation, the National Peanut Board and USDA National Institute of Food and Agriculture Hatch project TEX08835 to MDB. The Peanut Foundation specifically supported the targeted resequencing and provided graduate student support. Grant funds allow for payment of publication cost. Genotyping and technical support were provided by funding from the National Peanut Board and NIFA. Mention of trade names or commercial products in this article is solely for the purpose of providing specific information and does not imply recommendation or endorsement by the U.S. Department of Agriculture. Any opinions, findings, conclusions or recommendations expressed in this publication are those of the author(s) and do not necessarily reflect the view of the U.S. Department of Agriculture. The authors wish to thank Petar Grozdanov at the Texas Tech Health Sciences Core Facilities Center and Steven Hoffman of Fluidigm Corporation for technical assistance.

Funding Information:
Funding: This research was supported by the Peanut Foundation, the National Peanut Board and USDA National Institute of Food and Agriculture Hatch project TEX08835 to MDB. The Peanut Foundation specifically supported the targeted resequencing and provided graduate student support. Grant funds allow for payment of publication cost. Genotyping and technical support were provided by funding from the National Peanut Board and NIFA. Mention of trade names or commercial products in this article is solely for the purpose of providing specific information and does not imply recommendation or endorsement by the U.S. Department of Agriculture. Any opinions, findings, conclusions or recommendations expressed in this publication are those of the author(s) and do not necessarily reflect the view of the U.S. Department of Agriculture.

Publisher Copyright:
© 2020 by the authors. Licensee MDPI, Basel, Switzerland.

Keywords

Allopolyploid
Heterozygous SNP calls
Targeted resequencing
Tetraploids

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title = "Use of targeted amplicon sequencing in peanut to generate allele information on allotetraploid sub-genomes",

abstract = "The use of molecular markers in plant breeding has become a routine practice, but the cost per accession can be a hindrance to the routine use of Quantitative Trait Loci (QTL) identification in breeding programs. In this study, we demonstrate the use of targeted re-sequencing as a proof of concept of a cost-effective approach to retrieve highly informative allele information, as well as develop a bioinformatics strategy to capture the genome-specific information of a polyploid species. SNPs were identified from alignment of raw transcriptome reads (2 × 50 bp) to a synthetic tetraploid genome using BWA followed by a GATK pipeline. Regions containing high polymorphic SNPs in both A genome and B genomes were selected as targets for the resequencing study. Targets were amplified using multiplex PCR followed by sequencing on an Illumina HiSeq. Eighty-one percent of the SNP calls in diploids and 68% of the SNP calls in tetraploids were confirmed. These results were also confirmed by KASP validation. Based on this study, we find that targeted resequencing technologies have potential for obtaining maximum allele information in allopolyploids at reduced cost.",

keywords = "Allopolyploid, Heterozygous SNP calls, Targeted resequencing, Tetraploids",

author = "Roshan Kulkarni and Ratan Chopra and Jennifer Chagoya and Simpson, {Charles E.} and Baring, {Michael R.} and Andrew Hillhouse and Naveen Puppala and Kelly Chamberlin and Burow, {Mark D.}",

note = "Funding Information: This research was supported by the Peanut Foundation, the National Peanut Board and USDA National Institute of Food and Agriculture Hatch project TEX08835 to MDB. The Peanut Foundation specifically supported the targeted resequencing and provided graduate student support. Grant funds allow for payment of publication cost. Genotyping and technical support were provided by funding from the National Peanut Board and NIFA. Mention of trade names or commercial products in this article is solely for the purpose of providing specific information and does not imply recommendation or endorsement by the U.S. Department of Agriculture. Any opinions, findings, conclusions or recommendations expressed in this publication are those of the author(s) and do not necessarily reflect the view of the U.S. Department of Agriculture. The authors wish to thank Petar Grozdanov at the Texas Tech Health Sciences Core Facilities Center and Steven Hoffman of Fluidigm Corporation for technical assistance. Funding Information: Funding: This research was supported by the Peanut Foundation, the National Peanut Board and USDA National Institute of Food and Agriculture Hatch project TEX08835 to MDB. The Peanut Foundation specifically supported the targeted resequencing and provided graduate student support. Grant funds allow for payment of publication cost. Genotyping and technical support were provided by funding from the National Peanut Board and NIFA. Mention of trade names or commercial products in this article is solely for the purpose of providing specific information and does not imply recommendation or endorsement by the U.S. Department of Agriculture. Any opinions, findings, conclusions or recommendations expressed in this publication are those of the author(s) and do not necessarily reflect the view of the U.S. Department of Agriculture. Publisher Copyright: {\textcopyright} 2020 by the authors. Licensee MDPI, Basel, Switzerland.",

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AU - Baring, Michael R.

AU - Hillhouse, Andrew

AU - Puppala, Naveen

AU - Chamberlin, Kelly

AU - Burow, Mark D.

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N2 - The use of molecular markers in plant breeding has become a routine practice, but the cost per accession can be a hindrance to the routine use of Quantitative Trait Loci (QTL) identification in breeding programs. In this study, we demonstrate the use of targeted re-sequencing as a proof of concept of a cost-effective approach to retrieve highly informative allele information, as well as develop a bioinformatics strategy to capture the genome-specific information of a polyploid species. SNPs were identified from alignment of raw transcriptome reads (2 × 50 bp) to a synthetic tetraploid genome using BWA followed by a GATK pipeline. Regions containing high polymorphic SNPs in both A genome and B genomes were selected as targets for the resequencing study. Targets were amplified using multiplex PCR followed by sequencing on an Illumina HiSeq. Eighty-one percent of the SNP calls in diploids and 68% of the SNP calls in tetraploids were confirmed. These results were also confirmed by KASP validation. Based on this study, we find that targeted resequencing technologies have potential for obtaining maximum allele information in allopolyploids at reduced cost.

AB - The use of molecular markers in plant breeding has become a routine practice, but the cost per accession can be a hindrance to the routine use of Quantitative Trait Loci (QTL) identification in breeding programs. In this study, we demonstrate the use of targeted re-sequencing as a proof of concept of a cost-effective approach to retrieve highly informative allele information, as well as develop a bioinformatics strategy to capture the genome-specific information of a polyploid species. SNPs were identified from alignment of raw transcriptome reads (2 × 50 bp) to a synthetic tetraploid genome using BWA followed by a GATK pipeline. Regions containing high polymorphic SNPs in both A genome and B genomes were selected as targets for the resequencing study. Targets were amplified using multiplex PCR followed by sequencing on an Illumina HiSeq. Eighty-one percent of the SNP calls in diploids and 68% of the SNP calls in tetraploids were confirmed. These results were also confirmed by KASP validation. Based on this study, we find that targeted resequencing technologies have potential for obtaining maximum allele information in allopolyploids at reduced cost.

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Use of targeted amplicon sequencing in peanut to generate allele information on allotetraploid sub-genomes

Abstract

Bibliographical note

Keywords

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