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Development and preliminary evaluation of a 90 K Axiom® SNP array for the allo-octoploid cultivated strawberry Fragaria × ananassa.

Bassil NV, Davis TM, Zhang H, Ficklin S, Mittmann M, Webster T, Mahoney L, Wood D, Alperin ES, Rosyara UR, Koehorst-Vanc Putten H, Monfort A, Sargent DJ, Amaya I, Denoyes B, Bianco L, van Dijk T, Pirani A, Iezzoni A, Main D, Peace C, Yang Y, Whitaker V, Verma S, Bellon L, Brew F, Herrera R, van de Weg E - BMC Genomics (2015)

Bottom Line: Strategies and filtering pipelines were developed to identify and incorporate markers of several types: di-allelic SNPs (66.6%), multi-allelic SNPs (1.8%), indels (10.1%), and ploidy-reducing "haploSNPs" (11.7%).The array's high success rate is likely driven by the presence of naturally occurring variation in ploidy level within the nominally octoploid genome, and by effectiveness of the employed array design and ploidy-reducing strategies.This array enables genetic analyses including generation of high-density linkage maps, identification of quantitative trait loci for economically important traits, and genome-wide association studies, thus providing a basis for marker-assisted breeding in this high value crop.

View Article: PubMed Central - PubMed

Affiliation: USDA-ARS, NCGR, Corvallis, OR, USA. nahla.bassil@ars.usda.gov.

ABSTRACT

Background: A high-throughput genotyping platform is needed to enable marker-assisted breeding in the allo-octoploid cultivated strawberry Fragaria × ananassa. Short-read sequences from one diploid and 19 octoploid accessions were aligned to the diploid Fragaria vesca 'Hawaii 4' reference genome to identify single nucleotide polymorphisms (SNPs) and indels for incorporation into a 90 K Affymetrix® Axiom® array. We report the development and preliminary evaluation of this array.

Results: About 36 million sequence variants were identified in a 19 member, octoploid germplasm panel. Strategies and filtering pipelines were developed to identify and incorporate markers of several types: di-allelic SNPs (66.6%), multi-allelic SNPs (1.8%), indels (10.1%), and ploidy-reducing "haploSNPs" (11.7%). The remaining SNPs included those discovered in the diploid progenitor F. iinumae (3.9%), and speculative "codon-based" SNPs (5.9%). In genotyping 306 octoploid accessions, SNPs were assigned to six classes with Affymetrix's "SNPolisher" R package. The highest quality classes, PolyHigh Resolution (PHR), No Minor Homozygote (NMH), and Off-Target Variant (OTV) comprised 25%, 38%, and 1% of array markers, respectively. These markers were suitable for genetic studies as demonstrated in the full-sib family 'Holiday' × 'Korona' with the generation of a genetic linkage map consisting of 6,594 PHR SNPs evenly distributed across 28 chromosomes with an average density of approximately one marker per 0.5 cM, thus exceeding our goal of one marker per cM.

Conclusions: The Affymetrix IStraw90 Axiom array is the first high-throughput genotyping platform for cultivated strawberry and is commercially available to the worldwide scientific community. The array's high success rate is likely driven by the presence of naturally occurring variation in ploidy level within the nominally octoploid genome, and by effectiveness of the employed array design and ploidy-reducing strategies. This array enables genetic analyses including generation of high-density linkage maps, identification of quantitative trait loci for economically important traits, and genome-wide association studies, thus providing a basis for marker-assisted breeding in this high value crop.

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Related in: MedlinePlus

Matching and non-matching HD-20 genotypes obtained by comparing sequence-derived to array-obtained genotypes. Genotype data was not available for HolKor 2557 as the genotype calls were < 97%. No genotype comparison was possible when read depth at the variant site was less than 20 (green bars).
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Fig13: Matching and non-matching HD-20 genotypes obtained by comparing sequence-derived to array-obtained genotypes. Genotype data was not available for HolKor 2557 as the genotype calls were < 97%. No genotype comparison was possible when read depth at the variant site was less than 20 (green bars).

Mentions: For seven of the eight HD-20 filtration panel members (excluding HolKor 2557 for which genotype data was not available), comparisons were made between sequence-derived genotype calls and those of 9,186 di-allelic PHR SNPs. In total, 64,302 comparisons were attempted (9,186 SNPs x 7 individuals). A comparison could not be made if the SNP had “Nocall” in the detection panel for a given individual, fewer than 20 sequencing reads were present at the SNP location, or the SNP had more than two alleles at the SNP location. At least one comparison was possible for all but 22 of the 9,186 SNPs. These SNPs failed to meet the requirements for all seven individuals of the HD-20 panel. In total, 50,457 comparisons were possible, and of these, 43,153 (85.5%) had the same genotype calls from both the Axiom array and sequencing data. Of the 7,304 (14.5% of total) comparisons that did not have matching genotype calls, 6,425 (87.9%) were heterozygous based on sequencing and had homozygous Axiom array genotype calls, likely caused by sequencing or genotyping error. Another 839 (11.5%) appeared homozygous based on sequencing, and were heterozygous based on array genotyping, possibly due to absence of allele representation of alternate allele in sequence or presence of signal from paralogous sequences; and 40 (0.5%) had alternative homozygous genotypes. The number of comparisons for an individual appeared to be highly correlated with sequence read coverage. A smaller number of SNPs were compared in the HD-20 member with the lowest genome sequence coverage (HolKor 2549), as opposed to the remaining six HD-20 with higher genome coverage (Table 1, Figure 13).Figure 13


Development and preliminary evaluation of a 90 K Axiom® SNP array for the allo-octoploid cultivated strawberry Fragaria × ananassa.

Bassil NV, Davis TM, Zhang H, Ficklin S, Mittmann M, Webster T, Mahoney L, Wood D, Alperin ES, Rosyara UR, Koehorst-Vanc Putten H, Monfort A, Sargent DJ, Amaya I, Denoyes B, Bianco L, van Dijk T, Pirani A, Iezzoni A, Main D, Peace C, Yang Y, Whitaker V, Verma S, Bellon L, Brew F, Herrera R, van de Weg E - BMC Genomics (2015)

Matching and non-matching HD-20 genotypes obtained by comparing sequence-derived to array-obtained genotypes. Genotype data was not available for HolKor 2557 as the genotype calls were < 97%. No genotype comparison was possible when read depth at the variant site was less than 20 (green bars).
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4374422&req=5

Fig13: Matching and non-matching HD-20 genotypes obtained by comparing sequence-derived to array-obtained genotypes. Genotype data was not available for HolKor 2557 as the genotype calls were < 97%. No genotype comparison was possible when read depth at the variant site was less than 20 (green bars).
Mentions: For seven of the eight HD-20 filtration panel members (excluding HolKor 2557 for which genotype data was not available), comparisons were made between sequence-derived genotype calls and those of 9,186 di-allelic PHR SNPs. In total, 64,302 comparisons were attempted (9,186 SNPs x 7 individuals). A comparison could not be made if the SNP had “Nocall” in the detection panel for a given individual, fewer than 20 sequencing reads were present at the SNP location, or the SNP had more than two alleles at the SNP location. At least one comparison was possible for all but 22 of the 9,186 SNPs. These SNPs failed to meet the requirements for all seven individuals of the HD-20 panel. In total, 50,457 comparisons were possible, and of these, 43,153 (85.5%) had the same genotype calls from both the Axiom array and sequencing data. Of the 7,304 (14.5% of total) comparisons that did not have matching genotype calls, 6,425 (87.9%) were heterozygous based on sequencing and had homozygous Axiom array genotype calls, likely caused by sequencing or genotyping error. Another 839 (11.5%) appeared homozygous based on sequencing, and were heterozygous based on array genotyping, possibly due to absence of allele representation of alternate allele in sequence or presence of signal from paralogous sequences; and 40 (0.5%) had alternative homozygous genotypes. The number of comparisons for an individual appeared to be highly correlated with sequence read coverage. A smaller number of SNPs were compared in the HD-20 member with the lowest genome sequence coverage (HolKor 2549), as opposed to the remaining six HD-20 with higher genome coverage (Table 1, Figure 13).Figure 13

Bottom Line: Strategies and filtering pipelines were developed to identify and incorporate markers of several types: di-allelic SNPs (66.6%), multi-allelic SNPs (1.8%), indels (10.1%), and ploidy-reducing "haploSNPs" (11.7%).The array's high success rate is likely driven by the presence of naturally occurring variation in ploidy level within the nominally octoploid genome, and by effectiveness of the employed array design and ploidy-reducing strategies.This array enables genetic analyses including generation of high-density linkage maps, identification of quantitative trait loci for economically important traits, and genome-wide association studies, thus providing a basis for marker-assisted breeding in this high value crop.

View Article: PubMed Central - PubMed

Affiliation: USDA-ARS, NCGR, Corvallis, OR, USA. nahla.bassil@ars.usda.gov.

ABSTRACT

Background: A high-throughput genotyping platform is needed to enable marker-assisted breeding in the allo-octoploid cultivated strawberry Fragaria × ananassa. Short-read sequences from one diploid and 19 octoploid accessions were aligned to the diploid Fragaria vesca 'Hawaii 4' reference genome to identify single nucleotide polymorphisms (SNPs) and indels for incorporation into a 90 K Affymetrix® Axiom® array. We report the development and preliminary evaluation of this array.

Results: About 36 million sequence variants were identified in a 19 member, octoploid germplasm panel. Strategies and filtering pipelines were developed to identify and incorporate markers of several types: di-allelic SNPs (66.6%), multi-allelic SNPs (1.8%), indels (10.1%), and ploidy-reducing "haploSNPs" (11.7%). The remaining SNPs included those discovered in the diploid progenitor F. iinumae (3.9%), and speculative "codon-based" SNPs (5.9%). In genotyping 306 octoploid accessions, SNPs were assigned to six classes with Affymetrix's "SNPolisher" R package. The highest quality classes, PolyHigh Resolution (PHR), No Minor Homozygote (NMH), and Off-Target Variant (OTV) comprised 25%, 38%, and 1% of array markers, respectively. These markers were suitable for genetic studies as demonstrated in the full-sib family 'Holiday' × 'Korona' with the generation of a genetic linkage map consisting of 6,594 PHR SNPs evenly distributed across 28 chromosomes with an average density of approximately one marker per 0.5 cM, thus exceeding our goal of one marker per cM.

Conclusions: The Affymetrix IStraw90 Axiom array is the first high-throughput genotyping platform for cultivated strawberry and is commercially available to the worldwide scientific community. The array's high success rate is likely driven by the presence of naturally occurring variation in ploidy level within the nominally octoploid genome, and by effectiveness of the employed array design and ploidy-reducing strategies. This array enables genetic analyses including generation of high-density linkage maps, identification of quantitative trait loci for economically important traits, and genome-wide association studies, thus providing a basis for marker-assisted breeding in this high value crop.

Show MeSH
Related in: MedlinePlus