K-seq, an affordable, reliable, and open Klenow NGS-based genotyping technology
Abstract Background K-seq, a new genotyping methodology based on the amplification of genomic regions using two steps of Klenow amplification with short oligonucleotides, followed by standard PCR and Illumina sequencing, is presented. The protocol was accompanied by software developed to aid with pr...
| Main Authors: | , , , , , , |
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| Format: | Article |
| Language: | English |
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BMC
2021-03-01
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| Series: | Plant Methods |
| Subjects: | |
| Online Access: | https://doi.org/10.1186/s13007-021-00733-6 |
| _version_ | 1818743288779243520 |
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| author | Peio Ziarsolo Tomas Hasing Rebeca Hilario Victor Garcia-Carpintero Jose Blanca Aureliano Bombarely Joaquin Cañizares |
| author_facet | Peio Ziarsolo Tomas Hasing Rebeca Hilario Victor Garcia-Carpintero Jose Blanca Aureliano Bombarely Joaquin Cañizares |
| author_sort | Peio Ziarsolo |
| collection | DOAJ |
| description | Abstract Background K-seq, a new genotyping methodology based on the amplification of genomic regions using two steps of Klenow amplification with short oligonucleotides, followed by standard PCR and Illumina sequencing, is presented. The protocol was accompanied by software developed to aid with primer set design. Results As the first examples, K-seq in species as diverse as tomato, dog and wheat was developed. K-seq provided genetic distances similar to those based on WGS in dogs. Experiments comparing K-seq and GBS in tomato showed similar genetic results, although K-seq had the advantage of finding more SNPs for the same number of Illumina reads. The technology reproducibility was tested with two independent runs of the tomato samples, and the correlation coefficient of the SNP coverages between samples was 0.8 and the genotype match was above 94%. K-seq also proved to be useful in polyploid species. The wheat samples generated specific markers for all subgenomes, and the SNPs generated from the diploid ancestors were located in the expected subgenome with accuracies greater than 80%. Conclusion K-seq is an open, patent-unencumbered, easy-to-set-up, cost-effective and reliable technology ready to be used by any molecular biology laboratory without special equipment in many genetic studies. |
| first_indexed | 2024-12-18T02:26:02Z |
| format | Article |
| id | doaj.art-07809e5349c54f4188b16a499bd7073f |
| institution | Directory Open Access Journal |
| issn | 1746-4811 |
| language | English |
| last_indexed | 2024-12-18T02:26:02Z |
| publishDate | 2021-03-01 |
| publisher | BMC |
| record_format | Article |
| series | Plant Methods |
| spelling | doaj.art-07809e5349c54f4188b16a499bd7073f2022-12-21T21:24:02ZengBMCPlant Methods1746-48112021-03-0117111110.1186/s13007-021-00733-6K-seq, an affordable, reliable, and open Klenow NGS-based genotyping technologyPeio Ziarsolo0Tomas Hasing1Rebeca Hilario2Victor Garcia-Carpintero3Jose Blanca4Aureliano Bombarely5Joaquin Cañizares6COMAV, Universitat Politècnica de ValènciaSchool of Plant and Environmental Sciences, Virginia TechCOMAV, Universitat Politècnica de ValènciaCOMAV, Universitat Politècnica de ValènciaCOMAV, Universitat Politècnica de ValènciaSchool of Plant and Environmental Sciences, Virginia TechCOMAV, Universitat Politècnica de ValènciaAbstract Background K-seq, a new genotyping methodology based on the amplification of genomic regions using two steps of Klenow amplification with short oligonucleotides, followed by standard PCR and Illumina sequencing, is presented. The protocol was accompanied by software developed to aid with primer set design. Results As the first examples, K-seq in species as diverse as tomato, dog and wheat was developed. K-seq provided genetic distances similar to those based on WGS in dogs. Experiments comparing K-seq and GBS in tomato showed similar genetic results, although K-seq had the advantage of finding more SNPs for the same number of Illumina reads. The technology reproducibility was tested with two independent runs of the tomato samples, and the correlation coefficient of the SNP coverages between samples was 0.8 and the genotype match was above 94%. K-seq also proved to be useful in polyploid species. The wheat samples generated specific markers for all subgenomes, and the SNPs generated from the diploid ancestors were located in the expected subgenome with accuracies greater than 80%. Conclusion K-seq is an open, patent-unencumbered, easy-to-set-up, cost-effective and reliable technology ready to be used by any molecular biology laboratory without special equipment in many genetic studies.https://doi.org/10.1186/s13007-021-00733-6Klenow polymeraseK-seqK-merLow costSNPGenotyping |
| spellingShingle | Peio Ziarsolo Tomas Hasing Rebeca Hilario Victor Garcia-Carpintero Jose Blanca Aureliano Bombarely Joaquin Cañizares K-seq, an affordable, reliable, and open Klenow NGS-based genotyping technology Plant Methods Klenow polymerase K-seq K-mer Low cost SNP Genotyping |
| title | K-seq, an affordable, reliable, and open Klenow NGS-based genotyping technology |
| title_full | K-seq, an affordable, reliable, and open Klenow NGS-based genotyping technology |
| title_fullStr | K-seq, an affordable, reliable, and open Klenow NGS-based genotyping technology |
| title_full_unstemmed | K-seq, an affordable, reliable, and open Klenow NGS-based genotyping technology |
| title_short | K-seq, an affordable, reliable, and open Klenow NGS-based genotyping technology |
| title_sort | k seq an affordable reliable and open klenow ngs based genotyping technology |
| topic | Klenow polymerase K-seq K-mer Low cost SNP Genotyping |
| url | https://doi.org/10.1186/s13007-021-00733-6 |
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