Genotyping by Sequencing for Crop Improvement. Группа авторов

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Название Genotyping by Sequencing for Crop Improvement
Автор произведения Группа авторов
Жанр Биология
Серия
Издательство Биология
Год выпуска 0
isbn 9781119745679



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SNP. The range of SNPs to be genotyped ranges from 10k to 2000k SNPs. As gene chip requires high‐quality DNA, so for GeneChip assays to work efficiently, the complexity of the genomic DNA must be reduced through digestion with restriction endonucleases and fractionation. In previous years, researchers used a high‐density oligonucleotide probes array to genotype the entire genome, paving the way for genome‐wide association studies (GWAS) (Kennedy et al. 2003; Matsuzaki et al. 2004; McGall and Christians 2002). Affymetrix GeneChip array contains a large number of synthetic fragments (25‐mer) probes immobilized on a solid substrate. First, the denatured ssDNA are hybridized with these probes, which takes place in a highly specific manner as noncomplementary alleles will not hybridize. Subsequently, the fragments of noncomplementary strands are washed away. The hybrid strands containing probes are stained and then the gene chip is subjected to the CCD imaging device for scanning. Each SNP is represented by a probe set that contains multiple probe pairs. The probe pairs differ in the location of SNP within the oligonucleotide sequence (5 location’s probes are selected). For each position, probes are included from the sense and antisense strand. Hence, the total probes for each allele are 40 probes per SNP.

Crop Species Array fsize References Trait
Pigeonpea (Cajanus cajan) 62K Singh et al. (2020) Genetic diversity
Wheat (Triticum) 55K Zhang et al. (2019) Adult‐plant resistance to leaf and stripe rust
Apple (Malus domestica) 480K Bianco et al. (2016) Phenology, fruit quality, disease resistance, or drought tolerance
Apple (Malus domestica) 8K Chagné et al. (2012) Quantitative traits
Rice (Oryza sativa) 50K Singh et al. (2015) Genetic diversity
Rice (Oryza sativa) 44 K Zhao et al. (2011) Plant morphology, grain quality, plant development
Rye (Secale cereale) 5 K Li et al. (2011) Diversity analysis
Barley (Hordeum vulgare) 50 K Bayer et al. (2017) Evaluation and use of barley genetic resources
Sweet Cherry (Prunus avium) 6K Peace et al. (2012) Fruit taste
Pear (Prunus) 70K Montanari et al. (2019) Genetic diversity studies
Potato (Solanum tuberosum) 22K Khlestkin et al. (2019) Starch phosphorylation
Pear (Pyrus) 200K Li et al. (2019) Flowering time and candidate genes linked to the size of fruit
Walnut (Juglans regia) 700K Marrano et al. (2019) Genetic diversity of germplasm
Cotton (Gossypium barbadense) 63K Kumar et al. (2019) Fiber quality

       2.2.2.1.4 Sequencing‐based Platforms

      The simplest method of genotyping is whole‐genome resequencing of genotypes followed by the reference‐based assembly for variant calling. But as the majority of the genome is noncoding, these noncoding regions are also reflected in the variant file generated by the mapping of the reads which in turn increases the complexity of analysis as well as interpretation. Reduced representation approaches like RAD‐seq and GBS methods have solved this issue as in spite of sequencing the whole genome, these methods are confined to coding/genic regions only. For this, we first digest the DNA with methylation‐sensitive restriction enzyme with the hypothesis that DNA is more methylated in the noncoding region hence the digestion will be limited to coding regions only. The restricted ends generated are then ligated with adapters and sequenced. Other widely used sequencing‐based technologies are exome sequencing (Exom‐seq) and double digest RAD‐seq (ddRAD‐seq).

      Restriction Site‐Associated DNA (RAD‐seq)