Single Nucleotide Polymorphism Discovery in Bovine Pituitary Gland Using RNA-Seq Technology

Abstract

Examination of bovine pituitary gland transcriptome by strand-specific RNA-seq allows detection of putative single nucleotide polymorphisms (SNPs) within potential candidate genes (CGs) or QTLs regions as well as to understand the genomics variations that contribute to economic trait. Here we report a breed-specific model to successfully perform the detection of SNPs in the pituitary gland of young growing bulls representing Polish Holstein-Friesian (HF), Polish Red, and Hereford breeds at three developmental ages viz., six months, nine months, and twelve months. A total of 18 bovine pituitary gland polyA transcriptome libraries were prepared and sequenced using the Illumina NextSeq 500 platform. Sequenced FastQ databases of all 18 young bulls were submitted to NCBI-SRA database with NCBI-SRA accession numbers SRS1296732. For the investigated young bulls, a total of 113,882,3098 raw paired-end reads with a length of 156 bases were obtained, resulting in an approximately 63 million paired-end reads per library. Breed-wise, a total of 515.38, 215.39, and 408.04 million paired-end reads were obtained for Polish HF, Polish Red, and Hereford breeds, respectively. Burrows-Wheeler Aligner (BWA) read alignments showed 93.04%, 94.39%, and 83.46% of the mapped sequencing reads were properly paired to the Polish HF, Polish Red, and Hereford breeds, respectively. Constructed breed-specific SNP-db of three cattle breeds yielded at 13,775,885 SNPs. On an average 765,326 breed-specific SNPs per young bull were identified. Using two stringent filtering parameters, i.e., a minimum 10 SNP reads per base with an accuracy ≥ 90% and a minimum 10 SNP reads per base with an accuracy = 100%, SNP-db records were trimmed to construct a highly reliable SNP-db. This resulted in a reduction of 95,7% and 96,4% cut-off mark of constructed raw SNP-db. Finally, SNP discoveries using RNA-Seq data were validated by KASP™ SNP genotyping assay. The comprehensive QTLs/CGs analysis of 76 QTLs/CGs with RNA-seq data identified KCNIP4, CCSER1, DPP6, MAP3K5 and GHR CGs with highest SNPs hit loci in all three breeds and developmental ages. However, CAST CG with more than 100 SNPs hits were observed only in Polish HF and Hereford breeds.These findings are important for identification and construction of novel tissue specific SNP-db and breed specific SNP-db dataset by screening of putative SNPs according to QTL db and candidate genes for bovine growth and reproduction traits, one can develop genomic selection strategies for growth and reproductive traits.

RNA-seq based SNP discovery in gluteus medius muscle of Polish Landrace pigs

“…iN VeTeRiNARy ScieNce Vol 2, No 1, 2019 Background With the rapid advancement in high-throughput (HT) next-generation genome sequencing (NGS), numerous studies were performed in last decade, to discovery the de novo and reference-based Single nucleotide polymorphisms (SNPs) discovery in cattle and pig domesticated animals [1][2][3][4][5][6][7][8][9]. The SNPs are generally the single nucleotide variations (SNVs) caused either by transitions (C/T or G/A) or transversions (C/G, C/A, or T/A, T/G), in the same position between individual genomic DNA sequences [10][11].…”

“…It has been proved that considerable effects on protein function and gene expression can be caused by the both synonymous and nonsynonymous cSNPs, particularly at the regulatory coding regions. That is why, cSNP as a genetic marker has great potential in genetics and breeding studies in veterinary sciences [3][4]. Due to the high density, scalability and genomewide distribution, SNPs are considered as ideal genetic markers to characterize the genetic resources and functional genes associated with economic traits [15].…”

“…Moreover, RNA-seq is also a capable to perform the transcriptome studies such as gene characterization, gene expression quantification as well as post translational process analysis [21]. In this presented paper, we have performed the transcriptome sequencing of the gluteus medius muscle of Polish Landrace pigs and utilized the MiSeq illumina NGS platform known for its good sequencing coverage, read quality and SNP detection [3][4]. Moreover, a detailed mapping statistics of gluteus medius muscle transcriptome of Polish Landrace pig is analyzed and presented in Table 2.…”

RNA-seq based SNP discovery in liver transcriptome of Polish Landrace pigs

“…High-throughput next-generation genome sequencing (HT-NGS) based RNA-seq studies in past decade, allow us to detect numerous de novo and reference-based single nucleotide polymorphisms (SNPs) discoveries in cattle and pig domesticated animals [1][2][3][4][5][6][7][8][9]. The SNPs detection in any transcriptome are the single nucleotide variations (SNVs) caused by transitions (C/T or G/A) or transversions (C/G, C/A, or T/A, T/G) in the genome [10][11].…”

“…In mammals, synonymous cSNPs (affecting coding region) and nonsynonymous cSNPs (affecting protein sequence) have considerable effects on protein function and gene expression, particularly at the regulatory coding regions. Therefore, cSNP as a genetic marker has great impact and significance in genetics and breeding studies in veterinary sciences [3][4]. Due to the high density, scalability in the genome and genome-wide distribution, SNPs are the best genetic TRANSLATioNAL ReSeARch iN VeTeRiNARy ScieNce Vol 2, No 1, 2019 markers to characterize the genetic resources and functional genes associated with economic traits [15] and to identify the trait-associated variation within the domesticated animals [7][8][9], as well as, to discover genes linked to complex genetic traits in human [16][17].…”