Transcriptome analysis of  sex sorted and unsorted  spermatozoa of indicus cattle (Bos indicus)
Purpose:The transcriptome of sexed (X-chromosome and Y-chromosome bearing) spermatozoa of indicus cattle (Bos indicus) have not been known yet. This study aims to reveal the transcriptome of indicus cattle for better understanding of genetics, physiology and reproduction.Methods:NEBNext Ultra II Directional RNA library Prep Kit was used to prepare the high quality libraries and paired end sequencing reads of read length 151 bp were generated using Illumina HiSeq-X sequencing platform. The raw reads were checked individually by using FASTQC. Quality control and preprocessing of FASTQ files were done to clean data for downstream analysis. A cutoff of 30 was set for the quality Phred score and only high quality reads were retained. The high quality reads were aligned against NCBI Reference of Bos indicus (assembly Bos_indicus_1.0) by using ultra-fast, splice-ware aligner Hisat2.Results :The X-chromosome bearing spermatozoa, Y-Chromosome bearing spermatozoa and unsorted spermatozoa aligned >71%, >74% and >80% respectively with reference Bos indicus assembly (Bos_indicus_1.0). The transcript counts, obtained from each sample were further used for analysis of differential expression of transcripts between alternate conditions by using DESeq2. A p-value cut-off of 0.05 and less was used to identify the significantly expressed transcripts and a log2 Fold change cut-off (+2) and higher for upregulated transcripts and (-2) and less for down regulated transcripts were used.Conclusion:We provide here the complete and comprehensive transcriptome of X- and Y-sorted spermatozoa of indicus cattle (Bos indicus) which is the first reported X- and Y-sorted spermatozoal transcriptome among any eutherian mammals. This will help to focus on subset of genes and transcripts of specific interest, characterize transcriptional activities, other than serving a global picture of cell function.