Advancing the Indian Cattle Pangenome: Characterizing Non-Reference Sequences in Bos indicus

BackgroundIndia, with the worlds largest cattle population and more than 50 registered breeds of Bos indicus, stands as a vital reservoir of genetic diversity. However, the abundant diversity among Indian cattle breeds highlights the inadequacy of a single reference sequence to represent the entire genomic content of desi cattle. We recognize the need to capture the genomic differences within the Bos indicus population as a whole, and specifically within the dairy cattle subset by identifying non-reference sequences and constructing a pangenome. FindingFive representative genomes of prominent dairy breeds, including Gir, Kankrej, Tharparkar, Sahiwal, and Red Sindhi, were sequenced using 10X Genomics Linked-Read technology. Assemblies generated from these linked-reads ranged from 2.70 Gb to 2.77 Gb,comparable to the Bos indicus Brahman reference genome. A pangenome of Bos indicus cattle was constructed by comparing the newly assembled genomes with the reference using alignment and graph-based methods, revealing 8 Mb and 17.7 Mb of novel sequence respectively. A confident set of 6,844 Non-Reference Unique Insertions (NUIs) spanning 7.57 Mbs was identified through both methods, representing the pangenome of Indian Bos indicus breeds. Comparative analysis with previously published pangenomes unveiled 2.8 Mb (37%) commonality with the Chinese indicine pangenome and only 1% commonality with the Bos taurus pangenome. Among these, 2,312 NUIs - encompassing [~]2 Mb, were commonly found in 98 samples of the 5 breeds and designated as Bos indicus Common Insertions (BICIs) in the population. Furthermore, 926 BICIs were identified within 682 protein-coding genes, 54 long non-coding RNAs (LncRNA), and 18 pseudogenes. These protein-coding genes were enriched for functions such as chemical synaptic transmission, cell junction organization, cell-cell adhesion, and cell morphogenesis. The protein-coding genes were found in various prominent Quantitative Trait Loci (QTL) regions, suggesting potential roles of BICIs in traits related to milk production, reproduction, exterior, health, meat, and carcass. Notably, 63.21% of the bases within the BICIs call set contained interspersed repeats, predominantly LINEs. Additionally,70.28% of BICIs are shared with other domesticated and wild species, highlighting their evolutionary significance. ConclusionThis is the first report unveiling a robust set of NUIs defining the pangenome of Bos indicus breeds of India. The analyses contribute valuable insights into the genomic landscape of desi cattle breeds.