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United States Department of Agriculture

Agricultural Research Service

Title: Characterization of Boivne Micrornas Expressed in Immune and Gut Tissues by Deep Sequencing

Authors
item Matukumalli, Lakshmi - GEORGE MASON UNIVERSITY
item Coutinho, Luiz - UNIVERSITY OF SAO PAULO
item SONSTEGARD, TAD
item LIU, GE
item Gasbarre, Louis
item SMITH, TIMOTHY

Submitted to: BARC Poster Day
Publication Type: Abstract Only
Publication Acceptance Date: March 8, 2006
Publication Date: April 16, 2006
Citation: Matukumalli, L.K., Coutinho, L.L., Sonstegard, T.S., Liu, G., Gasbarre, L.C., Smith, T.P. 2006. Characterization of boivne micrornas expressed in immune and gut tissues by deep sequencing [abstract] 17th Annual BARC Poster Day. Beltsville, MD. No 27.

Technical Abstract: Genetic research in cattle is currently more focused on feed optimization and better animal health as these traits have higher potential for yield improvements. In order to facilitate better understanding of the digestive and immune systems, we comprehensively searched and characterized MicroRNAs (miRs) expressed in the tissues involved in these systems. miRs are small RNA transcripts (~22 nt long) that modulate gene expression by interfering with protein synthesis and/or targeting mRNA for degradation. miRs have been shown to be involved in regulating cellular proliferation, differentiation and immune response. Only 332 miRs have been described for humans, while estimates based on prediction algorithms identify approximately 800 microRNA stem loop loci from human genome sequence. Because of the paucity of information on bovine miRs, we initiated characterization of the miR portion of the transcriptome. Small RNA derived from bovine thymus, abomasal and mesenteric lymph nodes, spleen, bone marrow, Peyer’s patches, fundic abomasum, and small intestine was isolated by gel-based size fractionation. Samples were pooled to increase the potential diversity of miRs prior to ligation of 5’ and 3’ RNA adaptors and cDNA synthesis. This cDNA was cloned and sequenced by Massively Parallel Signature Sequencing (MPSS) to generate approximately 1 million sequence tags ranging from 17 to 20 nt in length. A total of 3,001 distinct tags were identified and these tags could be clustered into 1,692 sequences. Subsequent filtering against rRNA, tRNA and snoRNA (Small Nucleolar RNA) sequences and alignment to the bovine genome assembly yielded 1,252 tags potentially corresponding to miRs. Comparison of these 1,252 tags against the database for experimentally validated miRs (miRBase) resulted in the identification of 178 putative bovine miRs with 130 of these having very significant matches to known vertebrate miRs. The tags that matched miRBase were 83% and 68% of the total tag counts, respectively. The remainder of the sequence tags are being analyzed as potential novel miRs. These results indicate a moderate diversity of miR expression in adult cattle tissues and suggest that less intensive sequencing of different tissues will be sufficient for miR discovery and expression profiling. The sequenence data generated in this study will be useful in generating a high throughput platform for elucidating differences in miR expression that affect function of the bovine digestive and immune systems.

Last Modified: 8/19/2014