| Mastitis of dairy cows is a complex disease, which remains the most costly disease to the worldwide dairy industry, despite the widespread implementation of mastitis control strategies. Genetic differences of mastitis resistance between breeds and within breed in dairy cows are significant, so genetic selection to improve mastitis resistance may be the best long-term strategy bacause it offers the possibility for permanent resolution of this complex disease problem.The aim of the current research was to use the tools of comparative genomics on functional genome study of dairy cows to isolate and clone their disease resistance relational genes, look for molecular events and establish fundamental materials and techniques for further study and genetic selection. The main results obtained are as follows:The experiment used the blood of Hostein cows with mastitis. After extracting RNA from blood leucocytes, we designed primers according to cDNA sequence of bovineβ-defensins in GenBank and amplified the cDNA template of BNBD5 by RT-PCR technology and constructed pMD19-T vector finally. The cloned plasmid named BNBD5/pMD19-T.The amino acid sequences ofβ-defensin in Bos Taurus, Homo sapiens and Mus musculus tended to middle conservation. The positives score was 74.3 and identity score was 20.0. The analysis of conserved domain indicated thatβ-defensin proteins of above three species were members ofβ-defensin superfamily. The three dimensional structure was predicted by SWISS-MODEL (http://swissmodel.expasy.org/workspace/index.php). A structure of threeβ-strands which were arranged in reverse orde was showed by PyMol software. Three disulfide bonds with 6 cysteines fixed the structure. Our data also suggested thatβ-defensin was an antibacterial peptide with microbial colony resistance on the surface in mammary epithelial cells.The amino acid sequences of AHCY in Bos Taurus, Homo sapiens and Mus musculus were significantly high conservative. The positives score was 99.5 and identity score was 95.1. Bovine AHCY protein had 3 conserved domains. The predicted 3D structure included two parts, ecch had a fewβ-strands that were involved inα-helix of two sides.The amino acid sequences of MPIF-1 in Bos Taurus, Homo sapiens and Mus musculus were high conservative. The positives score was 37.8 and identity score was 29.7. Bovine MPIF-1 protein included a large number of functional regions, e.g. dimmer interface (I and P form), tetramer interface, putative receptor binding cleft, putative GAG binding site and N-loop etc. The predicted 3D structure revealed that bovine MPIF-1 was a monomer and adopted the chemokine fold of threeβ-strands and an overlyingα-helix. In addition to the four cysteines that characterize most chemokines, MPIF-1 had two additional cysteines that formed a disulfide bond. A distinct positively charged pocket was also found in this structure.
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