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Bustamante-Filho, I. C.; Salton, G. D.; Munari, F. M.; Schneider, M. R.; Mattos, R. C.; Laurino, J. P.; Cirne-Lima, E. O. and Jobim, M. I. M. (2014): Recombinant expression and purification of the bovine acidic Seminal Fluid Protein. In: Animal Reproduction, Vol. 11, No. 2: pp. 96-103

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The acidic Seminal Fluid Protein (aSFP),a 12.9 kDa protein is a maker for bovine semen freezability possibly due to its antioxidant activity and effect on sperm mitochondrial function. However, its precise function on sperm preservation during freezing-thaw is poorly understood. The use of recombinant DNA technology allows new approaches on the study of function and structure of proteins, and its production in procaryote systems offers several advantages. The present work describes the recombinant expression of the bovine aSFP and its binding properties. A cDNA library from the bovine seminal vesicle was used as template for amplification of the aSFP coding region. The amplicon was cloned into a pET23a(+) vector and transformed into E. coli BL21 pLysS strain. The recombinant expression was obtained in E coli. One step ion immobilized affinity chromatography was performed, resulting in high yield of purified protein. To determine the bioactivity of the raSFP, the protein was incubated in different concentrations with 107 spermtozoa at 37 degrees C for 5 h. Western blotting and fluorescence microscopy analyses showed the ability of the recombinant aSFP to attach to the spermatozoa. Based on our results, the described method can be used to obtain mg levels of recombinant aSFP.

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