Polyclonal antibody preparation and identification of rabbit anti-mouse MORF4L1 isoform a

doi:10.16352/j.issn.1001-6325.2022.05.025

Abstract

Abstract: Objective To prepare rabbit anti-mouse polyclonal antibody against MORF4L1 isoform to differentiate MORF4L1 isoform a and isoform b. Methods According to the antigenicity and autoploidy analysis of amino acid sequence of mouse MORF4L1 isoform a and isoform b,the N-terminal 52-66 amino acids peptide sequence (a 15 peptide,KSAVRPRRSEKSLKT) of mouse MORF4L1 isoform a was synthesized. Then the peptide was linked to a carrier protein KLH (keyhole limpet hemocyanin) as an antigen to prepare the antibody against MORF4L1 isoform a. The antiserum of rabbit 1# with good specificity was purified by antigen affinity chromatography and the antibody was titered by ELISA. In addition, the prokaryotic and eukaryotic expression plasmids of MORF4L1 isoform a and b were constructed. Western blot was used to identify the specificity of the antibody by using the MORF4L1 isoform a and b. Results The antiserum of rabbit 1# recognized polypeptide specifically. The titer of purified antibodywas 1:512 000.pGEX-4T-2-M4S,pGEX-4T-2-M4L prokaryotic expression plasmids and pCMV-3Tag-6-M4S,pCMV-3Tag-6-M4L eukaryotic expression plasmids were successfully constructed and expressed by prokaryotic and eukaryotic expression system. The antibody binds specifically to MORF4L1 isoform a. Conclusions Rabbit anti-mouse MORF4L1 a antibody with high titer and high specificity is successfully prepared, which lay an experimental foundation for further revealing the biological significance of MORF4L1 alternative splicing and the biological functions of different MORF4L1 isoforms.

Key words: MORF4L1, polyclonal antibody, isoform

CLC Number:

R392-33

LI Fang, ZHANG Yuan, ZHANG Ye. Polyclonal antibody preparation and identification of rabbit anti-mouse MORF4L1 isoform a[J]. Basic & Clinical Medicine, 2022, 42(5): 726-731.

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