猪CPB2基因可变剪接体的克隆及生物信息学研究

doi:10.11843/j.issn.0366-6964.2022.10.011

Abstract

Abstract: The objective of this study was to clone different alternative splices of pig CPB2 gene, so as to predict the functional properties of the corresponding proteins and study the expression characteristics of different transcripts. Three 12-month-old Bama miniature pigs with the same health status and body weight were used as experimental animal, the CDS region and part of non-coding region of CPB2 gene were amplified by RT-PCR and gene flat terminal cloning technology, while the basic biological characteristics of CPB2 coding protein were predicted by bioinformatics tools. The expression characteristics of CPB2 gene in liver, heart, kidney, subcutaneous fat, mesenteric fat, biceps femoris muscle and longissimus dorsi muscle were detected by RT-PCR. qRT-PCR was used to detect the expression characteristics of CPB2 gene in liver of metabolic disease susceptible pigs prepared in the laboratory under the effect of nutrient-rich diet.Three kinds of alternative splicing isoforms (CPB2-1, CPB2-2 and CPB2-3) of pig CPB2 gene were cloned successfully. CPB2-1 was identical with the NCBI sequence XM_001929144.6. CPB2-2 and CPB2-3 were newly discovered transcripts. Sequence alignment showed that alternative 5' splice site (A5SS) events occurred in CPB2-2 and CPB2-3 compared with CPB2-1. Bioinformatics showed that CPB2-1 encoded an acid unstable protein with 423 amino acids, and CPB2-2 and CPB2-3 encoded an alkaline unstable protein with 281 amino acids; Compared with CPB2-1, CPB2-2 and CPB2-3 lacked signal and activating peptides, but had the same carboxypeptidase activity domain. The three CPB2 transcripts were only expressed in liver and kidney, no expression was found in other tissues. In liver of metabolic disease susceptible pigs induced by nutrient-rich diet, CPB2-1 (P<0.01) and CPB2-2 (P<0.01) significantly reduced, CPB2-3 was not significantly reduced (P=0.14). In conclusion, three kinds of alternative splicing isoforms of pig CPB2 were successfully cloned in liver in this study, suggesting that CPB2-1 is a normal transcript of fibrinolysis and coagulation. Newly discovered transcripts of CPB2-2 and CPB2-3 may have carboxypeptidase activity and important physiological functions when retained in liver and kidney. CPB2 gene may be associated with metabolic related chronic liver disease.

Key words: pig, CPB2, alternative splicing, function prediction, expression characteristics, chronic liver disease

CLC Number:

S828.2

XIA Boce, ZHANG Kaiyi, MIAO Jiakun, YANG Yu, PENG Huanqi, WANG Yanfang, YANG Shulin. Cloning and Bioinformatics Study of Alternative Splicing Isoforms of Pig CPB2 Gene[J]. Acta Veterinaria et Zootechnica Sinica, 2022, 53(10): 3377-3390.

References 33

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Cloning and Bioinformatics Study of Alternative Splicing Isoforms of Pig CPB2 Gene

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