动物造模 z-bio 2022-06-07 240

　　Objective: To distinguish and identify multiple alternative splices of APP gene mRNA in tree shrew, describe the characteristics of APP gene, and determine its expression distribution in various tissues.

　　Methods: Referring to the known human and predicted APP gene sequences from tree shrew genomes, common-specific primers for the spliced exons of tree shrew APP gene mRNA were designed. Total RNA was extracted from different tissues of tree shrew, reverse transcribed into cDNA, and the target spliced DNA was amplified by high-fidelity enzyme. The type of spliceosome was preliminarily determined according to the presence and size of the electrophoretic bands of the PCR amplification product. Finally, the PCR product was recovered by gel and sequenced for identification. distribution in different organizations.

　　RESULTS: The results showed that the full length of the tree shrew APP spliceosome was 3514 bp, with a 5'-UTR of 109 bp and a 3'-UTP of 1092 bp. The APP gene was highly homologous and conserved among the nine species investigated, indicating that tree shrew and primates have a relatively close relationship. Four domains shared by tree shrew and human APP gene were obtained by 3D modeling. At the same time, the distribution and expression of these four APP alternative spliceosomes generated by exon skipping in different tissues were confirmed. The four detected spliceosomes, APP770, APP695, APP751 and APP677, were all expressed in lung, kidney and intestine, with the highest expression in lung, muscle and testis.

　　Conclusion: The expression study of the alternative spliceosome of the APP gene in tree shrew will help promote the in-depth study of the disease mechanism and drug development of tree shrew as an Alzheimer's disease model.