Cloning of three newly-identified alternatively splicing isoforms of mouse Pax-8 and their spatial and temporal expression patterns
1.Department of Cardiology, the First Affiliated Hospital of Wenzhou Medical University, Wenzhou, 325035; 2.Department of Cardiology, the Affiliated Hospital of Hangzhou Normal University, Hangzhou, 310015
Abstract：Objective: Pax-8 gene plays a pivotal role in embryonic development and tumorgenesis. This study aims to identify and clone different alternatively splicing isoforms of mouse Pax-8, and to detect their spatial and temporal expression patterns. Methods: Hearts from wild-type prenatal and postnatal mice of C57BL/6 background were carefully excised and total RNA was extracted using Trizol reagent. The cDNA fragment of Pax-8 was reverse transcribed and amplified by Q5 Taq polymerase and cloned into pcDNA3.1(-) vector. The recombinant vectors were validated by enzyme digestion and sequencing. Results: In comparison to their cognate full-length Pax-8a, three alternatively splicing isoforms of mouse Pax-8, designated Pax-8b (exon4 deleted), Pax-8c (exon 4 and 11 deleted) and Pad-8d (exon 4 and 9 deleted), were identified in different periods of embryogenesis and successfully cloned. Conclusion: Mouse heart has four alternatively splicing isoforms of mouse Pax-8 that express in different spatial and temporal expression patterns. Such findings open a bright avenue for the further functional study.
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