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[Cloning of full-length Dnmt3b cDNA and its alternative splicing isoforms in mouse embryo]
1National Laboratory of Medical Molecular Biology, Institute of Basic Medical Sciences, CAMS and PUMC, Beijing 100005.
Objective:
To clone the novel full-length cDNA related to mouse embryonic development.
Methods:
Starting from a new expression sequence (clone 2 #) out of a mouse embryonic cDNA library, its full-length cDNA were obtained by rapid amplification of cDNA ends with polyA RNA isolated from mouse 8-9 day embryonic tissue as template, combined with screening of mouse embryonic cDNA library by PCR. Based on the sequences at the two ends of the cDNA, the primers for long range RT-PCR were designed to confirm its factuality.
Results:
The cDNA isolated in our hands was one of the alternative splicing isoforms of mouse de novo DNA cytosine 5'-specific methyltransferase gene (Dnmt3b) reported in July, 1998. A new alternative splicing region upstream of its initiate codon was found 5' forwards 152 bp over the Dnmt3b cDNA sequence. Eight alternative splicing isoforms of Dnmt3b from mouse embryonic tissue were cloned (Genbank accession number are AF151969, AF151970, AF151971, AF151972, AF151973, AF151974, AF151975, AF151976 respectively).
Conclusions:
The 4 kb full-length Dnmt3b cDNA was isolated. Dnmt3b may have novel exons at its 5' upstream and present a complex alternative splicing pattern during mouse embryonic development.