PMC:441373 / 42989-44651 JSONTXT

{"target":"https://pubannotation.org/docs/sourcedb/PMC/sourceid/441373","sourcedb":"PMC","sourceid":"441373","source_url":"http://www.ncbi.nlm.nih.gov/pmc/441373","text":"Cloning of vertebrate TACC cDNAs\nThe rabbit TACC3 was cloned by rt-PCR using the \"TACC4\" specific primer T4RACE2 [9] (5'-cccgaactgctccaggtaatcgatctc-3') and a consensus primer, T3con2, designed to the region encompassing the vertebrate TACC3 initiator methionine (5'-tatgagtctgcaggtcttaaacgac-3'). For cloning the mouse Tacc1X cDNA, the primers used were based upon the genomic sequence reported, and the sequence of the IMAGE cDNA clone 4933429K08: T1XF (5'-ccatgttcagtcattggcaggtc-3'), T1XF2 (5'-ctgcagaaccaacagttcaag-3'), T1XR1 (5'-agatctgtgacatcacagctc-3'), T1XR2 (5'-ctcgagtcagttagtcttatccagctt-3'), BB617F (5'-accaccaacttgagtacctg-3') and BB617R (5'-gtatcttgaactgttggttctg-3'). For analysis of TACC1 splice variants, the forward primers used were located in exon 1b: EF (5'-gagagatgcgaaatcagcg-3'), Exon 1d: X3F (5'-agtcaaagaaggcatctgcag-3'), Exon 1a: 1DF (5'-ccaagttctgcgccatggg-3'). The reverse primers used were: Exon 1: X1R (5'-ggatttggtctcggcttgcgaatc-3'), Exon 2: BX647R (5'-cttgtgattcttggcttttgg-3'), Exon 3: 6SPR (5'-gtcatcgcctcgtcctggagggc-3'), Exon 4a: 1DR (5'-aatttcacttgttcagtagtc-3'), Exon 5: 128R (5'-cctgcttctgaggatgaaaacgc-3'). Rabbit brain poly A+ mRNA, mouse testis and human brain total RNA were obtained from BD Bioscience Clontech (Palo Alto, CA, U.S.A.). Reverse transcription and PCR was performed as previously described, using either 1 μg of total RNA or 50 ng of poly A+ mRNA as template for first strand cDNA synthesis. PCR products were cloned into pCR2.1 (Invitrogen, Carlsbad CA, U.S.A.) and transformed into InvαF' competent cells. 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