title: [Bufo gargarizans mcl-1 cloning and its prokaryotic recombinant protein expression].
abstract: MCL-1 is encoded by myeloid cell leukemia-1 gene (mcl-1), which is one of the anti-apoptotic members of bcl-2 cell apoptotic gene superfamily. ChanSu is made of dorsal secretions of several Bufo species, commonly used in the prescriptions of traditional Chinese medicine for treating many diseases including cancer. To clarify if mcl-1 is expressed in the dorsal skin of B. gargarizans, the PCR (polymerase chain reaction) was performed with its dorsal skin first strand cDNA as the template and a pair of specific primers of mcl-1, and PCR products were cloned into the pGM-T vector. DNA sequencing indicated that the ORF length was 639 bp encoding 212 amino acid residues, and the homology of 44%-95% with the MCL-1 of several other animals. For the further studies on MCL-1 biological functions during the oncogenesis and preparation of its antibody, the prokaryotic expression construct of pET-28b-mcl-1 was prepared which was confirmed by DNA sequencing, and its recombinant protein expression (0.02% wet weight) in E. coli BL21 (DE3) strain was confirmed by SDS-PAGE and Western blotting.
authors.affiliation: College of Animal Science and Technology, Zhejiang A & F University, Lin 'an 311300, China.
authors.forename: Qiao-Ling
authors.initials: QL
authors.lastname: Hu
authors.affiliation: The Nurturing Station for the State Key Laboratory of Subtropical Silviculture, Zhejiang A & F University, Lin 'an 311300, China.
authors.forename: Shu-Fang
authors.initials: SF
authors.lastname: Zhang
authors.affiliation: College of Animal Science and Technology, Zhejiang A & F University, Lin 'an 311300, China.
authors.forename: Xian-Yu
authors.initials: XY
authors.lastname: Yang
authors.affiliation: College of Animal Science and Technology, Zhejiang A & F University, Lin 'an 311300, China.
authors.forename: Mei-Hua
authors.initials: MH
authors.lastname: Yu
authors.affiliation: The Nurturing Station for the State Key Laboratory of Subtropical Silviculture, Zhejiang A & F University, Lin 'an 311300, China.
authors.forename: Hui
authors.initials: H
authors.lastname: Zhu-Ge
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D002025:Bufonidae / Q000145:classification / Q000235:genetics*; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics; D004926:Escherichia coli / Q000378:metabolism*; D064549:Myeloid Cell Leukemia Sequence 1 Protein / Q000235:genetics* / Q000378:metabolism; D016366:Open Reading Frames / Q000235:genetics; D010802:Phylogeny; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism*; D016415:Sequence Alignment; D017385:Sequence Homology; D012867:Skin / Q000378:metabolism
chemical_list: D018076:DNA, Complementary; D064549:Myeloid Cell Leukemia Sequence 1 Protein; D011994:Recombinant Proteins