Cloning and sequencing of mature fragment of BMP-4 gene
- VernacularTitle:人骨形态发生蛋白4(2b)成熟区cDNA的克隆和序列分析
- Author:
Xinlu WANG
;
Miao LIU
;
Guangfu YANG
- Publication Type:Journal Article
- Keywords:
Bone morphogenetic proteins;
Cloning,molecular;
Sequence analysis,DNA;
Reverse transcription polymerase chain reaction
- From:
Chinese Journal of Orthopaedics
1996;0(09):-
- CountryChina
- Language:Chinese
-
Abstract:
Objective To analyse the cloning and sequencing of mature fragment of human bone morphogenetic protein-4 gene. Methods The template DNA was extract from the human osteosarcoma cells line U-2OS by the single-step isolation method with isothiocyanic acid guanidine, the cDNA coding for the mature fragment of BMP-4 was amplified by the reverse transcription-polymerase chain reaction(RT-PCR). The mature fragment of BMP-4 was cloned into the vector pUC19, and sequenced by Sanger dideoxy-mediated chain termination method. Results The mature fragment of BMP-4 cDNA was obtained by RT-PCR and identified by sequencing. The computer search was done on Genebank, and the published DNA sequence of BMP-4 from Genebank (D30751) was chosen for a reference. Analysis showed that the homology and similarity of nucleotides and amino acids between cDNA of rhBMP-4 mature fragment and the published sequence of BMP-4 were both 99％ . Sequence analysis revealed that there were two bases mutations, one was at base 1 154(201) G C, this had no influence on the corresponding amino acids(Val), another was at base 1 222(269) C T, the mutation at the base 1 222 had turned the Ala into Val. Conclusion The mature fragment of BMP-4 gene has been cloned. The gene is of great significance in treatment of skeletal injuries and diseases.
