Cloning of integral mature peptide gene of human GDF-5

Abstract

The integral mature peptide gene of human growth differentiation factor-5 (GDF-5) was cloned to provide the essential foundation for study on the biological characteristics of GDF-5 at gene and protein levels. Two primers were chemosynthesized according to the hGDF-5 sequence reported in Genbank. The hDGF-5 gene was gained by TR-PCR methods from the total RNA extracted from human fetus cartilage tissue, and was cloned into vector pMD18-T. The sequence of recombinant plasmid pMD18-T-hGDF-5 was analyzed by sequence analysis. DNA agarose gel electrophoresis showed that the product of TR-PCR was about 380bp, and double enzyme digestion of the recombinant plasmid corresponded with it. The result of sequence assay was in agreement with the reported hGDF-5 sequence in Genbank. Our results showed that the integral mature peptide gene of human GDF-5 was cloned successfully from human fetal cartilage tissue, and totally identified with the sequence of human GDF-5 in Genbank.

Keywords

growth differentiation factor-5 / cloning

References

[1] HottenG, NeidhardtH, JacobowskyB, et al. . Cloning and expression of recombinant human growth/differentiation factor 5. Biochem Biophys Res Commun, 1994, 204(2): 646-646

[2] YangL, LiH Y, GuoS Z, et al. . Transforming growth factor-β-1 on expression of SMAD3 and SMAD7 in keloid-derived fibroblasts. Zhongguo Linchuang Kangfu, 2003, 7(14): 2004-2004

[3] BuxtonP, EdwardsC, ArcherC W, et al. . Growth/differentiation factor-5 (GDF-5) and skeletal development. J Bone Joint Surg Am, 2001, 83-A(Suppl 1): 23-23

[4] SambrookJ, RussellD W, SambrookJ. Molecular cloning: Alahoratory Manual, 2002, New York, Cold Spring Harbor Laboratory Press, Cold Spring Harbor

[5] ColemanC M, TuanR S. Functional role of growth/differentiation factor 5 in chondrogenesis of limb mesenchymal cells. Mech Dev, 2003, 120(7): 823-823

[6] SettleS H, RountreeR B. Multiple joint and skeletal patterning defects caused by single and double mutations in the mouse Gdf6 and Gdf5 genes. Dev Biol, 2003, 254(1): 116-116

[7] SimankH G, ManggoldJ. Bone morphogenetic protein-2 and growth and differentiation factor-5 enhance the healing of necrotic bone in a sheep model. Growth Factors, 2001, 19(4): 247-247

[8] SpiroR C, LiuL, HeidaranM A, et al. . Inductive activity of recombinant human growth and differentiation factor-5. Biochem Soc Trans, 2000, 28(4): 362-362