Amplification of Human Nerve Growth Factor cDNA in Vitro

Objective RNA was extracted from human placenta.Human βNGFcDNA was amplified by using mRNA as template and then nucleotide sequence was deter mined.All of this work is the basis of further study of cloning and expression of βNGFcDNA.Methods RNA was extracted by guanidine isothiocyanate method;human βNGFcDNA was amplified by RT-PCR technology;nucleoside sequence was identified using Sanger double deoxgen end method.Results RNA was extracted from the tissue of human placenta;human βNGFcDNA was amplified,mRNA of RNA served as template.The segment was identified as aimed DNA by agarose electrophoresis,and necleotide sequence accorded to actual sequence.Conclusion Human βNGFcDNA was amplified by RT-PCR technology using mRNA from human placenta as template and cDNA 5' end was modified with cutting-site of restriction endonuclease,which is convenient for the cloning and expression of βNGF gene.