Total DNA synthesis and cloning in Escherichia coli of a gene coding for the human growth hormone releasing factor.
A DNA containing a sequence coding for the human growth hormone releasing factor (hGRF) has been obtained by enzymatic assembly of chemically synthesized DNA fragments. The synthetic gene consists of a 140 base-pair fragment containing initiation and termination signals for translation and appropriate protruding ends for cloning into a newly constructed plasmid vector (pULB1219). Eleven oligodeoxyribonucleotides, from 14 to 31 bases in length, sharing pairwise stretches of complementary regions of at least 13 bases were prepared by phosphotriester solid-phase synthesis. The DNA sequence was designed to take into account the optimal use of E. coli codons. Oligomers were annealed in one step and assembled by ligation. The DNA fragment of the expected size (140 bp) was recovered and cloned into the pULB1219 vector. The expected sequence was confirmed by DNA sequencing.
D000595:Amino Acid Sequence; D001483:Base Sequence; D003001:Cloning, Molecular; D004262:DNA Restriction Enzymes; D005796:Genes; D005813:Genes, Synthetic; D013007:Growth Hormone-Releasing Hormone / Q000235:genetics*; D006801:Humans; D009838:Oligodeoxyribonucleotides / Q000138:chemical synthesis; D010957:Plasmids
D009838:Oligodeoxyribonucleotides; D013007:Growth Hormone-Releasing Hormone; D004262:DNA Restriction Enzymes