A large fragment approach to DNA synthesis: total synthesis of a gene for the protease inhibitor eglin c from the leech Hirudo medicinalis and its expression in E. coli
Biotechnology article published in Nucleic Acids Research (1984)
Abstract
A DNA containing the coding sequence for the proteinase inhibitor protein, eglin c, from the leech Hirudo medicinalis has been obtained by enzymatic assembly of chemically synthesized DNA fragments. The synthetic gene consists of a 232 base-pair fragment containing initiation and termination codon signals with restriction enzyme recognition sites conveniently placed for cloning into a plasmid vector. Only six oligonucleotides from 34 to 61 bases in length, sharing pairwise stretches of complementary regions at their 3'-termini, were prepared by phosphotriester solid-phase synthesis. The oligomers were annealed pairwise and converted into double stranded DNA fragments by DNA polymerase I mediated repair synthesis. The fragments were assembled by ligation, and the synthetic gene was expressed in high yield in E. coli under the transcriptional control of the E. coli tryptophan promoter. The expression product was purified to homogeneity and was shown to have similar physicochemical and identical biological properties as the authentic protein isolated from the leech.
Abstract sourced from PubMed (NCBI) for the cited record. See the original publication for the authoritative version.
Резюме
Enzymatic assembly of chemically synthesized oligonucleotides created a synthetic gene encoding leech eglin c that was cloned and expressed at high yield in E. coli under the tryptophan promoter.
Почему это важно для гирудотерапии
This paper reports the total chemical synthesis and expression of a gene encoding eglin c—the proteinase inhibitor from Hirudo medicinalis—by assembling six chemically synthesized oligonucleotides into a 232 base-pair fragment with appropriate codon signals, cloning it under the E. coli tryptophan promoter, and expressing it in high yield in E. coli; the purified product matched the authentic leech protein in physicochemical and biological properties. For ASH's domain, this is relevant as a production method for a defined leech-secretome bioactive, potentially enabling scalable supply independent of live leeches. Caveat: this is a molecular-biotechnology/gene-synthesis study; it contains no therapeutic, in-vivo, or hirudotherapy data.
Цитирование
A large fragment approach to DNA synthesis: total synthesis of a gene for the protease inhibitor eglin c from the leech Hirudo medicinalis and its expression in E. coli.
Rink H, Liersch M, Sieber P, Meyer F · Nucleic acids research, 1984
Связанный клинический контекст
Узнайте, как это исследование связано с клинической практикой
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