geoscience.net logo
+ Resolve Article
+ Follow Us
Follow on FacebookFollow on Facebook
Follow on TwitterFollow on Twitter

+ Translate
+ Subscribe to Site Feed
GeoScience Most Shared ContentMost Shared Content

FokI method of gene synthesis


, : FokI method of gene synthesis. Gene 68(1): 101-107

An accurate, fast and simple method is presented for synthesis of a gene, or any DNA fragment with a defined sequence. The method is based on the observation that large (approx. 100 bp long) inserts can be cloned into a plasmid using a technique of oligodeoxynucleotide (oligo)-directed double-strand (ds) break repair. The procedure involves transformation of Escherichia coli with a denatured mixture of an insert-carrying oligo and linearized plasmid DNA [Mandecki, Proc. Natl. Acad. Sci. USA 83 (1986) 7177-7181]. The nucleotide (nt) sequences are inserted between two FokI restriction nuclease sites in one of four pUC-derived plasmids. Since FokI makes a staggered ds break at a DNA site 9 and 13 nt away from its recognition site, upon cleavage of the plasmid DNA with FokI, a restriction fragment is liberated that by design contains unique 4-nt-long 5'-protruding ends. The uniqueness of ends permits efficient and directed simultaneous ligation of several restriction fragments to form a gene. The method offers flexibility due to the modular-type assembly and does not require any restriction sites within the constructed gene. The sequence error rate is low: about one error per 4000 bp of DNA cloned. Synthetic DNA for only one DNA strand needs to be provided. The method was applied to the synthesis of a gene fragment encoding the N-terminal 143 amino acid residues of the human immunodeficiency virus transmembrane protein (p41).

US$29.90

PMID: 3265397


Other references

Khidekel, N.; Hsieh-Wilson, L.C., 2004: A 'molecular switchboard'--covalent modifications to proteins and their impact on transcription. Proteins undergo a remarkable variety of posttranslational modifications, with more than 200 distinct modifications identified to date. Increasing evidence suggests that many proteins bear multiple, distinct modifications, and the ability of one m...

Guadagni, S.; D.B.rnardinis, G.; Pavone, P.; Catarci, M.; Agnifili, A.; Carboni, M., 1992: A rare cause of digestive hemorrhage: an aneurysm of the superior pancreaticoduodenal artery rupturing into the duodenal stump of a Billroth II partial gastrectomy. We report herein a case of an iatrogenic superior pancreaticoduodenal arterial aneurysm which ruptured into the duodenal stump of a Billroth II partial gastrectomy. Superselective angiography was used for the diagnosis, and successful embolization...

Platt H.W., 1992: Response of potato breeding program advanced selections and cultivars to late blight 1990. Wysong, D S (Ed ) Biological And Cultural Tests For Control Of Plant Diseases, Vol 7 Iii+137p Aps Press: St Paul, Minnesota, Usa Paper : 33

Maruyama, K., 1966: Effect of beta-actinin on the particle length of F-actin. Biochimica et Biophysica Acta 126(2): 389-398

de la Sierra, A.; Pintó, X.; Guijarro, C.; Miranda, Jé.López.; Callejo, D.; Cuervo, Jús.; Subirà, R.; Rubio, M., 2016: Prevalence, Treatment, and Control of Hypercholesterolemia in High Cardiovascular Risk Patients: Evidences from a Systematic Literature Review in Spain. Cardiovascular diseases (CVDs) represent a major Public Health burden. High serum cholesterol levels have been linked to major CV risk. The objectives of this study were to review the epidemiology of hypercholesterolemia in high risk CV patients f...

Tayler, J.C.; Rudman, J.E., 1963: The assessment of conformation and fatness in beef carcases. The paper reviews various methods of carcass evaluation available to the research worker, and reports some results of studies carried out at Hurley.

Lin, W.Q.; van Dyke, R.A.; Marsh, H.M.; Trudell, J.R., 1994: Time course of 72-kilodalton heat shock protein induction and appearance of trifluoroacetyl adducts in livers of halothane-exposed rats. Previous studies have shown that exposure of phenobarbital pretreated rats to halothane in 10% 02 causes centrilobular necrosis, induces expression of the 72-kDa heat shock protein (HSP72), and produces several trifluoroacetylated adducts. In the...

Stephanie Widdison; Michael Watson; Jenny Piercy; Chris Howard; Tracey J.C.ffey, 2008: Granulocyte chemotactic properties of M. tuberculosis versus M. bovis-infected bovine alveolar macrophages. Molecular Immunology 45(3): 0-749

Goenadi, D.H., 1990: Effect of parent material and topography on soil fertility. I. The Cisampora-Sumberagung Estate, West Java. The effects of topography and the complex geology of mountainous West Java on soil fertility were studied. Eight pedons examined met the criteria for Latosol in the 1961 Indonesia classification, although some had features indicating their develop...

Pevzner L.A.; Shatsilov V.I.; Erenburg M.S., 1987: Complex study methods of longitudinal, transverse and composite waves in studies of the Earth's crust in Kazakhstan. Pages 137-141 1987