Literature DB >> 10631686

Troubleshooting in gene splicing by overlap extension: a step-wise method.

F J Mergulhão1, A G Kelly, G A Monteiro, M A Taipa, J M Cabral.   

Abstract

A step-wise method for cloning intron-containing genes from genomic DNA is described. The two exons of the human proinsulin gene were separately amplified in two steps using, in the first step, completely homologous primers. This reduces unwanted interactions between mismatched primers and a complex DNA template such as genomic DNA. The fragments were amplified in a second step polymerase chain reaction (PCR) using mismatched primers that incorporated additional bases complementary to the other exon, and these products were spliced together in a third step PCR.

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Year:  1999        PMID： 10631686     DOI： 10.1385/MB:12:3:285

Source DB:  PubMed          Journal:  Mol Biotechnol        ISSN： 1073-6085            Impact factor:   2.695


  5 in total

1.  Recombination and amplification of multiple portions of genomic DNA by a modified polymerase chain reaction.

Authors:  K Uchida
Journal:  Anal Biochem       Date:  1992-04       Impact factor: 3.365

2.  Splicing by overlap extension by PCR using asymmetric amplification: an improved technique for the generation of hybrid proteins of immunological interest.

Authors:  A N Warrens; M D Jones; R I Lechler
Journal:  Gene       Date:  1997-02-20       Impact factor: 3.688

3.  Gene splicing by overlap extension: tailor-made genes using the polymerase chain reaction.

Authors:  R M Horton; Z L Cai; S N Ho; L R Pease
Journal:  Biotechniques       Date:  1990-05       Impact factor: 1.993

4.  Engineering hybrid genes without the use of restriction enzymes: gene splicing by overlap extension.

Authors:  R M Horton; H D Hunt; S N Ho; J K Pullen; L R Pease
Journal:  Gene       Date:  1989-04-15       Impact factor: 3.688

5.  Nucleotide sequence of a cDNA clone encoding human preproinsulin.

Authors:  G I Bell; W F Swain; R Pictet; B Cordell; H M Goodman; W J Rutter
Journal:  Nature       Date:  1979-11-29       Impact factor: 49.962
  5 in total
  2 in total

1.  A quantitative ELISA for monitoring the secretion of ZZ-fusion proteins using SpA domain as immunodetection reporter system.

Authors:  F J Mergulhão; G A Monteiro; J M Cabral; M A Taipa
Journal:  Mol Biotechnol       Date:  2001-11       Impact factor: 2.695

2.  The acidic domain of cytochrome c₁ in paracoccus denitrificans, analogous to the acidic subunits in eukaryotic bc₁ complexes, is not involved in the electron transfer reaction to its native substrate cytochrome c(552).

Authors:  Michela Castellani; Jeffrey Havens; Thomas Kleinschroth; Francis Millett; Bill Durham; Francesco Malatesta; Bernd Ludwig
Journal:  Biochim Biophys Acta       Date:  2011-08-11
  2 in total

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