Site-directed, Ligase-Independent Mutagenesis (SLIM): a single-tube methodology approaching 100% efficiency in 4 h

Site-directed, Ligase-Independent Mutagenesis (SLIM): a single-tube methodology approaching 100% efficiency in 4 h.

Site-directed, Ligase-Independent Mutagenesis (SLIM) is a novel PCR-mediated mutagenesis approach that can accommodate all three sequence modification types (insertion, deletion and substitution). The method utilizes an inverse PCR amplification of the template by two tailed long primers and two short primers in a single reaction with all steps carried out in one tube. The tailed primers are de...

Ligase chain reaction for site-directed in vitro mutagenesis.

Construction of a recombinant vector for site-directed mutagenesis in Salmonella typhimurium

BACKGROUND: Among all common techniques in sitedirectedmutagenesis, λ Red recombinase system has beenwidely used to knock out chromosomal genes in bacteria. In thismethod, there is always the risk of DNA Linear digestion byhost's restriction enzymes that leads to the low frequency ofrecombination. OBJECTIVES:To overcome this, we constructeda recombinant vector to disrupt phoP gene in Salmonella...

Site-Directed Mutagenesis Study

Structural elements of the rat 1.t-opioid receptor important in ligand receptor binding and selectivity were examined using a site-directed mutagenesis approach. Five single amino acid mutations were made, three that altered conserved residues in the ~ 8, and K receptors (Asn 15°t Ala, His297 to Ala, and Tyr326 to Phe) and wo de igned to test for ~i/8 selectivity (lie198 to Val and Va1202 to li...

Cloning independent site-directed mutagenesis using total RNA as template.

Several site-directed mutagenesis methods have been developed and are being currently used in research. All these methods apply three or four primers, including one or two bearing the appropriate mutation, and most use PCR to amplify the desired sequence and to incorporate the mutation (1–6). Although several developments have improved the robustness and reliability of the method, all of them r...