Random mutagenesis libraries: optimization and simplification by PCR.
نویسندگان
چکیده
We thank John Connolly for helpful discussions and critical reading of the manuscript. We are very grateful to Fabrice Chareyre, Laetitia Gressin, Catherine Giudicelli, Jean Christophe Beaudoin and Isabelle Legall for help in the BAC screening, sequencing and genotyping. This work was supported by a grant from the Association pour la Recherche sur le Cancer (ARC) and the Ministère de l’Education et de la Recherche. Address correspondence to Jessica Zucman-Rossi, INSERM U434 CEPH/ Fondation Jean Dausset, 27 Rue Juliette Dodu, 75010 Paris, France. Internet: [email protected]
منابع مشابه
Thermostable DNA Ligase-Mediated PCR Production of Circular Plasmid (PPCP) and Its Application in Directed Evolution via In situ Error-Prone PCR
Polymerase chain reaction (PCR) is a powerful method to produce linear DNA fragments. Here we describe the Tma thermostable DNA ligase-mediated PCR production of circular plasmid (PPCP) and its application in directed evolution via in situ error-prone PCR. In this thermostable DNA ligase-mediated whole-plasmid amplification method, the resultant DNA nick between the 5' end of the PCR primer and...
متن کاملA computer program for the estimation of protein and nucleic acid sequence diversity in random point mutagenesis libraries
A computer program for the generation and analysis of in silico random point mutagenesis libraries is described. The program operates by mutagenizing an input nucleic acid sequence according to mutation parameters specified by the user for each sequence position and type of point mutation. The program can mimic almost any type of random mutagenesis library, including those produced via error-pr...
متن کاملRandom mutagenesis by whole-plasmid PCR amplification.
Random mutagenesis has become a powerful means of studying the effects of a large number of permutations of a particular DNA sequence. As a prime example, libraries of randomized cDNA clones, when translated into their corresponding proteins, can be useful in investigating the functional contributions of a mutagenized region to the overall properties of a protein. Existing molecular cloning tec...
متن کاملCreating random mutagenesis libraries using megaprimer PCR of whole plasmid.
The conventional method for cloning a DNA fragment is to insert it into a vector and ligate it. Although this method is commonly used, it is labor intensive because the ratio and concentrations of the DNA insert and the vector need optimizing. Even then, the resultant library is often plagued with unwanted plasmids that have no inserts or multiple inserts. These species have to be eradicated to...
متن کاملUV mutagenesis for the overproduction of xylanase from Bacillus mojavensis PTCC 1723 and optimization of the production condition
Objective(s):[p1] This study highlights xylanase overproduction from Bacillus mojavensis via UV mutagenesis and optimization of the production process. Materials and Methods:Bacillus mojavenis PTCC 1723 underwent UV radiation. Mutants’ primary screening was based on the enhanced Hollow Zone Diameter/ Colony Diameter Ration (H/C ratios) of the colonies in comparison with the wild strain on Xyla...
متن کاملذخیره در منابع من
با ذخیره ی این منبع در منابع من، دسترسی به آن را برای استفاده های بعدی آسان تر کنید
عنوان ژورنال:
- BioTechniques
دوره 27 6 شماره
صفحات -
تاریخ انتشار 1999