Assembly PCR for targeted random mutagenesis
Expression-independent gene trap vectors for random and targeted mutagenesis in embryonic stem cells
... of random insertions generated with these vectors show a significantly reduced insertional bias and the vectors can be targeted directly to a 5 ’ ...insertional mutagenesis that can be used for ...
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Systematic analysis of the kalimantacin assembly line NRPS module using an adapted targeted mutagenesis approach
... Han et al. 2012; Thirlway et al. 2012). In this study, we attempted to rationally and systematically change the speci- ficity of the glycine- incorporating A domain in the NRPS module of the kalimantacin biosynthetic ...
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CRISPR/Cas9-Targeted Mutagenesis in Caenorhabditis elegans
... by PCR, using a forward primer (5’-AAGAATTCATGCAAGATCCTTTCAAGCA), and reverse primer (5’-AAGAGCTCTGATCCACGATCTGGAAGATT) with EcoRI and SacI restriction sites, ...resulting PCR product was cloned into ...
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PRINCIPLES OF TARGETED MUTAGENESIS IN THE MOSS PHYSCOMITRELLA PATENS.
... at random locations occurs orders of magnitude more frequently than targeted integration at homologous ...of targeted to random integration upon transgenesis correlates with the major pathway ...
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Directed evolution of green fluorescent protein by a new versatile PCR strategy for site-directed and semi-random mutagenesis
... multiple-site mutagenesis, we have substantially modified the Quik-Change™ Site- Directed Mutagenesis Kit protocol (Stratagene, La Jolla, ...a PCR reaction using an in vitro technique, LDA (ligation- ...
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Targeted Mutagenesis of Beta Lactoglobulin Gene in Caprine Fetal Fibroblasts by Context Dependent Assembly Zinc Finger Nucleases
... Abstract Targeted mutagenesis by zinc-finger nucleases (ZFNs) can be used to generate knock-out mamma- lian cell lines with high ...and assembly of gene-specific ZFNs, making them easily accessible ...
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Highly Efficient Targeted Mutagenesis in Mice Using TALENs
... for mutagenesis we enhanced the activity of TALEN mRNAs to optimize nuclease expression upon pronuclear delivery, such that one or more knockin or knockout alleles are obtained among a group of mice derived from a ...
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Insights into CO2 Fixation Pathway of Clostridium autoethanogenum by Targeted Mutagenesis
... Construction of C. autoethanogenum ClosTron strains. Following conjugation of ClosTron retargeting plasmids into C. autoethanogenum by using E. coli strain CA434 as the donor, thiamphenicol- and trimethoprim-resistant ...
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Insights into CO2 fixation pathway of Clostridium autoethanogenumby targeted mutagenesis
... Construction of C. autoethanogenum ClosTron strains. Following conjugation of ClosTron retargeting plasmids into C. autoethanogenum by using E. coli strain CA434 as the donor, thiamphenicol- and trimethoprim-resistant ...
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FASTPCR software for PCR, in silico PCR, and oligonucleotide assembly and analysis
... Oligonucleotide Assembly and Analysis Ruslan Kalendar, David Lee, and Alan ...for PCR primer and probe ...most PCR applications and their combinations, including standard, multiplex, long-distance, ...
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Darwin Assembly: fast, efficient, multi site bespoke mutagenesis
... at 72 ◦ C was routinely included. Annealing temperature and extension time varied according to the primer pair and the size of vector being prepared. Amplification reactions were purified, and treated con- comitantly ...
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Darwin Assembly: fast, efficient, multi-site bespoke mutagenesis
... the targeted codons ...residues targeted. Outnest PCR of the T7 RNA polymerase library (expected product of 2840 bp) confirms that as little as 10 minutes of isothermal assembly were ...
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Efficient targeted mutagenesis in the monarch butterfly using zinc finger nucleases
... zinc-finger assembly assistance; Sriramana Kanginakudru for initial work on this project; Ann Ryan for diet and monarch butterflies; Antonia Monteiro for egg in- jection advice; and Robert Agate, Jerome Menet, ...
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Targeted Chromosomal Cleavage and Mutagenesis in Drosophila Using Zinc-Finger Nucleases
... by PCR. In three cases, the binding site for one of the DSBs in the Drosophila genome. Other investigators have produced hybrid transcription regulators by fusing primers had been deleted, and amplification was ...
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CRISPR/Cas-induced targeted mutagenesis with Agrobacterium mediated protein delivery
... Stability of the octopine Ti plasmid As the previous experiments showed that Agrobacterium can be cured of RP4 we performed an experiment to test if Agrobacterium can be similarly cured of its octopine Ti plasmid. ...
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Marker-fusion PCR for one-step mutagenesis of essential genes in yeast
... Our results show the multiple advantages of this method that let us manipulate essential genes at the endogenous locus in a fast, easy and efficient way. We routinely use this approach to transfer alleles between strains ...
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PTO-QuickStep : a fast and efficient method for cloning random mutagenesis libraries
... conventional PCR that uses phosphorothioate (PTO) oligos to generate megaprimers with 3 ′ ...a random mutagenesis library for directed ...ligation-free random mutagenesis library ...
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Generation of novel random mutagenesis lipase libraries via directed evolution
... Shuffling PCR products of ANL and BTL2 genes, which are amplified with the PCR cycle men- tioned above, are purified using Qiagen PCR Purification ...
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Development of an inducible transposon system for efficient random mutagenesis in Clostridium acetobutylicum
... When colonies were visible on the plates after incubation at 37 ◦ C for 48–72 h, replica cultures of over 200 mutant colonies were inoculated into fresh CBM liquid broth (O’Brien and Morris 1971) in 96-well microtitre ...
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RAPID SOUTHERN BLOT PROTOCOL FOR THE ANALYSIS OF TARGETED MUTAGENESIS IN THE MOUSE IGHG1 LOCUS
... Different probes have been designed to monitor the replacement of the secretory form of the Ighg1 gene. The sequence of the primers used for the PCR amplification of the probes was presented in the table 1. ...
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