The Polymerase Incomplete Primer Extension (PIPE) Method Applied to High-Throughput Cloning and Site-Directed Mutagenesis

2009 ◽  
pp. 91-103 ◽  
Author(s):  
Heath E. Klock ◽  
Scott A. Lesley
Keyword(s):  
High Throughput ◽  
Primer Extension ◽  
Site Directed Mutagenesis ◽  
Directed Mutagenesis

A one-tube site-directed mutagenesis method using PCR and primer extension

2008 ◽  
Vol 372 (2) ◽  
pp. 255-257 ◽  
Author(s):  
Nava Chapnik ◽  
Hadas Sherman ◽  
Oren Froy
Keyword(s):  
Primer Extension ◽  
Site Directed Mutagenesis ◽  
Directed Mutagenesis

HTP-OligoDesigner: An Online Primer Design Tool for High-Throughput Gene Cloning and Site-Directed Mutagenesis

2016 ◽  
Vol 23 (1) ◽  
pp. 27-29 ◽  
Author(s):  
Cesar M. Camilo ◽  
Gustavo M.A. Lima ◽  
Fernando V. Maluf ◽  
Rafael V.C. Guido ◽  
Igor Polikarpov
Keyword(s):  
Gene Cloning ◽  
High Throughput ◽  
Primer Design ◽  
Design Tool ◽  
Site Directed Mutagenesis ◽  
Directed Mutagenesis

scholarly journals High-throughput site-directed mutagenesis using oligonucleotides synthesized on DNA chips

BioTechniques ◽  
2005 ◽  
Vol 39 (3) ◽  
pp. 363-368 ◽  
Author(s):  
Didier Saboulard ◽  
Vincent Dugas ◽  
Mehdi Jaber ◽  
Jérôme Broutin ◽  
Eliane Souteyrand ◽  
...  
Keyword(s):  
High Throughput ◽  
Site Directed Mutagenesis ◽  
Directed Mutagenesis ◽  
Dna Chips

scholarly journals High throughput mass spectrometry-based characterisation of Arabidopsis thaliana group H glycosyltransferases

RSC Advances ◽  
2018 ◽  
Vol 8 (53) ◽  
pp. 30080-30086 ◽  
Author(s):  
Aishat Akere ◽  
Qian Liu ◽  
Shibo Wu ◽  
Bingkai Hou ◽  
Min Yang
Keyword(s):  
Mass Spectrometry ◽  
Arabidopsis Thaliana ◽  
Sequence Alignment ◽  
High Throughput ◽  
Site Directed Mutagenesis ◽  
Directed Mutagenesis ◽  
Substrate Specificities

We cloned and characterised four group H glycosyltransferases by studying their substrate specificities and kinetics. Sequence alignment and site-directed mutagenesis studies showed that serine is a crucial residue for UDPGlcNAc and UDPGal activity.

scholarly journals High-throughput antibody engineering in mammalian cells by CRISPR/Cas9-mediated homology-directed mutagenesis

2018 ◽  
Author(s):  
Derek M Mason ◽  
Cédric R Weber ◽  
Cristina Parola ◽  
Simon M Meng ◽  
Victor Greiff ◽  
...  
Keyword(s):  
Directed Evolution ◽  
High Throughput ◽  
High Throughput Screening ◽  
Mammalian Cells ◽  
Affinity Maturation ◽  
Antibody Engineering ◽  
Site Directed Mutagenesis ◽  
Directed Mutagenesis ◽  
Variant Discovery ◽  
Novel Variant

ABSTRACTAntibody engineering is performed to improve therapeutic properties by directed evolution, usually by high-throughput screening of phage or yeast display libraries. Engineering antibodies in mammalian cells offers advantages associated with expression in their final therapeutic format (full-length glycosylated IgG), however, the inability to express large and diverse libraries severely limits their potential throughput. To address this limitation, we have developed homology-directed mutagenesis (HDM), a novel method which extends the concept of CRISPR/Cas9-mediated homology-directed repair (HDR). HDM leverages oligonucleotides with degenerate codons to generate site-directed mutagenesis libraries in mammalian cells. By improving HDM efficiency (>35-fold) and combining mammalian display screening with next-generation sequencing (NGS), we validated this approach can be used for key applications in antibody engineering at high-throughput: rational library construction, novel variant discovery, affinity maturation, and deep mutational scanning (DMS). We anticipate that HDM will be a valuable tool for engineering and optimizing antibodies in mammalian cells, and eventually enable directed evolution of other complex proteins and cellular therapeutics.

High-Throughput Site-Directed Mutagenesis in ES Cells

2001 ◽  
Vol 289 (2) ◽  
pp. 329-336 ◽  
Author(s):  
Alexandra Höllrigl ◽  
Alexander Hergovich ◽  
Irene Görzer ◽  
Alice Bader ◽  
Günther Ellersdorfer ◽  
...  
Keyword(s):  
High Throughput ◽  
Es Cells ◽  
Site Directed Mutagenesis ◽  
Directed Mutagenesis
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