Features | Partner Sites | Information | LinkXpress
Sign In
GLOBETECH PUBLISHING
GLOBETECH PUBLISHING
JIB

Robust Site-Directed Mutagenesis Kit Enables Standard and Novel Applications

By BiotechDaily International staff writers
Posted on 22 May 2013
A new kit that enables rapid site-directed mutagenesis (SDM) of plasmid DNA has been designed to include unique features that expand the range of applications, including a more effective method for introduction of long insertions and deletions. A new SDM primer design web tool that supports the kit has also been developed.

New England Biolabs (NEB; Ipswich, MA, USA) introduces the “Q5 Site-Directed Mutagenesis Kit,” capable of making standard single-base modifications as well as novel manipulations such as deletions or additions of almost any length, including for the addition of nuclear localization signals to cloned proteins. It also includes unique features for difficult-to-amplify GC-rich and repetitive sequences. "The Q5 SDM Kit makes applications possible for which you might not have thought of using SDM. For example, SDM can be a much quicker and less error-prone way to add tags, make large deletions, add or remove restriction sites, and many other sequence modifications," said John Pezza, PhD, scientist at NEB’s Applications and Product Development.

NEB adopted a nonoverlapping primer design to allow for longer modifications. The most popular SDM methods use overlapping primers that limit the length of insertions or deletions. NEB's approach enables exponential amplification and more efficient plasmid transformation, which also facilitates the successful modification of highly repetitive or GC-rich sequences. NEB has also released “NEBaseChanger,” an SDM primer design web tool that supports the Q5 SDM Kit. Developed by Sanjay Kumar, PhD, Senior Research Scientist at NEB, it includes a novel feature - the application of nearest neighbor thermodynamics for matched and mismatched bases to design mutational SDM primers and calculate custom annealing temperatures. "These adjustments increase the odds that the experiment will work. Most, if not all other design tools, are not able to take mismatches into account," said Dr. Pezza. NEBaseChanger also provides live primer design updates, so users can make iterative changes to their primers and watch how details like reading frame and Tm are affected.

The Q5 SDM Kit includes the robust Q5 Hot Start High-Fidelity Polymerase, which has been tested to amplify sequences with a fidelity 100-times greater than Taq and twice that of “Phusion” DNA Polymerase. The kit also includes high-efficiency NEB 5-alpha Competent E. coli. Robust results have been observed with plasmids up to at least 14 kb.

Related Links:

New England Biolabs
Q5 Site-Directed Mutagenesis Kit



comments powered by Disqus

Channels

Genomics/Proteomics

view channel
Image: Alternative splicing produces two protein isoforms (Photo courtesy of Wikimedia Commons).

Key Regulator of Cancer-Inducing Alternative Splicing Identified

Cancer researchers have identified the splicing factor RBM4 (RNA-binding protein 4) as a key determinant in processes that prevent tumor development and spread. RBM4 is known to be crucial to gene splicing... Read more

Drug Discovery

view channel

Ibuprofen May Restore Immune Function in Older Individuals

New research suggests that macrophages from the lungs of old mice respond differently to infections than those of young mice, and ibuprofen given to the mice reversed these changes. New research using lab mice suggests that the solution to more youthful immune function might already be a common over-the-counter pain reliever.... Read more

Therapeutics

view channel
Image: Hair follicle (blue) being attacked by T cells (green) (Photo courtesy of Christiano Lab/Columbia University Medical Center).

Hair Restoration Method Clones Patients’ Cells to Grow New Hair Follicles

Researchers have developed of a new hair restoration approach that uses a patient’s cells to grow new hair follicles. In addition, the [US] Food and Drugs Administration (FDA) recently approved a new drug... Read more

Business

view channel

Partnership Established to Decode Bowel Disease

23andMe (Mountain View, CA,USA), a personal genetics company, is collaborating with Pfizer, Inc. (New York, NY, USA), in which the companies will seek to enroll 10,000 people with inflammatory bowel disease (IBD) in a research project designed to explore the genetic factors associated with the onset, progression, severity,... Read more
 
Copyright © 2000-2014 Globetech Media. All rights reserved.