By A Max Juchheim
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When facing a cloning project, scientists are no longer limited to traditional restriction enzyme cloning. Instead, you can choose a molecular cloning technique that will work well with a given set of resources, time, and experimental needs. Since its invention in the late ...
If you’re into cloning, you’re probably aware that there are several methodologies currently available for approaching it. These include the traditional restriction enzyme/ligase-mediated method, the more recently developed Gibson Assembly Cloning and Gateway® cloning ...
One of the most powerful strategies to investigate a gene's function is to inactivate, or "knockout", the gene by replacing it or disrupting it with an piece of DNA designed in the lab. Specially constructed plasmids can be used to replace genes in yeast, mice, or Drosophila ...
Topoisomerase based cloning (TOPO cloning) is a DNA cloning method that does not use restriction enzymes or ligase, and requires no post-PCR procedures. Sounds easy right? The technique relies on the basic ability of complementary basepairs adenine (A) and thymine (T) to ...
Have you ever tried digesting with XbaI or ClaI restriction enzymes and gotten unusual or unexpected results? Or considered why DpnI will degrade your template DNA from a PCR reaction but not the newly synthesized product from a site-directed mutagenesis experiment? The answer ...
This post was updated on March 21, 2018. Most of the time, plasmid prepping is a breeze. You get your stab from Addgene, streak for single colonies, sub-culture, and prep with a DNA prep kit or your lab's favorite in-house protocol. DNA yields for this procedure are typically in ...
Molecular cloning requires some method of screening colonies for the presence of an insert. Traditionally this has been done with restriction enzyme digest; however colony PCR can accomplish the same thing in less time and for less money. The key steps to colony PCR are: 1) ...
