...these include, TA cloning, ligation independent cloning, TOPO cloning, one step cloning, and overlap extension...Standard Restriction Cloning and Fast Cloning Requirement Restriction Cloning Fast Cloning Choose Restriction...restriction cloning is still routinely performed by many labs, a variety of other cloning techniques with...primer used to amplify the vector is located in the 3’ end of the multi-cloning site, and the reverse primer...your vector and insert. Once you’ve confirmed that the PCR was successful, mix together the vector and ... PCR with vector specific primers to confirm that the products are the correct size (vector + insert)....Transformation + + The FastCloning process in brief In FastCloning, you PCR both the vector and the insert in...

...Expression Vectors Find Bacterial Two Hybrid Plasmids Read our Blog Post About Yeast Vectors Read our Blog...activation (TA) domains retain their individual activities when separated (2), and that DBD and TAs from different...yeast strain that also contains a reporter gene cloned downstream of a promoter containing the target ... is to use a number of different bait and prey vectors. This has been shown to be as effective as using...encoding proteins of interest(POI’s) are fused to TAs and DBDs (see Figure 2a) (4). These fusions are then...if the two POI’s interact, this will bring the POI-TA fusion (commonly referred to as the “prey”) in close...

... TOPO-TA) cloning; however, the TOPO cloning technique has also be adapted for blunt end cloning. So how...Topoisomerase based cloning (TOPO cloning) is a DNA cloning method that does not use restriction enzymes... commercially available TOPO kits provide vectors or cloning arms with overhanging 3´ deoxythymidine (...positive clones. Once you introduce your 3’-end “A” overhang insert, the magic of TOPO cloning happens....protocol.  2. Set Up TOPO Cloning Reaction: Mix together the PCR product and TOPO Vector.  3. Incubate 5 Minutes...Topoisomerase I is released from the vector when the PCR product and vector ligate; however, it can potentially... how does TOPO cloning work? As illustrated in the Figure below, the “A” overhang on the blue PCR product...

...modular vector assembly process designed for a quick, plug-and-play approach to cloning CRISPR vectors (McGee...for each module, generate the vector file, and quickly clone their vector of interest. And it truly is ...working, or spend a few weeks designing and cloning the vectors you actually want. But now you can use the..., were able to use Fragmid to design and clone their vectors in five days. Did you say… easier? Fragmid...destination vectors for various delivery methods, along with compatible inserts. All destination vectors contain...simply assemble them into a destination vector via Golden Gate cloning, perform an exonuclease cleanup, transform...heavy lifting out of CRISPR vector design, making it easier to get the vectors that are just right for your...

...other high throughput cloning strategies There are several high-throughput cloning strategies available...between various cloning methods are listed in Table 2. For example,  chaining-based cloning methods require... used to clone cDNAs from them. These enzymes might not, however, be appropriate for cloning genes from...chaining-based cloning methods are listed in Table 3. The goal of high throughput cloning is to facilitate...High-throughput cloning, in a nutshell, is the systematic combination of different genetic sequences ...sequences into plasmid DNA. In high throughput cloning techniques, although the specific sequences of the genetic... set of various mammalian promoters), the same cloning procedure can be used to incorporate each element...

...Molecular Biology Protocols Find Empty Vectors for Your Next Cloning Experiment ...as Taq generate A-overhangs (a feature used in TA-cloning). Such non-complementary bases at the 3’-end ...manner. The approach can be used in conventional cloning (to introduce or remove restriction sites), in ...the desired modification. Finally, the positive clones are sequenced to confirm the desired modification... addition to the “A” site already in the parent vector). Also shown are the “B” restriction sites, which...subsequent process of screening for succesfully mutated clones. Forward and reverse primers are designed to be... to sequence), it may be easier to subsequently clone a smaller region (containing the specific modification...

... are cloned into Golden Gate entry vectors via traditional restriction enzyme cloning, TA cloning, or ...recombination vectors. The efficient and flexible cloning process provides improvements to classical cloning methods...homology between the vector and gene primer for SLIC cloning. Amplify the pCoofy vector and the sequence ...for recombination in SLIC cloning to form the desired vector. The pCoofy vectors contain the ccdB cassette... existing transposon and vector systems and creating an all-synthetic vector that included only the elements...constructs? A new cloning system combining the advantages of Golden Gate and Multisite Gateway cloning methods ...that you need. The Golden GATEway cloning kit simplifies the cloning process for complex DNA constructs...