precise genpome editing

Your colleagues submitted interesting questions during the webinar, “TAL Effectors Tool Box for Precision Genome Editing,” so we’ve collected and answered them here.

Find the answer quickly by clicking your desired question.

Q- How specific is TAL? Off-site targeting?

Q- Are TAL-meditated KO or KI strains considered to be GMO?

Q- How big is TAL? Or what’s the size of TALs?

Q- Can TALs be delivered by retroviruses?

Q- Can TALs recognize degenerate binding sites?

Q- Can or how Life Technologies guarantee that TALs will work in the first place?

Q- How fast can I get TALs made so that I can do my KO or KI experiments?

Q- What is the best delivery vehicle for TALs, transfection, electroporation or retroviruses?

 

Q-How specific is TAL? Off-site targeting?

A- Recent paper by Prashant Mali in Nature Biotechnology shows that TALs are tolerant to 1-2 mismatches and less to large majority of 3 bp mismatches.

Q- Are TAL-meditated KO or KI strains considered to be GMO?

A- KO and KI involves editing the native genetic code by either mutating or deleting a encoded message or inserting a new piece of information at a desired site. Although this does manipulate the native genetic information, this technology when used in a responsible manner has very useful applications like engineering yeasts for insulin production or engineering cells for more economiocally and clinically valuable products.

Q- How big is TAL? Or what’s the size of TALs?

A- 3.3 Kb TALDNA

 

Q- Can TALs be delivered by retroviruses?

A- If viral based delivery is the option then Adenoviral systems are better than lento.


Q- Can TALs recognize degenerate binding sites?

A- By careful designing they can be engineered to be very specific. Note: recent publications show that 1-3 base pair mismatch in target DNA sequence can be tolerated to a large extent.

 

Q- Can or how Life Technologies guarantee that TALs will work in the first place?

A-Careful design followed by in vitro validation as well as cleavage analysis to validate the designed TALs.

 

Q- How fast can I get TALs made so that I can do my KO or KI experiments?

A- Manufacturing typically 2 weeks after order has been placed.

Q- What is the best delivery vehicle for TALs, transfection, electroporation or retroviruses?

A- mRNA or DNA with lipid for standard test cell lines ex., 293 hela ets, mRNA delivery also circumvents the threat of transgene integration. Where as for for stem cell electroporation is comparatively a better option.