Preparation of DNA for Microinjection

Preparation of DNA for Microinjection
1. Isolate the DNA fragment to be microinjected by digesting in
appropriate restriction enzyme and isolating the insert by
electrophoresis on an agarose gel. Start with 50ug or more
2. Excise the band from the agarose gel in a minimum volume of
agarose using a clean, sharp scalpel.
3. Follow QIAEX II Agarose Gel Extraction Protocol except,
injection buffer must be used for elution step: Elute DNA by
adding 20ul of Injection Buffer (TAC prepared). Centrifuge for
30 sec. Carefully pipette the supernatant into a clean tube. This
supernatant contains the DNA.
4. Determine the concentration. Bring the DNA and gel picture to
TAC at 408. TAC staff will do the dilution for injection.
Note: All buffers must be prepared with high quality water such as
Milli-Q, etc. All solutions added to the DNA sample should be
prepared in laminar flow hood
QIAEX II Spin Column, QIAGEN; Catalog Number: 20051