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Team:Mississippi State/Isolated Vector
From 2008.igem.org
(Difference between revisions)
| Line 4: | Line 4: | ||
# Add 250ul p2 buffer, mix thoroughly by INVERTING 4-6 TIMES!!, don't vortex. | # Add 250ul p2 buffer, mix thoroughly by INVERTING 4-6 TIMES!!, don't vortex. | ||
# Immediately add 350ul N3 buffer, mix immediately by inverting 4-6 times, mix thoroughly. | # Immediately add 350ul N3 buffer, mix immediately by inverting 4-6 times, mix thoroughly. | ||
| - | # Centrifuge 10min @ 13000rpm (17900g | + | # Centrifuge 10min @ 13000rpm (17900g.) |
# Get 3 spin columns from kit. | # Get 3 spin columns from kit. | ||
# Put supernatant into spin column via pipette. | # Put supernatant into spin column via pipette. | ||
| Line 12: | Line 12: | ||
# Discard flow-through, centrifuge additional 1min to remove residual wash buffer. | # Discard flow-through, centrifuge additional 1min to remove residual wash buffer. | ||
# place QIAprep column in clean 1.5ml centrifuge tube. To elate DNA, add 50ul water to the center of each QIAprep spin column, let stand for 5-10min, centrifuge 1min. | # place QIAprep column in clean 1.5ml centrifuge tube. To elate DNA, add 50ul water to the center of each QIAprep spin column, let stand for 5-10min, centrifuge 1min. | ||
| - | # [[Team:Mississippi State/Running a Gel|Run Gel]] | + | # [[Team:Mississippi State/Running a Gel|Run Gel]]. |
Latest revision as of 16:10, 23 June 2008
- After centrifuge, empty supernatant into flask.
- Resuspend pellet in 250ul pl buffer.
- Vortex until cell pellet dissolves(looks milky.)
- Add 250ul p2 buffer, mix thoroughly by INVERTING 4-6 TIMES!!, don't vortex.
- Immediately add 350ul N3 buffer, mix immediately by inverting 4-6 times, mix thoroughly.
- Centrifuge 10min @ 13000rpm (17900g.)
- Get 3 spin columns from kit.
- Put supernatant into spin column via pipette.
- Centrifuge 30-60s, discard flow-through.
- Wash spin column, add 0.5ml PB buffer and centrifuge 30-60s, discard flow-through.
- Wash spin column, add .75ml PE buffer, centrifuge 60s.
- Discard flow-through, centrifuge additional 1min to remove residual wash buffer.
- place QIAprep column in clean 1.5ml centrifuge tube. To elate DNA, add 50ul water to the center of each QIAprep spin column, let stand for 5-10min, centrifuge 1min.
- Run Gel.
