SBB13Ntbk - Mar12: Difference between revisions
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(New page: ==Regular Zymo Cleanup== #Add 180 uL of Zymo ADB buffer (brown bottle) to a 33uL or 50uL reaction. #Transfer into the Zymo column (small clear guys) #spin through (top speed, 30 sec), dis...) |
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==Regular Zymo Cleanup== | ==Regular Zymo Cleanup== | ||
#Add 180 uL of Zymo ADB buffer (brown bottle) to a 33uL or 50uL reaction. | #Add 180 uL of Zymo ADB buffer (brown bottle) to a 33uL or 50uL reaction. | ||
#Transfer into the Zymo column (small clear guys) | #Transfer into the Zymo column (small clear guys) | ||
| Line 10: | Line 9: | ||
#spin for 90 seconds, full speed to dry. | #spin for 90 seconds, full speed to dry. | ||
#elute with water into a fresh Eppendorf tube, use the same volume of water as the volume of the original reaction | #elute with water into a fresh Eppendorf tube, use the same volume of water as the volume of the original reaction | ||
===Amplification=== | |||
# 1 ul each outer oligo (10 uM) | |||
# 1 ul purified pca product from March 8 | |||
# .5 ul phusion | |||
# 10 ul 5x phusion buffer | |||
# 5 ul 2mM dNTPs | |||
# 32.5 ul H2O | |||
Latest revision as of 23:58, 20 March 2013
Regular Zymo Cleanup
- Add 180 uL of Zymo ADB buffer (brown bottle) to a 33uL or 50uL reaction.
- Transfer into the Zymo column (small clear guys)
- spin through (top speed, 30 sec), discard waste.
- Add 200 uL of Zymo Wash Buffer (which is basically 70% ethanol)
- spin through, discard waste.
- Add 200 uL of Zymo Wash Buffer
- spin through, discard waste.
- spin for 90 seconds, full speed to dry.
- elute with water into a fresh Eppendorf tube, use the same volume of water as the volume of the original reaction
Amplification
- 1 ul each outer oligo (10 uM)
- 1 ul purified pca product from March 8
- .5 ul phusion
- 10 ul 5x phusion buffer
- 5 ul 2mM dNTPs
- 32.5 ul H2O
