Team:Hawaii/Notebook/2008-07-29

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= Things we did today =
== Wetlab work ==
===Gel purification of ccdB PCR product===
:<strong>Grace</strong>

===RE digests===
:<strong> Grace</strong>

:* Digested pRL1383a (from last night) with BamHI
:* Digested B0030 and B0024 (from last night) with EcoRI
:* Sequentially digested gel purified ccdB with HindIII and BamHI

===Construction of BioBricks===
:<strong>Grace</strong>

:* Purified RE digests on EtBr stained 3% agarose gel
::* Extracted bands for B0030 and B0024
::* Other lanes FAILED!

===Direct Band Extraction Method===
:* make two comb gels
:* run test UPA band along with ladder
:* attempt to pipette out DNA as it runs into the second well
:* no more gel cutting???

===PCR amplification of pRL1383a parts===
:<strong>Margaret</strong>

:* Made a reaction buffer containing {38.5ul, 20ul accusure, 10ul dNTPs, & 5ul 5XBuffer}. This should be good for 20 50ul reactions.

:*Amplified with Accusure: aadA ([http://partsregistry.org/Part:BBa_J23012 BBa_J23012]), aadaA (pRL1383a), rep region, oriV
:*Amplified with Green Taq: rep region

:*On [[Team:Hawaii/Notebook/2008-07-30|7-30]] I ran a gel.

===Media===

:*started to make LB amp100 plates, but autoclave room is closed. Remember to do it tomorrow.



= Discussion =
''' SYBR Safe'''
:* Blue light still doesn't illuminate the bands well. No bands (not even the ladder) are visible. Resolution *much* better with short wave UV. EtBr better bet for now?
'''Restriction enzymes'''
:* HindIII and BamHI in the lab -20C freezer (in box labeled "TKW Restriction Enzymes") fail. They're really old and don't appear to cut well anymore. Digestion of pRL1383a resulted in a long smear.

= Quote of the Day =
LOL! http://www.youtube.com/watch?v=J0s0Y3-BCaw

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