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Things we did today

Wetlab work

Verification of Transformants

EtBr stained 2% agarose gel ran at 95V for 1 hour. Five microliters of PCR reaction were loaded into each well.

Grace

• Colony PCR of transformants and restreaked colonies from yesterday

• Ran on 2.5% agarose gel at 95V for 1 min.

• Restreaked:

• slr1+GFPf colony 5
• pilA+GFPf colony 4

• GFPf likely J33207 due to mix up -- see below

Drylab Work

Sequencing

Grace

• Checked sequencing results returned from CORE Hawaii

• Confirmed:

• B0015
• B0030
• B0034 (one bp off)
• nir
• slr

• slr1 = slr2 huh?

• Other:

• nir+rbs

• No rbs present; only nir

• Huh? We ligated nir INTO rbs

• I14032+rbs

• No plac present; only rbs

• BB-pRL1383a

• E0040 (GFP) went in successfully, not J33207

• WTF? Our plasmid preps were/are mislabeled
• Redo ligation to get blue/white screen?

• GFP+tt (reverse only)

• Low quality read; GFP is not present (segment too small)

• GFPf+tt (reverse only)

• lacZ fragment present; no tt

• Huh? We ligated the part INTO tt

• GFPf+tt and J33207+tt samples switched?

• J33207+tt

• Part did not go in; tt only

Discussion

Quote of the Day

History is the only laboratory we have in which to test the consequences of thought. - Étienne Gilson

[http://manoa.hawaii.edu/ ][http://manoa.hawaii.edu/ovcrge/ ][http://www.ctahr.hawaii.edu ]

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