Jessica Karen Wong/Notebook/2007-8-8

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  • Minipreped 2 samples of I2055-SX-B0030 for seq
    • Sent both samples (1 was fluor and 1 wasn't) for seq w/ VF & VR
  • PCR cleaned I2056 digests (E/X, S/X, both N)
  • Ligated I2056-ENX closed, I2056-SNX closed, I2056-EX-B34, I20550EX-B34
  • Ligated in the presence of Xba and Spe restriction enzymes: I2055-SX-B32, I2056-SX-B32, I2056-SX-B34, I2055-SX-B34
    • Transformed w/ top10
    • Ran out of B0032 cut E/S so made an overnight of B0032 to later cut and ligate
  • Digested E0240 S/N to go into F2620, prep I2055 S/X, prep I2055 E/X


PCR Digest Gel
PCR Digest Gel


  • Ran a gel of the PCR product of each construct next to the Digest of that PCR to make sure digests were cutting in the right place
    • Loaded: lad sp I2057-EX, EX Digest, I2057-ES, ES digest, I2056-EX, EX digest, I2056-SX, SX digest, I2055-EX, I2055 EX digest, sp, I2055 SX digest
    • Didn't have any more PCR product of I2055-SX



Seq

  • I2055-EX seems to have a 1bp mutation at 892 - bad
  • I2055-SX has a few bp differences, breaks down towards 806 even w/ VR - bad
  • I2057-EX-J116 is actually J102, I2057 part is fine after bp 13
    • I2057 first 12 bp's have strange double peaks in chromatogram, large peaks are wrong, small are right
  • I2056-7, I2057-ENS, I2057-ENX all failed again

Made overnights of I2055-EX-B0032, I2056 colony 7, I2057-ENS, I2057-ENX, I2055-ENX-2, I2055-SNX-6, I2055-SNX-7 to sequence

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