Jessica Karen Wong/Notebook/2007-8-9

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  • All overnight plates (I2056's and I2055's from yesterday's ligations) have a good amount of growth, very little fluorescence though
    • However, last time they got a lot more red after being in the fridge for a night
    • Will check on them again tomorrow and then make overnights of bright colonies
  • Minipreped overnight cultures (B0032 w/ EB, rest w/ 8.5 water)
  • Sent for sequencing, each with VF and VR
    • I2055-ENX colony 2
    • I2055-SNX (colonies 6 and 7)
    • I2055-EX-B0032
    • I2057 sealed EX and ES to redo
    • I2056 (Scar) colony 7 to redo
I2056 Colony PCR Gel


  • Colony PCRed sealed I2056 8 of each ENX and SNX
    • Ran on gel loaded I2056-ENX (1-8) sp lad sp I2056-SNX (1-8)
    • Top row was PCR's w/ too dilute primers, bottom was normal (exact same cell samples loaded same order)
  • Made overnights of I2056-SX colonies 1,5,6,8 and I2056-EX colonies 3,5,7,8


  • Digested B0032 E/S (b/c had run out of DNA)
  • PCR cleaned I2055 prep S/X and E/X digests and E0240 S/N digest
  • Realized the reason we didn't get many F2620 colonies was b/c we didn't ligate into DB3.1
    • Retransformed all 3 ligations of F2620 into DB3.1 and plated


Seq

  • E0240-ES - E2 and M1 still have mutation, M2 failed - not sure
  • E0240-EX - E2 was perfect, has correct seq!
    • Other sample had an insertion
  • F2620 - was just p1010 mutated
    • Have already set up correct transformations to fix
  • I2055-SX-B31 was perfect, complete coverage!
  • I2057-ES-R0040 - sample 1 was perfect
    • Other 2 samples were wrong
  • I2057-EX-J100 - bad read, mixed signal for both
    • Probably contains 2 different samples
  • I2057-EX-J116 - both samples match J102 instead of J116
    • Sample 1 matches I2057 only after 15, is mixed before that probably contains 2 different strains

I2057-EX-R0040 - both samples contain all of r0040 and all of i2057 and have the right spacing