Samantha Liang Notebook


Revision as of 00:00, 10 June 2007 by Samanthaliang (Talk | contribs)

My Construction Files
My Sequencing Files
My Biobricks

Samanthaliang 20:00, 9 June 2007 (EDT)

  • Miniprepped 209, 210, 211, 212, 213 and sent them out for sequencing

Will try to come in on Sunday too to grow up the 214 and 215 cultures - i moved the plates to the fridge for now.

Samanthaliang 17:58, 8 June 2007 (EDT)

  • Made a ridiculously large batch of DH10B competent cells
  • Miniprepped 207 and 208 clones, clone saved them, and sent them out for sequencing
  • Used those preps to subclone I716214 and I716215 (two versions of each)
  • Grew up cultures of 209, 210, 211, 212, 213

Samanthaliang 17:58, 7 June 2007 (EDT)

  • Chris gave me the wrong template for Cre and I didn't notice until today so I redid those 2 PCRs
  • All the other PCRs looked fine on a gel
  • Cleaned up Digested and inserted all PCR products into Biobrick form (I716210 through I716113)
  • Got sequencing data back and only one of the [rbs][atg] ones came out good, but the [lox][terms]s all came out good - just forget about the [rbs][atg] ones for now. More detail - the ones that failed also had really tiny colonies and were in Bca1101 form with a promoter in front, so it might be translating some nonsense that is toxic to the e.coli - but since I have one of the rbs's already I'm just going to proceed with that
  • Picked colonies from pBca9145-I716207-1 and pBca9145-I716208-1
  • Austin grew up 10ml of DH10B for me so I can make competent cells tomorrow

Samanthaliang 17:31, 6 June 2007 (EDT)

  • miniprepped 201 through 206 and sent them out for sequencing
  • put the clones on "clone saver" paper
  • subcloned 207 and 208 with all clones (have 2 versions of each just in case)
  • PCRed ceaB, BamHI, Barnase, Cre with both alternate stop codons

Samanthaliang 19:20, 5 June 2007 (EDT)

  • Grew up 2 colonies each of I716201 through I716206
  • Made a lot of construction files and entered things into the registry
  • My oligos were ordered today!
  • Demonstrated liquid media making and registry entering
  • Now will also make an RFP variant as a reporter on how well Cre is working

To do tomorrow:

  • subclone 207 and 208
  • Run PCRs if they come in the afternoon
  • Send things out for sequencing and make -80s
  • Fill out and sent those survey forms

Samanthaliang 15:29, 4 June 2007 (EDT)

  • Oligos to make biobricks of toxins are not yet in - maybe Wednesday or Thursday?

For cassette between Cre and toxin: [rbs][ATG][Lox][Term][Lox]

  • Made [rbs][ATG] variants with different ribosome binding sites
       I716201 is Bca1106A.Bca1112 
I716202 is Bca1106B.Bca1112
I716203 is Bca1128A.Bca1112
I716204 is Bca1090.Bca1112
  • Made [Lox][Term] variants with 2 different terminators (will probably use Bca1092 version)
     I716205 is Bca1114.Bca1124 
I716206 is Bca1114.Bca1092
  • Tomorrow will make [Lox][Term][Lox] part
  • Then will put the rbsATG together with LoxTermLox

eek so many construction files and ape files to make.

to do