Samantha Liang Notebook

From 2007.igem.org

(Difference between revisions)
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*All the other PCRs looked fine on a gel
*All the other PCRs looked fine on a gel
*Cleaned up Digested and inserted all PCR products into Biobrick form (I716210 through I716113)
*Cleaned up Digested and inserted all PCR products into Biobrick form (I716210 through I716113)
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*Got sequencing data back and only one of the [rbs][atg] ones came out good, but the [lox][terms]s all came out good
+
*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
*Picked colonies from pBca9145-I716207-1 and pBca9145-I716208-1

Revision as of 22:08, 7 June 2007

My Construction Files
My Sequencing Files
My Biobricks


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


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