Team:UNIPV-Pavia/Calendar/September/settimana5

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(September, 30th)
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==September, 30th==
==September, 30th==
Transformation of I73 (1ul) into ''E. coli'' TOP10. It was plated on LB+Amp 100 ug/ml agar plate.
Transformation of I73 (1ul) into ''E. coli'' TOP10. It was plated on LB+Amp 100 ug/ml agar plate.
 +
Since I55 was negative again we decided to ligate it again (probably we had a bad digest for vector - I20 - and a good one for insert - we used I38 in other ligations without problems). We diegested already miniprepped DNA  for three hours with 1 ul of enzymes:
Since I55 was negative again we decided to ligate it again (probably we had a bad digest for vector - I20 - and a good one for insert - we used I38 in other ligations without problems). We diegested already miniprepped DNA  for three hours with 1 ul of enzymes:

Revision as of 09:56, 30 September 2010


SEPTEMBER: WEEK 4



September, 27th

MiniPrep for I62-a, I62-b, I62-c, I62-d, I62-e, I62-f

(foto)

I62-e was positive.

DNA pcr for MyYeast 1-2-3

ColonyPCR for I55?? (insert: about 1600bp...)

Protein electrophoresis for phasins (Fede)

September, 28th

Tecan Test Giacomo

September, 29th

Tecan Test Giacomo


Miniprep and quantification for:

  • I1: 59,6 ng/ul
  • I4: 109,9 ng/ul
  • I53: 160,3 ng/ul
  • <partinfo>BBa_J04450</partinfo>-1:55,9 ng/ul
  • <partinfo>BBa_J04450</partinfo>-2: 47 ng/ul

ON digestion of:

  • I1 (E-S)
  • I4 (E-S)
  • I53 (E-X)
  • <partinfo>BBa_J04450</partinfo>-1 (E-P)
  • <partinfo>BBa_J04450</partinfo>-2 (E-P)
  • MyYeast-3 (DNA already available) (E-S)
  • <partinfo>pSB4C5</partinfo> (DNA already available) (E-P)

Gel run for all digested parts and gel run/cut/purification for:

  • 4C5 (E-P): 19,3 ng/ul
  • My Yeast 3 (E-S): 20,8 ng/ul
  • I1: 3,8 ng/ul
  • I4 (E-S): 14,2 ng/ul
  • I53 (E-X): 21,6 ng/ul
  • <partinfo>BBa_J04450</partinfo> (pLac-RBS-mRFP-TT) (E-P): 5,6 ng/ul
  • <partinfo>pSB1C3</partinfo>: 10 ng/ul


Ligation of:

I70=MyYeast 3(E-S)+I4(E-S)
I71=MyYeast 3(E-S)+I1(E-S)
I72=<partinfo>BBa_J04450</partinfo>(E-P)+<partinfo>pSB4C5</partinfo>
I73= I38 (E-S) + I53 (E-X)


September, 30th

Transformation of I73 (1ul) into E. coli TOP10. It was plated on LB+Amp 100 ug/ml agar plate.


Since I55 was negative again we decided to ligate it again (probably we had a bad digest for vector - I20 - and a good one for insert - we used I38 in other ligations without problems). We diegested already miniprepped DNA for three hours with 1 ul of enzymes:

  • I20: (E-X)

Gel run/cut

(foto)

We repeated I55 ligation using newly digested vector and already digested I38 (E-S)

Inoculum into 5ml LB+Amp of:

  • I31
  • I35
  • I37
  • I57
  • I60
  • INTEIN
  • 3x <partinfo>pSB1C3</partinfo>

Cultures were let grow ON at 37°C, 220 rpm.



October, 1st

October, 2nd