Team:Potsdam Bioware/Lab/Labjournal/October

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Contents

AID

2012-10-10

Inoculation of plasmid samples of the 48h retransformation plates (after FACS)

Investigators: Tom


Time: 2012-10-10


Materials:

  • LB medium
  • Amp stock solution
  • plate with cultures:

EGFR-C-AID without NES, with NLS+Kozak sequence+eGFP

(after FACS selection)

Method:

Inoculation of:

5 cultures per plate in 5 ml LB medium + 5µL Amp.

(--> 5 cultures)

Further tasks:

  • Miniprep


Antibody

2012-10-12

cell culture


Investigator:Kerstin

Topic: cell-culture

  • passaging of CHO w Zeocin
  • passaging of HT1080
  • passaging of HEK AAV 293
  • passaging of HeLa


Topic: planning new primer for integration of new RAGE-transmembrane domain and RAGE-signalpeptide

Investigator: Sascha

Materials and Methods: Geneious



Topic: Primer design and ordering for integration of RAGE-transmembrane domain into scFVconstruct and nanobody-geneart construct

Investigator: Sascha

Materials and Methods: Geneious

Results:

  • scFv:startprimer-fw
    • GCCTCTAGAGCTAGCATGGCAGC
  • rvp:scfv+overhang to TEV/TMD
    • CGCCCAAGATTCCCAGGGCCAGGGCGAGGGTGCCGCTTCCGCCGCCACCGCTTCCCCCTTGAAAATATAAATTCTCTCCAGATCCCCGTTT

GATCTCCAGTTCTGTCC

  • fwp:nterm. of yfp to TMD
    • CCCTGGGAATCTTGGGCGGCCTGGGGACCGCTGCCCTCTTGATCGGCGTGATCCTGTGGCAGAGAAGGTCTGGAGTGAGCAAGGGCGAG

GAGC

  • scFv:endprimer-rv
    • GAGCTGCAGCGGCCG
  • rage:startprimer-fw_nano/Fc
    • CTTGAATTCGCGGCCG
  • rage-rv:Fc to rageTMD
    • CGCCCAAGATTCCCAGGGCCAGGGCGAGGGTGCCGCTTCCTAATAACTTCGTATAATGTATGCTATACGAAGTTATCCC
  • rage-fwp:mcherry/rageTMD
    • CCCTGGGAATCTTGGGCGGCCTGGGGACCGCTGCCCTCTTGATCGGCGTGATCCTGTGGCAGAGAAGGGGCTCCATGGTGTCCAAGG
  • rage:endprimer in mcherry/loxp
    • AAGTCACTGCAGCGGCC


2012-10-15

cell culture


Investigator:Kerstin

Topic: cell-culture

  • passaging of CHO w Zeocin
  • passaging of HT1080
  • passaging of HEK AAV 293
  • passaging of HeLa


2012-10-17

cell culture


Investigator:Kerstin

Topic: cell-culture

  • passaging of CHO w Zeocin
  • passaging of HT1080
  • passaging of HEK AAV 293
  • passaging of HeLa
  • passaging of stably transfected Clones 4.1, 4.2, 4.3, 4,4, 29.1, 29.2, 29.3 with 550µg/ml Hygromycin

2012-10-19

cell culture


Investigator:Stefan/Kerstin

Topic: cell-culture

  • passaging of CHO w Zeocin
  • passaging of HT1080
  • passaging of HEK AAV 293
  • passaging of HeLa

2012-10-22

cell culture


Investigator:Kerstin

Topic: cell-culture

  • passaging of CHO w Zeocin
  • passaging of HT1080
  • passaging of HEK AAV 293
  • passaging of HeLa
  • transfection of stably transfected clone 29.1 with Cre Recombinase
  • seeding of HT1080 in Ibidi Dish for infection with virus

2012-10-23

cell culture


Investigator:Stefan

Topic: cell-culture

  • seeding of CHO and HeLa in Ibidi Dishes for transfection with new Nanobody RAGE construct
  • infection with Virus (CFP on surface and YFP as GOI, AAV with Sortase motif)

2012-10-24

cell culture


Investigator:Stefan/Kerstin

Topic: cell-culture

  • Transfection of new Nanobody RAGE construct in CHO and HeLa
  • seeding of CHO and HeLa in Ibidi Dishes for transfection with new scFv RAGE construct
  • passaging of CHO w Zeocin
  • passaging of HT1080
  • passaging of HEK AAV 293
  • passaging of HeLa

2012-10-25

cell culture


Investigator:Stefan/Kerstin

Topic: cell-culture

  • transfection of new scFv RAGE construct
  • transfection of TAL-AID from cooperation with Freiburg iGEM Team in CHO cells (co-transfection with clone 4)
  • purification of Nanobody from supernatant of Cre recombinase transfected cells with magnetic beads
  • Western Blot of purified Nanobody

Virus

XXX

XXX

Investigator: XXX

Aim: XXX

Materials:

XXX

Method:

XXX

Results:

XXX

Further tasks

XXX
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