Team:Frankfurt/Notebook

From 2012.igem.org

(Difference between revisions)
(Kit)
(Kit)
Line 118: Line 118:
=Kit=
=Kit=
-
PCR Purification Kit
+
PCR Purification Kit <br>
-
Gel Extraction Kit
+
Gel Extraction Kit<br>
-
Midi Plasmid Preparation Kit
+
Midi Plasmid Preparation Kit<br>

Revision as of 15:03, 14 September 2012

Frankfurt logo.png
Home Team Project Organisms New Yeast RFC Notebook Registered Parts Modeling Safety Attributions Official Team Profile

You should make use of the calendar feature on the wiki and start a lab notebook. This may be looked at by the judges to see how your work progressed throughout the summer. It is a very useful organizational tool as well.

Contents

Methods and Protocols

Culture Medium

Full Medium (YEPD) for Yeast
Yeast Extract1 % (weight/volume)
Pepton2 % (w/v)
Glucose2 % (w/v)
Synthetic Complete Medium (SC) for Yeast
Yeast Nitrogen Base0.17 % (w/v)
Ammoniumsulfate0.5 % (w/v)
Glucose2 % (w/v)
Amino Acid Mix*50 ml/l
Histidin**0.25 mM
Tryptophan**0.19 mM
Leucin**0.35 mM
Uracil**0.44 mM

pH has to be regulated with KOH to pH=6.3
!* contains no His, Leu, Trp and Uracil
** addition of this components depents on the respective selection medium

SOC-Medium for Regeneration of transformed Escherichia coli`s after Electroporation
Trypton2 % (w/v)
Yeast Extract0.5 % (w/v)
NaCl10 mM
KCl2,5 mM
MgCl210 mM
MgSO410 mM
Glucose20 mM

pH has to be regulated to pH=6.8-7.0

Full Medium (LB) for E. coli
Yeast Extract0.5 % (w/v)
Trypton1 % (w/v)
NaCl0.5 % (w/v)

pH has to be regulated with NaOH to pH=7.5

Every cluture medium has to be autoclaved to be sterile.

Agar Plate

LBampicillin-Agar

Add 2 % agar to LB-medium. After autoclaving and cooling-down to 60 °C steril ampicillin is added. Plates were poured.

SCD-Agar

Add 2 % agar to SCD-medium. After autoclaving and cooling-down steril amino acid solution is added. Dependent on the respective selective medium Histidin (0.25 mM), Trypthophan (0.19 mM), Uracil (0.44 mM) or Leucin (0.35 mM) are added. Plates were poured.

YEPDG418-Agar

Add 2 % agar to YEPD-medium. After autoclaving and cooling-down sterile G418 (final concentration 2g/l) is added. Plates were poured.

Gel Electrophoresis

Agarose Gel (1x)
TAE puffer1x
Agarose1 % (w/v)

Solve agarose in TAE by boiling it. After cooling-down to 55-60 °C gel is poured.

TAE Puffer (50x) for Gel Electrophoresis
EDTA18,6 g
Tris242g
Glacial Acetic Acid57,2 ml
Purified Water1000ml

pH has to be regulated with glacial acetic acid to pH=8.

Kit

PCR Purification Kit
Gel Extraction Kit
Midi Plasmid Preparation Kit