Team:Freiburg/Notebook/20 July

From 2011.igem.org

(Difference between revisions)
(→NAME OF YOUR EXPERIMENT)
(→Precipitator)
Line 123: Line 123:
S43 + M14+P28+P20
S43 + M14+P28+P20
 +
 +
|}
 +
 +
 +
 +
 +
'''Transformation '''
 +
 +
 +
{| style="border-spacing:0;"
 +
| style="border:0.0069in solid #00000a;padding-top:0in;padding-bottom:0in;padding-left:0.075in;padding-right:0.075in;"| Name:
 +
| style="border:0.0069in solid #00000a;padding-top:0in;padding-bottom:0in;padding-left:0.075in;padding-right:0.075in;"| Date:
 +
 +
|-
 +
| colspan="2"  style="border:0.0069in solid #00000a;padding-top:0in;padding-bottom:0in;padding-left:0.075in;padding-right:0.075in;"| Continue from Date Name
 +
 +
Experiment
 +
 +
|-
 +
| colspan="2"  style="border:0.0069in solid #00000a;padding-top:0in;padding-bottom:0in;padding-left:0.075in;padding-right:0.075in;"| Project Name:
 +
 +
|}
 +
Procedure
 +
 +
 +
# take cells from -80°C freezer and put them on ice! (every eppi contains about 400 μl cells)
 +
# thaw cells on ice 20 minutes
 +
# pipette 50 μl cells and 2 μl DNA into eppi still on ice!
 +
# Incubate for 30 minutes on ice
 +
# Heat at 42°C for 60 sec
 +
# Incubate on ice for 5 minutes
 +
# Add 200 μl LB Broth
 +
# Incubate for 2 hours at 37°C (cells with lysis cassette at 30°C!!)
 +
# Plate 50 μl and 200μl on two different LB/Agar plates with appropriate antibiotic resistance
 +
 +
'''Documentation:'''
 +
 +
Why are you doing this experiment? Name of the sample? Where are they stored? Name the vector with inserts, antibiotika resistance etc.
 +
 +
 +
 +
{| style="border-spacing:0;"
 +
| style="border:0.0069in solid #00000a;padding-top:0in;padding-bottom:0in;padding-left:0.075in;padding-right:0.075in;"| 1 S39 + M14+P28+P18
 +
 +
2 S39 + M14+P28+P19
 +
 +
3 S39 + M14+P28+P20
 +
 +
4 S43 + M14+P28+P18
 +
 +
5 S43 + M14+P28+P19
 +
 +
6 S43 + M14+P28+P20
 +
 +
 +
All have cm resistance
 +
 +
in incubator 37°C overnight
 +
 +
|-
 +
| style="border-top:none;border-bottom:0.0069in solid #00000a;border-left:0.0069in solid #00000a;border-right:0.0069in solid #00000a;padding-top:0in;padding-bottom:0in;padding-left:0.075in;padding-right:0.075in;"|
|}
|}

Revision as of 15:54, 20 July 2011

Contents

green light receptor

NAME OF YOUR EXPERIMENT

Investigators:NAME


blue light receptor

NAME OF YOUR EXPERIMENT

Investigators:NAME



red light receptor

NAME OF YOUR EXPERIMENT

Investigators:NAME



Lysis cassette

NAME OF YOUR EXPERIMENT

Investigators:NAME


Precipitator

Ligation


Name: Ruediger Date: 20.07
Continue from Digestion Date 19.07 Name Ruediger

Experiment

Project Name: GFP Pbd

Procedure


PCR tube:

total volume 20 μl


  1. add H2O (17 μl -X-Y-Z)
  2. add 2 μl Ligase Buffer 10x
  3. add Insert 1, Insert 2(when proceeding from 3A digestion use 2 μl of each)
  4. add Vector (20ng needed. When proceeding from 3A digestion use 2 μl)
  5. Add 1 μl T4-DNA Ligase
  6. Incubate 10-30 min at room temperature
  7. heat for 20 minutes at 80°C
  8. store at -20°C or directly proceed to transformation


Name of part Ratio Insert:Vector

= 3:1 or 1:1

Volume (μl)
X insert 1 -- -- --
Y insert 2 M14+P28+P18/19/20 2.7:1 2 each
Z vector S39 / S40 2 each
H2O

Documentation:

Why are you doing this experiment? Where are your parts stored? Name the parts for ligation etc.


Ligation of 3 different GFP Pbd part (M14+P28+P18/19/20) into two vector with Promotor and RBS


S39 + M14+P28+P18

S39 + M14+P28+P19

S39 + M14+P28+P20


S43 + M14+P28+P18

S43 + M14+P28+P19

S43 + M14+P28+P20



Transformation


Name: Date:
Continue from Date Name

Experiment

Project Name:

Procedure


  1. take cells from -80°C freezer and put them on ice! (every eppi contains about 400 μl cells)
  2. thaw cells on ice 20 minutes
  3. pipette 50 μl cells and 2 μl DNA into eppi still on ice!
  4. Incubate for 30 minutes on ice
  5. Heat at 42°C for 60 sec
  6. Incubate on ice for 5 minutes
  7. Add 200 μl LB Broth
  8. Incubate for 2 hours at 37°C (cells with lysis cassette at 30°C!!)
  9. Plate 50 μl and 200μl on two different LB/Agar plates with appropriate antibiotic resistance

Documentation:

Why are you doing this experiment? Name of the sample? Where are they stored? Name the vector with inserts, antibiotika resistance etc.


1 S39 + M14+P28+P18

2 S39 + M14+P28+P19

3 S39 + M14+P28+P20

4 S43 + M14+P28+P18

5 S43 + M14+P28+P19

6 S43 + M14+P28+P20


All have cm resistance

in incubator 37°C overnight

Retrieved from "http://2011.igem.org/Team:Freiburg/Notebook/20_July"