Difference between revisions of "Team:Paris Saclay/Notebook/June/28"

(→‎Puc19-gBlocks ligation)
Line 2: Line 2:
 
=Tuesday 28th June=
 
=Tuesday 28th June=
 
==Lab work==
 
==Lab work==
 +
 +
===Interlab study===
 +
====Cytometer====
 +
"By Caroline, Alice, Lea and Charlene, with Isabelle help''
 +
 +
Cells culture created on the 27/06/2016 were analysed with cytometer.
 +
Results were as expected.
 +
 +
<!---picture--->
  
 
===Visualization===
 
===Visualization===
Line 10: Line 19:
 
====Puc19-gBlocks ligation====  
 
====Puc19-gBlocks ligation====  
 
''By Charlene''
 
''By Charlene''
 +
 
GBlocks were resuspended into 100µL of TE buffer (except for 4-1 which was resuspended into 50µL of TE) in order to get a concentration of 10ng/mL after a quick spin.
 
GBlocks were resuspended into 100µL of TE buffer (except for 4-1 which was resuspended into 50µL of TE) in order to get a concentration of 10ng/mL after a quick spin.
 
Solutions were vortexed, incubated for 20min at 50°C, vortexed another time and quickly spun down.
 
Solutions were vortexed, incubated for 20min at 50°C, vortexed another time and quickly spun down.
Line 50: Line 60:
 
# 1µL digested vector + 1µL ligase + 1µL ligation buffer + 7µL H2O
 
# 1µL digested vector + 1µL ligase + 1µL ligation buffer + 7µL H2O
 
# 1µL digested vector + 1µL ligase + 1µL ligation buffer + 7µL insert
 
# 1µL digested vector + 1µL ligase + 1µL ligation buffer + 7µL insert
 +
 +
====Transformation with ligation products====
 +
''By Caroline and Charlene''
 +
 +
DH5α cells were transformed with ligation products using the same protocol as on 24/06/2016
 +
Transformations were displayed on Petri dishes with LB medium containing 50µg/mL of ampicillin and covered with 0.5µL of 80mg/mL XGal and 0.5µL of 1M IPTG.
 +
 +
 +
===Bringing DNA closer===
 +
====Plasmid extraction====
 +
''By Naiane and Lea''
 +
 +
Plasmids from cell culture of the 27/06/2016 were extracted following the same protocol as on 24/06/2016.
 +
 +
====Cells transformation====
 +
''By Naiane''
 +
 +
Extracted plasmids (2µL) were transformed into DH5α competent cells (50µL) using the same protocol as on 24/06/2016.
 +
 +
 +
===BioBrick characterization===
 +
====Plasmid extraction====
 +
''By Lea''
 +
 +
The same protocol as on 24/06/2016 was used to extract K1372001 from DH5α cells, except the phenol chloroform step was not done.
  
 
{{Team: Paris_Saclay/notebook_footer}}
 
{{Team: Paris_Saclay/notebook_footer}}

Revision as of 14:51, 4 July 2016

Template:Team: Paris Saclay/notebook header

Tuesday 28th June

Lab work

Interlab study

Cytometer

"By Caroline, Alice, Lea and Charlene, with Isabelle help

Cells culture created on the 27/06/2016 were analysed with cytometer. Results were as expected.


Visualization

Puc19 digestion

Puc19 was digested as expected.

Puc19-gBlocks ligation

By Charlene

GBlocks were resuspended into 100µL of TE buffer (except for 4-1 which was resuspended into 50µL of TE) in order to get a concentration of 10ng/mL after a quick spin. Solutions were vortexed, incubated for 20min at 50°C, vortexed another time and quickly spun down.

gBlock 1-1 1-2 2-1 3-1 3-2 4-1 4-2 GFP1-9
Size (bp) 960 960 1023 960 960 706 1288 862
vector size/insert size x 3.5 9.84 9.84 9.24 9.84 9.84 13.39 7.34 10.96

3 controls were made :

  1. 1µL digested vector + 9µL H2O
  2. 1µL digested vector + 1µL ligase + 1µL ligation buffer + 7µL H2O
  3. 1µL digested vector + 1µL ligase + 1µL ligation buffer + 7µL insert

Transformation with ligation products

By Caroline and Charlene

DH5α cells were transformed with ligation products using the same protocol as on 24/06/2016 Transformations were displayed on Petri dishes with LB medium containing 50µg/mL of ampicillin and covered with 0.5µL of 80mg/mL XGal and 0.5µL of 1M IPTG.


Bringing DNA closer

Plasmid extraction

By Naiane and Lea

Plasmids from cell culture of the 27/06/2016 were extracted following the same protocol as on 24/06/2016.

Cells transformation

By Naiane

Extracted plasmids (2µL) were transformed into DH5α competent cells (50µL) using the same protocol as on 24/06/2016.


BioBrick characterization

Plasmid extraction

By Lea

The same protocol as on 24/06/2016 was used to extract K1372001 from DH5α cells, except the phenol chloroform step was not done.

Template:Team: Paris Saclay/notebook footer