Difference between revisions of "Team:Paris Saclay/Notebook/August/29"

(→‎Visualization)
(→‎PCR of the plasmid extraction from clonies 4,5,9,12 containing FRB sequence in pJET plasmid)
Line 73: Line 73:
 
|}
 
|}
  
−
[[File:T--Paris Saclay--160826 gel ligation fragm1 et 2.jpg.png|400px|thumb|center|Result of the migration]]
+
GEL GEL GEL
  
 
all 4 plasmids were sent for sequencing.
 
all 4 plasmids were sent for sequencing.

Revision as of 08:30, 16 September 2016

Monday 29th August

Lab work

Visualization

PCR of the plasmid extraction from clonies 4,5,9,12 containing FRB sequence in pJET plasmid

By Mahnaz

  • 2.5 µL DreamTaq Buffer
  • 0.5 µL of dNTPs (10mM)
  • 1 µL of each primer (10µM) primer pJET R and F
  • 0.13 μl of DreamTaq Pol
  • 20 µl H2O

PCR was performed as follow:

Step Temperature Time
Initial denaturation 95°C 3 min
30 cycles 95°C 30 sec
52 30 sec
72°C 1 min
Final Extension 72°C 7min
Hold 4°C \infinity\ 
Primers used were:
Matrix Clones containing dCas9 NM - GFP 10 in pSB1C3
Primers pJET R and F
Tm 52°C
t 1 min 30

After amplification, 3 µL of each PCR products and 5 µL of DNA ladder were placed in wells and migrated at 100V during 30 min.

PCR products expected were :

PCR products Expected band size (bp)
FRB in pJET 474

GEL GEL GEL

all 4 plasmids were sent for sequencing.

NanoDrop Measurements

By Mahnaz

Sample Concentration (ng/µL)
PCR fragment FRB clone 4
 58.43
PCR fragment FRB clone 5
 320.75
PCR fragment FRB clone 9
 830.99
PCR fragment FRB clone 12
 473.37