Paris/October 20

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yesterday -- tomorrow

Contents

Digestion Reaction

PCR reaction

PCR : PrefixBB-frt1-CmR-frt2-SufixBB
PCR Settings Buffer (5x) (5x) 10µL Expected size
Annealing (°C) MgCl2 10µM 10µM 0µL (no extra) 1000bp
/ dNTP 10µM 10µM 1µL Success
Time Elongation Oligo F 10µM O54 10µM 4µL YES
72°C 1'40" Oligo R 10µM O55 10µM 4µL Image (click to enlarge)
Number cycles Water qsp 50µL [[Image:|30px]]
30 Polymerase Phusion 0,5µL Band (0=ladder)
DNA pY-2P-IntC 5µL
PCR : PrefixBB-frt1-CmR-frt2-SufixBB
PCR Settings Buffer (5x) (5x) 10µL  ! GC one ! Expected size
Annealing (°C) MgCl2 10µM 10µM 0µL (no extra) 1000bp
/ dNTP 10µM 10µM 1µL Success
Time Elongation Oligo F 10µM O54 10µM 4µL NO
72°C 1'40" Oligo R 10µM O55 10µM 4µL Image (click to enlarge)
Number cycles Water qsp 50µL [[Image:|30px]]
30 Polymerase Phusion 0,5µL Band (0=ladder)
DNA pY-2P-IntC 5µL
PCR : PrefixBB-frt1-CmR-frt2-SufixBB
PCR Settings Buffer (5x) (5x) 10µL ! GC one ! Expected size
Annealing (°C) MgCl2 10µM 10µM 0µL (no extra) 1000bp
/ dNTP 10µM 10µM 1µL Success
Time Elongation Oligo F 10µM O54 10µM 4µL NO
72°C 1'40" Oligo R 10µM O55 10µM 4µL Image (click to enlarge)
Number cycles Water qsp 50µL [[Image:|30px]]
30 Polymerase Phusion 0,5µL Band (0=ladder)
DNA pY-2P-IntC 5µL
PCR : PrefixBB-frt1-CmR-frt2-T1T2-ufixBB
PCR Settings Buffer (5x) (5x) 10µL Expected size
Annealing (°C) MgCl2 10µM 10µM 0µL (no extra) 1000bp
/ dNTP 10µM 10µM 1µL Success
Time Elongation Oligo F 10µM O54 10µM 4µL YES
72°C 1'40" Oligo R 10µM O56 10µM 4µL Image (click to enlarge)
Number cycles Water qsp 50µL [[Image:|30px]]
30 Polymerase Phusion 0,5µL Band (0=ladder)
DNA pY-2P-IntC 5µL
PCR : PrefixBB-frt1-CmR-frt2-T1T2-SufixBB
PCR Settings Buffer (5x) (5x) 10µL  ! GC one ! Expected size
Annealing (°C) MgCl2 10µM 10µM 0µL (no extra) 1000bp
/ dNTP 10µM 10µM 1µL Success
Time Elongation Oligo F 10µM O54 10µM 4µL NO
72°C 1'40" Oligo R 10µM O56 10µM 4µL Image (click to enlarge)
Number cycles Water qsp 50µL [[Image:|30px]]
30 Polymerase Phusion 0,5µL Band (0=ladder)
DNA pY-2P-IntC 5µL
PCR : PrefixBB-frt1-CmR-frt2-T1T2-SufixBB
PCR Settings Buffer (5x) (5x) 10µL ! GC one ! Expected size
Annealing (°C) MgCl2 10µM 10µM 0µL (no extra) 1000bp
/ dNTP 10µM 10µM 1µL Success
Time Elongation Oligo F 10µM O54 10µM 4µL YES
72°C 1'40" Oligo R 10µM O56 10µM 4µL Image (click to enlarge)
Number cycles Water qsp 50µL [[Image:|30px]]
30 Polymerase Phusion 0,5µL Band (0=ladder)
DNA pY-2P-IntC 5µL

Purification of Digestion and PCR products

Ligation Reactions

Culture

Launching of 80mL culture of the strain countaining the plasmid pY-2P-IntC, last minipreps revealed the weak nomber of plasmid recover with a classic miniprep on 5mL culture.