Exemple de solution complète avec le SDS :
5 X SDS-PAGE Gel Running Buffer
15.1 g Tris base72.0 g glycine
5.0 g SDS
Bring to 1 liter with distilled water and dissolve. * Store at room temperature 1 month.
mercredi 6 juillet 2011
Préparation du running buffer pour les gels SDS
Il faut un tampon tris glycine 5x par exemple et mettre du SDS en poudre ou en liquide à 0.1% pour 1L
Exemple de solution complète avec le SDS :
72.0 g glycine
5.0 g SDS
Bring to 1 liter with distilled water and dissolve. * Store at room temperature 1 month.
Exemple de solution complète avec le SDS :
72.0 g glycine
5.0 g SDS
Bring to 1 liter with distilled water and dissolve. * Store at room temperature 1 month.
mardi 7 décembre 2010
Sequence pour tag 6xHIs
Sequence nucléotidique pour tag 6xHis : à respecter pour ne pas affecter la machinerie de transcription de E.coli
CAC CAC CAT CAT CAC CAT
CAC CAC CAT CAT CAC CAT
mercredi 17 novembre 2010
Préparation IPTG 0.5 M
Préparer de l'IPTG 0.5M, pour au final le mettre a 0.5mM dans les erlens de 1L
m= PM X V(L) X Cm (M)
m= 238 X 10 ml X 0.5M
m= 238 X 0.01 X 0.5 M
m= 1.19g
m= PM X V(L) X Cm (M)
m= 238 X 10 ml X 0.5M
m= 238 X 0.01 X 0.5 M
m= 1.19g
vendredi 15 octobre 2010
Methodes de purification des proteines menbranaires, corps d'inclusion, et complexes
Methodes de purification des proteines menbranaires, corps d'inclusion, et complexes
ici
ici
vendredi 6 mars 2009
Optimisation PCR
PCR Primer Design Guidelines
Length : 18 to 30 nucleotides
GC Content: 40% to 60%
Annealing: For annealing temperature, use 5°C to 10°C below melting temperature (Tm).
Temperature: Example Tm calculation :
Tm (°C) = 2( #A + #T ) + 4( #G + #C )
Generally, a Tm between 55°C and 80°C will yield the best results.
Sequence: Avoid polybase sequences (3 or more) Gs and Cs at the 3' end.
Avoid mismatches at the 3' end. +
Avoid complementary sequences within the primer. +
Avoid primer-dimer formation by complementarity at 3' ends of primer pairs.
PCR Reaction Condition Guidelines
Primer concentration : - Final primer concentration should be 100 - 500 nM, which is equivalent to -100 to 250 ng of an 18- to 25-mer oligonucleotide primer in a 100-ul reaction volume. +
- Primers should be salt-free and gel-purified.
Length : 18 to 30 nucleotides
GC Content: 40% to 60%
Annealing: For annealing temperature, use 5°C to 10°C below melting temperature (Tm).
Temperature: Example Tm calculation :
Tm (°C) = 2( #A + #T ) + 4( #G + #C )
Generally, a Tm between 55°C and 80°C will yield the best results.
Sequence: Avoid polybase sequences (3 or more) Gs and Cs at the 3' end.
Avoid mismatches at the 3' end. +
Avoid complementary sequences within the primer. +
Avoid primer-dimer formation by complementarity at 3' ends of primer pairs.
PCR Reaction Condition Guidelines
Primer concentration : - Final primer concentration should be 100 - 500 nM, which is equivalent to -100 to 250 ng of an 18- to 25-mer oligonucleotide primer in a 100-ul reaction volume. +
- Primers should be salt-free and gel-purified.
Pourcentage gel d'acrylamide en fonction de la taille
| Recommended Aery | amide Gel |
| Percentages for Resolution of Protein | |
| Gel percentage | DNA size range |
| 8% | 40-200 kD |
| 10% | 21-100 kD |
| 12% | 10-40 kD |
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