Lab Solutions and Reagents: Difference between revisions

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Sive, Hazel (Author), Grainger, Robert M (Author), and Harland, Richard M (Author).
Sive, Hazel (Author), Grainger, Robert M (Author), and Harland, Richard M (Author).


Additional methodology adapted from Khokha lab protocols [http://khokha.medicine.yale.edu/protocols_general/protocols_general.html]


'''please click each bulleted list heading to see solution/reagent recipes'''
 
'''click each bulleted list heading to view solution/reagent recipes'''


*'''[[culture media]]'''
*'''[[culture media]]'''
Line 53: Line 55:
:Bleaching solution (Mayor et al. 1995)
:Bleaching solution (Mayor et al. 1995)
*'''[[fixatives]]'''
*'''[[fixatives]]'''
:MEMFA
:Bouin’s Fixative
:Dent's Fix (Methanol/DMSO)
:Paraformaldehyde(PFA), 20%
:PTw Containing 4% Paraformaldehyde
*'''[[DNA/RNA preparation]]'''
*'''[[DNA/RNA preparation]]'''
:10x mRNA Transcription Buffer
:2x mRNA Nucleotide Triphosphate Mix
:TE
:Homoginization Buffer
:Reticulocyte Standard Buffer (RSB)
:Nuclei Freezing Buffer
*'''[[embryo manipulation/dissection]]'''
*'''[[embryo manipulation/dissection]]'''
:Agarose Plates, 1% (for use with trops or laevis)
:Dehydrating embryos
:Rehydrating Embryos(for embryos stored in 100% methanol)
*'''[[microinjection]]'''
*'''[[microinjection]]'''
:Ficoll, 2.5%
:2.5% Ficoll in 1/3X MR (for use with X. laevis embryos)
:3% Ficoll in 1/9XMR (for use with X.tropicalis embryos)
*'''[[general buffers and salt solutions]]'''
*'''[[general buffers and salt solutions]]'''
:HEPES 1 M, pH 7.4
:1M MgCl2
:10x Phosphate Buffered Saline (PBS)
:20x Phosphate Buffered Saline (PBS)
:PBT
:Proteinase K(PK), 10 mg/ml
:PTw
:4M NaCl, 1 liter
:SDS 10%
:TAE 50X
:1M Tris pH 9.5




'''Additional Solutions/Reagents:'''


*[[benzocaine stock, 100x (Khokha lab)]]
*[[benzocaine stock, 100x (Khokha lab)]]


*[[high-salt solution for egg laying (Zorn lab/Ron Laskey)]]
*[[high-salt solution for egg laying (Zorn lab/Ron Laskey)]]
*[[dejellying solution (Zorn lab)]]


*[[recipes for miscellaneous salts, solutions, etc. (Khokha lab)]]
*[[recipes for miscellaneous salts, solutions, etc. (Khokha lab)]]
:agarose plates 1% - for use with laevis or tropicalis
:Alkaline Phosphatase (AP) Buffer, with Levamisol and Tween
:amphibian Ringers, 5 gal (19 L)
:bleaching solution (Sive et al. 2000, Mayor et al. 1995)
:4% BMB blocking reagent in MAB
:Bouin’s fixative
:cysteine, 2.0%
:dehydrating embryos
:Denhardt’s solution, 100X
:dNTPs 10 mM
:2.5% Ficoll
:2.5% Ficoll in 1/3X MR (for use with X. laevis embryos)
:3% Ficoll in 1/9XMR (for use with X.tropicalis embryos)
:gentamicin stock, Sigma (1000X, 100 mg/ml)
:HEPES 1 M, pH 7.4
:hybridization buffer
:levamisol 1 M
:MAB, 10X
:1M MgCl2
:MEMFA* (Sive et al. 2000)
:100X MR salts
:1/3X Modified Ringers
:1/9X MR + 1% Gentamicin
:1/20X MR
:modified Ringers
:paraformaldehyde(PFA), 20%
:phosphate buffered saline (PBS), 10X
:proteinase K(PK), 10 mg/ml
:PTw
:rehydrating embryos (for embryos stored in 100% methanol)
:RNase A (Sigma R 5000)
:RNase T1 (Sigma R8251)
:4M NaCl, 1 liter
:SDS 10%
:Sodium Phosphate Buffer(1M)
:SSC, 20X
:TAE 50X
:Triethanolamine 0.1 M (pH7.0-8.0)
:1M Tris pH 9.5

Latest revision as of 12:54, 16 December 2011

most information adapted from:

Early Development of Xenopus Laevis: A Laboratory Manual.[1]

2000; (First ed.). Cold Spring Harbor: Cold Spring Harbor Laboratory Press.

Sive, Hazel (Author), Grainger, Robert M (Author), and Harland, Richard M (Author).

Additional methodology adapted from Khokha lab protocols [2]


click each bulleted list heading to view solution/reagent recipes

Modified Barth's Saline (MBS)
Anitbiotics
Gentamycin
Oxytetracycline
Penicillin
Streptomycin Sulfate
High-salt MBS
Amphibian Ringers
Modified Frog Ringers (MR)
Marc's Modified Ringer's (MMR)
Normal Amphibian Medium (NAM)
Holtfreter's Solution
MEMFA
Dent's Fix (Methanol/DMSO)
Bleaching solution (Mayor et al. 1995)
Goat Serum (GS)
PBT
DAB
H2O2
Secondary Antibodies Coupled to Horseradish Peroxidase
Murray's Clearing Medium
Nucleotide Stocks
5x SP6 Buffer
MEMFA
PTw
Triethanolamine 0.1 M
PTw Containing 4% Paraformaldehyde
Hybridization Buffer
SSC, 20X
Denhardt’s solution, 100X
RNase A
RNase T1
4% BMB Blocking Reagent in MAB
Alkaline Phosphatase Buffer
Levamisol 1 M
NBT/BCIP
PVA Alkaline Phosphatase Buffer
Bouin’s Fixative
Bleaching solution (Mayor et al. 1995)
MEMFA
Bouin’s Fixative
Dent's Fix (Methanol/DMSO)
Paraformaldehyde(PFA), 20%
PTw Containing 4% Paraformaldehyde
10x mRNA Transcription Buffer
2x mRNA Nucleotide Triphosphate Mix
TE
Homoginization Buffer
Reticulocyte Standard Buffer (RSB)
Nuclei Freezing Buffer
Agarose Plates, 1% (for use with trops or laevis)
Dehydrating embryos
Rehydrating Embryos(for embryos stored in 100% methanol)
Ficoll, 2.5%
2.5% Ficoll in 1/3X MR (for use with X. laevis embryos)
3% Ficoll in 1/9XMR (for use with X.tropicalis embryos)
HEPES 1 M, pH 7.4
1M MgCl2
10x Phosphate Buffered Saline (PBS)
20x Phosphate Buffered Saline (PBS)
PBT
Proteinase K(PK), 10 mg/ml
PTw
4M NaCl, 1 liter
SDS 10%
TAE 50X
1M Tris pH 9.5


Additional Solutions/Reagents:


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