Lab Solutions and Reagents

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

• culture media

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

• immunohistochemistry

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

• in situ hybridization

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)

• fixatives

MEMFA

Bouin’s Fixative

Dent's Fix (Methanol/DMSO)

Paraformaldehyde(PFA), 20%

PTw Containing 4% Paraformaldehyde

• 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

Agarose Plates, 1% (for use with trops or laevis)

Dehydrating embryos

Rehydrating Embryos(for embryos stored in 100% methanol)

• 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

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)

• high-salt solution for egg laying (Zorn lab/Ron Laskey)

• dejellying solution (Zorn lab)

• recipes for miscellaneous salts, solutions, etc. (Khokha lab)