29 Spectral Imaging Analysis of Metaphase and Interphase Cells
367
- Thymidine (Life Technologies), 105
M.
- Tween 20 (Sigma).
- Yeast artificial chromosome (YAC) library (Research Genetics). Store
at -80°e.
- Zymolase (Sigma), 70,000 U/g, 10 mglml in 50 mM KH 2 P0 4 (pH 7.8),
50% glycerol. Store at -20°e.
- AHC medium (BIO 101): Add 36.7 gIl of purified water. Autoclave at
121°C for 15 min.
- AHC agar medium (BIO 101): Add 53.7 gil of purified water. Mix well
to dissolve, then autoclave at 121°C for 15 min. Cool to 50°C, and pour
plates. Store plates at 4°e.
- Fixative: acetic acid/methanol: 1/3 (vol/vol). Make fresh just before use.
- Denaturing solution: add 35 ml formamide, 10 ml distilled water, 5 ml
20xSSe. Store at 4°e. Prepare fresh every 2-4 weeks.
- ES Buffer: 0.5 M EDTA (pH 8.0),1% Sarcosyl. Store at room temperature.
- Ethanol series: prepare 70%, 80%, and 100% ethanol in distilled water.
- 1% Formaldehyde: add 1.35 ml of 37% Formaldehyde to 50 ml of
1xPBS/MgCh. Store at 4°C.
- FHM: 1xFHM HEPES Buffered Media (Speciality Media). Store at
-20°C.
- Formamide/SSC wash solution: ad$l 15 ml 20xSSC, 60 ml distilled
water, 75 ml formamide. pH 7.0. Store at 4°e.
- Hank's BSS (Life Technologies).
- Hybridization Master Mix (MM 2.1): 14.3% w/vol dextran sulfate,
78.6% formamide, 2.9xSSC, pH 7.0. Prepare by combining 1.45 ml
of20x SSC, 0.7 ml ultrapure water, 1.43 g dextran sulfate (Calbiochem),
and 7.86 ml formamide. Heat to 70°C to dissolve dextran sulfate. Aliquot in 1.5-ml tubes and store at -20°e.
- Modified nucleotide mix (lOx A4) for labeling in combination with
1 mM digoxigenin-ll-dUTP, 1 mM fluorescein-12-dUTP or 1 mM
Cy3-dUTP: combine 5 ~l each of 100 mM dATP, 100 mM dGTP
Buffers and other
solutions
367
- Thymidine (Life Technologies), 105
M.
- Tween 20 (Sigma).
- Yeast artificial chromosome (YAC) library (Research Genetics). Store
at -80°e.
- Zymolase (Sigma), 70,000 U/g, 10 mglml in 50 mM KH 2 P0 4 (pH 7.8),
50% glycerol. Store at -20°e.
- AHC medium (BIO 101): Add 36.7 gIl of purified water. Autoclave at
121°C for 15 min.
- AHC agar medium (BIO 101): Add 53.7 gil of purified water. Mix well
to dissolve, then autoclave at 121°C for 15 min. Cool to 50°C, and pour
plates. Store plates at 4°e.
- Fixative: acetic acid/methanol: 1/3 (vol/vol). Make fresh just before use.
- Denaturing solution: add 35 ml formamide, 10 ml distilled water, 5 ml
20xSSe. Store at 4°e. Prepare fresh every 2-4 weeks.
- ES Buffer: 0.5 M EDTA (pH 8.0),1% Sarcosyl. Store at room temperature.
- Ethanol series: prepare 70%, 80%, and 100% ethanol in distilled water.
- 1% Formaldehyde: add 1.35 ml of 37% Formaldehyde to 50 ml of
1xPBS/MgCh. Store at 4°C.
- FHM: 1xFHM HEPES Buffered Media (Speciality Media). Store at
-20°C.
- Formamide/SSC wash solution: ad$l 15 ml 20xSSC, 60 ml distilled
water, 75 ml formamide. pH 7.0. Store at 4°e.
- Hank's BSS (Life Technologies).
- Hybridization Master Mix (MM 2.1): 14.3% w/vol dextran sulfate,
78.6% formamide, 2.9xSSC, pH 7.0. Prepare by combining 1.45 ml
of20x SSC, 0.7 ml ultrapure water, 1.43 g dextran sulfate (Calbiochem),
and 7.86 ml formamide. Heat to 70°C to dissolve dextran sulfate. Aliquot in 1.5-ml tubes and store at -20°e.
- Modified nucleotide mix (lOx A4) for labeling in combination with
1 mM digoxigenin-ll-dUTP, 1 mM fluorescein-12-dUTP or 1 mM
Cy3-dUTP: combine 5 ~l each of 100 mM dATP, 100 mM dGTP
Buffers and other
solutions