270
JULIANNE MEYNE, SUSAN M. BAILEY, and EDWIN H. GOODWIN
Results
CO-FISH and COD-FISH have been used in a variety of studies of chromosome structure and organization. Figure 2 shows images of chromosomes after strand-specific hybridizations with various repetitive DNA
probes. Additional images and details of using CO-FISH and CODFISH for cytogenetic analysis of inversions, isochromosomes and lateral
asymmetry are available in the references listed at the end of this chapter.
Recently, the CO-FISH method has been used in combination with wholechromosome paint probes for measuring chromosome aberration frequencies in first-versus-second division cells for biological dosimetry (Jordan et al., 1999).
Comments
1. Long-wave UV (about 365 nM) is preferable for inducing nicks in the
Hoechst 33258 stained DNA, as short-wave UV can nick unsubstituted
DNA. The exposure time will vary with the type and wavelength of the
UV lamp used. If a sunlamp is used, place a thin sheet of plastic between the UV source and the slides to protect the DNA from dimer
formation that will interfere with removal of the nicked DNA strand.
Use caution to avoid optic exposure when using UV light. An enclosed
light source is best. We use a Stratalinker 1800 UV Irradiator (Stratagene, La Jolla, CA). Alternately, wear eyeglasses or a face shield specifically designed for use with UV light.
2. There are a variety of methods for preparation of single-stranded DNA
probes, including oligonucleotide synthesis and PCR. We have worked
primarily with repetitive DNA sequences and find that synthetic oligonucleotide probes provide the brightest signals with the lowest background levels. This is especially true when probes are labeled directly
with fluorochrome-tagged molecules.
References
Bailey SM, Meyne J, Cornforth MN, McConnell IS, Goodwin EH (1996) A new method
for detecting pericentric inversions using COD-FISH. Cytogenetics and Cell Genetics 75:248-253
Bailey SM, Goodwin EH, Meyne J, Cornforth MN (1996) CO-FISH reveals inversions
associated with isochromosome formation. Mutagenesis 11:139-144
JULIANNE MEYNE, SUSAN M. BAILEY, and EDWIN H. GOODWIN
Results
CO-FISH and COD-FISH have been used in a variety of studies of chromosome structure and organization. Figure 2 shows images of chromosomes after strand-specific hybridizations with various repetitive DNA
probes. Additional images and details of using CO-FISH and CODFISH for cytogenetic analysis of inversions, isochromosomes and lateral
asymmetry are available in the references listed at the end of this chapter.
Recently, the CO-FISH method has been used in combination with wholechromosome paint probes for measuring chromosome aberration frequencies in first-versus-second division cells for biological dosimetry (Jordan et al., 1999).
Comments
1. Long-wave UV (about 365 nM) is preferable for inducing nicks in the
Hoechst 33258 stained DNA, as short-wave UV can nick unsubstituted
DNA. The exposure time will vary with the type and wavelength of the
UV lamp used. If a sunlamp is used, place a thin sheet of plastic between the UV source and the slides to protect the DNA from dimer
formation that will interfere with removal of the nicked DNA strand.
Use caution to avoid optic exposure when using UV light. An enclosed
light source is best. We use a Stratalinker 1800 UV Irradiator (Stratagene, La Jolla, CA). Alternately, wear eyeglasses or a face shield specifically designed for use with UV light.
2. There are a variety of methods for preparation of single-stranded DNA
probes, including oligonucleotide synthesis and PCR. We have worked
primarily with repetitive DNA sequences and find that synthetic oligonucleotide probes provide the brightest signals with the lowest background levels. This is especially true when probes are labeled directly
with fluorochrome-tagged molecules.
References
Bailey SM, Meyne J, Cornforth MN, McConnell IS, Goodwin EH (1996) A new method
for detecting pericentric inversions using COD-FISH. Cytogenetics and Cell Genetics 75:248-253
Bailey SM, Goodwin EH, Meyne J, Cornforth MN (1996) CO-FISH reveals inversions
associated with isochromosome formation. Mutagenesis 11:139-144