Published in Methods (San Diego, Calif.)
Genotyping methods based on nonradioactive detection of PCR products and suitable for large-scale mapping projects are described. Two alternative techniques are proposed for the genotyping of polymorphic short tandem repeats or microsatellite markers. The first is designed for investigators who do not have access to automatic sequencing machines. T...
Published in Methods (San Diego, Calif.)
Microcell fusion is a technology that involves the transfer of single or small clusters of intact chromosomes from one cell to another. The transferred chromosome can be stably retained in the recipient cell background under dominant selective pressure. Hybrid clones generated by this method result in karyotypically simple and homogeneous populatio...
Published in Methods (San Diego, Calif.)
Primed in situ labeling (PRINS) has become a routine technique for the microscopical staining of specific DNA sequences in cells and nuclei. For this purpose, the technique has the general advantage of being fast, simple, and sensitive. Furthermore, the reaction characteristics of the technique enable it to discriminate efficiently among closely re...
Published in Methods (San Diego, Calif.)
Chicken cell lines derived from avian leukosis virus-induced B-cell tumors are highly recombination-proficient. In these cells, homologous recombination between transfected DNA sequences and the corresponding chromosomal alleles occurs at high frequency, and targeting efficiencies of 10-90% can be obtained. We previously described a chromosome shut...
Published in Methods (San Diego, Calif.)
Somatic cell genetic studies gave the first proof that functional tumor suppressor genes exist in mammalian genomes. Initial studies showed that whole-cell hybrids between tumorigenic mouse or human cell lines and their normal counterparts became nontumorigenic upon inoculation into animals. However, identification of the operative tumor suppressor...
Published in Methods (San Diego, Calif.)
The combination of synthetic oligonucleotide probes and DNA ligases is central to several recently developed genetic assays. Among the advantages of ligase-mediated gene detection is that ligation of probe pairs provides highly specific detection of unique DNA sequences in genomic samples. The technique also allows for convenient distinction betwee...
Published in Methods (San Diego, Calif.)
Microcell-mediated chromosome transfer (MMCT) offers a unique method for introducing tagged individual human chromosomes from mouse/human monochromosomal hybrids into cell lines displaying recessive mutant phenotypes. Functional analysis of the resultant microcell hybrids bearing different tagged individual human chromosomes permits identification ...
Published in Methods (San Diego, Calif.)
Genetic isolates represent exceptional resources in the identification of disease loci. Not only have specific monogenic diseases become enriched and families frequent enough to be utilized in linkage analyses, but also the vast majority of patients carry the same mutation and share identical chromosomal haplotypes at significant distances. Additio...
Published in Methods (San Diego, Calif.)
A rate-limiting step in the analysis of large segments of genomic DNA is the generation of a set of representative short single-copy sequences that can be used for development of fine-structure maps. In direct response to this need, we have developed interspersed repetitive DNA sequence (IRS)-bubble PCR. IRS-bubble PCR was designed to amplify the h...
Published in Methods (San Diego, Calif.)
Fluorescence in situ hybridization (FISH) applied to metaphase chromosomes provides a mapping resolution of 1 to 3 Mb. FISH applied to interphase nuclei has a resolution of 50 kb and ranges 1-2 Mb. This better resolution is attributed to the higher degree of chromatin decondensation. Here, we describe FISH applied to naked DNA fibers (fiber FISH) a...
