Ebook BRS Genetics: Part 2
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Ebook BRS Genetics: Part 2
chapter10ChromosomalMorphology MethodsI.STUDYING HUMAN CHROMOSOMES■Mitotic chromosomes are fairly easy to study because they can be observed in any ce Ebook BRS Genetics: Part 2ell undergoing mitosis.■Melotlc chromosomes are much more difficult to study because they can be observed only in ovarian or testicular samples. In the female, meiosis is especially difficult because meiosis occurs during fetal development. In the male, meiotic chromosomes can be studied only in a t Ebook BRS Genetics: Part 2esticular biopsy of an adult male.■Any tissue that can be grown in culture can be used for karyotype analysis, but only certain tissue samples are suiEbook BRS Genetics: Part 2
table for some kinds of studies. For example, chorionic villi or amniocytes from amniotic fluid are used for prenatal studies; bone marrow is usually chapter10ChromosomalMorphology MethodsI.STUDYING HUMAN CHROMOSOMES■Mitotic chromosomes are fairly easy to study because they can be observed in any ce Ebook BRS Genetics: Part 2nexplained mental retardation, or any other suspected generic conditions.■Whatever the (issue used, the cells must be grown in tissue culture for some period of time until optimal growth occurs. Blood cells must have a mitogen added to the culture media to stimulate the mitosis of lympocytes, but ot Ebook BRS Genetics: Part 2her tissues can be grown without such stimulation.■Once a tissue has reached its optimal time for a harvest, colchicine (Colcemid) is added to the medEbook BRS Genetics: Part 2
ia, which arrests the cells in metaphase.■The cells are then concentrated, treated with a hypotonic solution, which aids in the spreading of the chromchapter10ChromosomalMorphology MethodsI.STUDYING HUMAN CHROMOSOMES■Mitotic chromosomes are fairly easy to study because they can be observed in any ce Ebook BRS Genetics: Part 2of methods (see below).■It is often preferable to use prometaphase chromosomes in cytogenetic analysis as they are less condensed and therefore show more detail. In cytogenetic analysis, separated prometaphase or metaphase chromosomes are identified and photographed or digitized.■The chromosomes in Ebook BRS Genetics: Part 2the photograph of the metaphase are then cut out and arranged in a standard pattern called the karyotype, or in the case of digital images, arranged iEbook BRS Genetics: Part 2
nto a karyotype with the assistance of a computer.II.STAINING OF CHROMOSOMESMetaphase or prometaphase chromosomes may be prepared for karyotype analyschapter10ChromosomalMorphology MethodsI.STUDYING HUMAN CHROMOSOMES■Mitotic chromosomes are fairly easy to study because they can be observed in any ce Ebook BRS Genetics: Part 2mosomes are not required.94Chapter 10Chromosomal Morphology Methods95A.Chromosome Banding. Chromosome banding techniques are based on denaturation and/or enzymatic digestion of DNA, followed by incorporation of a DNA-binding dye. This results in chromosomes staining as a series of dark and light ban Ebook BRS Genetics: Part 2ds.1G-Banding. G-banding uses trypsin denaturation before staining with the Giemsa dye and is now the standard analytical method in cytogenetics.a.GieEbook BRS Genetics: Part 2
msa staining produces a unique pattern of dark bands (Giemsa positive; G bands) which consist of heterochromatin, replicate in the late s phase, are rchapter10ChromosomalMorphology MethodsI.STUDYING HUMAN CHROMOSOMES■Mitotic chromosomes are fairly easy to study because they can be observed in any ce Ebook BRS Genetics: Part 2chromatin. replicate in the early s phase, are rich in G-C bases, and contain many genes.2R-Banding. R-banding uses the Giemsa dye (as above) to visualize light bands (Giemsa negative; R bands) which are essentially the reverse of the G-banding pattern. R-banding can also be visualized by G-C specif Ebook BRS Genetics: Part 2ic dyes (e.g., chromomycin A3, oligomycin. or mithramycin).3Q-Banding. Q-banding uses the fluorochrome ợuinacrine (binds preferentially to A-T bases)Ebook BRS Genetics: Part 2
to visualize Q bands which are essentially the same as G bands.4T-Banding. T-banding uses severe heat denaturation prior to Giemsa staining or a combichapter10ChromosomalMorphology MethodsI.STUDYING HUMAN CHROMOSOMES■Mitotic chromosomes are fairly easy to study because they can be observed in any ce Ebook BRS Genetics: Part 2xide denaturation prior to Giemsa staining to visualize c bands, which are constitutive heterochromatin, located mainly at the centromere.B.Fluorescence in situ Hybridization (FISH).■The FISH technique is based on the ability of single stranded DXA (i.e., a DNA probe) to hybridize (bind or anneal) t Ebook BRS Genetics: Part 2o its complementary target sequence on a unique DNA sequence that one is interested in localizing on the chromosome.■Once this unique DNA sequence isEbook BRS Genetics: Part 2
known, a fluorescent DNA probe can be constructed.■The fluorescent DNA probe is allowed to hybridize with chromosomes prepared for karyotype analysis chapter10ChromosomalMorphology MethodsI.STUDYING HUMAN CHROMOSOMES■Mitotic chromosomes are fairly easy to study because they can be observed in any ce Ebook BRS Genetics: Part 2ction of fluorescent DNA probes to a wide variety of different DNA fragments from a single chromosome.■The fluorescent DNA probes are allowed to hybridize with chromosomes prepared for karyotype analysis and thereby visualize many different loci spanning one whole chromosome (i.e., a chromosome pain Ebook BRS Genetics: Part 2t). Essentially, one whole particular chromosome will fluoresce.D.Spectral Karyotyping or 24 Color Chromosome Painting.■The spectral karyotyping technEbook BRS Genetics: Part 2
ique Is based on chromosome painting whereby DNA probes for all 24 chromosomes are labeled with five different fluorochromes so that each of the 24 chchapter10ChromosomalMorphology MethodsI.STUDYING HUMAN CHROMOSOMES■Mitotic chromosomes are fairly easy to study because they can be observed in any ce Ebook BRS Genetics: Part 2n analyze the different ratios and assign a pseudocolor for each ratio.chapter10ChromosomalMorphology MethodsI.STUDYING HUMAN CHROMOSOMES■Mitotic chromosomes are fairly easy to study because they can be observed in any ceGọi ngay
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