Karyotyping

To prepare a karyotype (the chromosome constitution of a cell) a series of metaphase preparations is photographed and the individual chromosomes cut out and arranged in order of decreasing size (an Idiogram — Fig.7.6). In most cases groups of chromosomes are apparent — e.g. in man there are seven groups — but seldom is it possible to identify individual chromosomes solely on the basis of size and position of the centromere. 

Karyotyping has advanced dramatically since it was discovered that certain stains stain individual chromosomes in characteristic ways. Caspersson (Caspersson et al., 1970a,b, 1971) showed that quinacrine mustard and quinacrine dihydrochloride (Fig. 7.7) produced a characteristic banding pattern (Q-banding), and later it was found that complementary bands could be formed with Giesma (G-bands) if the chromosomes are treated first with trypsin or mild alkali (Patil et al., 1971 Seabright, 1971 Wang and Federoff, 1972). 

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Fig. 2. Karyotype of 10 S. cerevisiae strains. Intact chromosomes have been separated electrophoreticaUy by size in a TAPE gel. (a), gel mn to separate most chromosomes. Large chromosomes are at the top, smaller at the bottom. Since most strains are polyploid, more than 16 bands may be observed, (b). Chromosomes from the same strains have been separated in a gel mn to enhance resolution of the smaller chromosomes, corresponding to the 4—5...

Figure 36-6. A human karyotype (of a man with a normal 46,XY constitution), in which the metaphase chromosomes have been stained by the Giemsa method and aligned according to the Paris Convention. (Courtesy of H Lawce and F Conte.)...

Cytogenetic Analysis and Sex Determination. Since amnlotlc fluid cells are fetal In origin, they have the chromosome complement of the fetus Itself Therefore, the chromosomes of the fetus can be examined by karyotyping the cultured amnlotlc fluid cells There are two principal reasons for examining fetal chromosomes One, the numerically less frequent reason. 

While all chromosomal abnormalities can In principle be detected by prenatal diagnosis, fetal karyotyping Is not at present being done as a general screening procedure Rather,... 

Zellweger, H. and Simpson, J. "Is Routine Prenatal Karyotyping Indicated in Pregnancies of Very Young Women "... 

Karyotypic Number, size, and structure of cell chromosomes. 

Figure 9.5 Karyotypic changes during initiation, promotion, and progression stages of carcinogenesis.

Goldstein, P. and Triantaphyllou, A.C. (1979) Karyotype analysis of the plant-parasitic nematode Heteroderaglycinesby electron microscopy. II. The tetraploid and an aneuploid hybrid. Journal of Cell Science 43, 225. 

Wolman, S. R., Karyotypic progression in human tumors, Cancer Metastasis Reviews, 2 257-293 (1983). 

Nadzhafova, R.S., N.S. Bulatova, A.I. Kozlovskii, and I.N. Ryabov. 1994. Identification of a structural chromosomal rearrangement in the karyotype of a root vole from Chernobyl. Russian Jour Genet. 30 318-322... 

Lamborot, M. and Alvarez-Sarret, E. (1989) Karyotypic characterization of some Liolaemus lizards in Chile (Iguanidae). Genome 32, 393-403. 

Lamborot, M., Espinoza, A. and Alvarez, E. (1979) Karyotypic variation in Chilean lizards of the genus Liolaemus (Iguanidae). Experientia 35, 593-595. 

The TK+/ fine was originally isolated as a spontaneously arising revertant clone from a UV-induced TK / clone. The parental TK+/+ cell and the heterozygote were then the only TK-competent mouse lymphoma cells that could be maintained in THMG medium (3 pg ml-1 thymidine, 5 pg ml-1 hypoxanthine, 0.1 pg ml-1 methotrexate and 7.5 pg ml-1 glycine) (Clive, 1987). Thus, like most established lines, these cells are remote from wild-type cells. The karyotype of the TK+/ —3.7.2C line has a modal chromosome number of 40 like wild-type, but has a variety of chromosomal rearrangements and centromeric heteromorphisms (Blazak et al., 1986). 

Acridinium ester—labeled chemiluminescent probes have been utilized to detect the specific protein-coding transcripts and to distinguish between transcripts that code for the 190-kDa protein and the two closely related 210-kDa proteins. The assay is called the hybridization protection assay (D3). In this assay, RNA isolated from the patient s white blood cells is first amplified by PCR. The amplified product is incubated with the chemiluminescent probe. The unhybridized probe is removed by selective hydrolysis in sodium tetraborate buffer, containing surfactant Triton X-100 at pH 8.5, in an incubation step at 60°C for 6 min. After the sample is cooled to room temperature, the chemiluminescence of the hybridized probe is measured in a luminometer. The procedure is reported to detect one leukemic cell in a population of a million or more normal cells. It is also rapid, requiring less than 30 min. Its reliability has been attested to by correlation with results obtained on karyotypic and Southern blot analysis (D3). 

This picture of the human chromosomes lined up in pairs is called a karyotype. 

Karyotype The characteristic chromosome complement of an individual, race, or species as defined by their number, size, shape, etc. [NIH]... 

Another important feature of the transformed/immortalized cells is their karyotype. While in the early stages of transformation, most of the cells will retain a near diploid karyotype. However, once the cells have gone through a crisis event or are growing well beyond the typical crisis period (— 15—30 subcultures), they are aneuploid with a characteristic distribution of individual chromosomes, that is, there are likely to be multiple copies of specific chromosomes while others are not cytogenetically detectable. 

Kazmierczak, B., B. Thode, S. Bartnitzke, J. Bullerdiek, and W. Schloot. 1992. Pleomorphic adenoma cells vary in their susceptibility to SV40 transformation depending on the initial karyotype. Genes Chromosomes Cancer 5(1) 35-9. 

During mitosis, aU the DNA is highly condensed to allow separation of the sister chromatids. This is the only time in the ceE cycle when the chromosome structure is visible. Chromosome abnormalities may be assessed on mitotic chromosomes by karyotype analysis (metaphase chromosomes) and by banding techniques (prophase or prometaphase), which identify aneu-ploidy, translocations, deletions, inversions, and duplications. 

A normal male karyotype is labeled 46,XY (46 chromosomes, including one X and one Y). Similarly, a normal female karyotype is labeled 46,XX. A complex nomenclature has been devised to describe deviations from these karyotypes. This nomenclature is given in Table II-3-1. 

Table II-3-1. Common Symbols Used in Karyotype Nomenclature...

Klinefelter syndrome (47,XXY) is seen in approximately 1 in 1,000 males (all individuals with this condition have a male phenotype because they have a Y chromosome). Klinefelter males are nearly always sterile because of atrophy of the seminiferous tubules. They tend to be taller than average, with abnormally long arms and legs. Breast development (gynecomastia) is seen in about one third of cases, and the IQ is on average 10-15 points below that of unaffected siblings. Individuals have also been seen with 48.XXXY and 49,XXXXY karyotypes the additional X chromosomes produce a more severely affected phenotype. 

Other common, but less clinically significant, sex chromosome aneuploidies are the 47,XXX (1/1,000 females) and 47,XYY (1/1,000 males) karyotypes. The former karyotype is associated with increased stature and a mild decrease in IQ, and the latter is associated with increased stature, mild decrease in IQ, acne, and a susceptibility to impulsive behavior (early studies suggesting a propensity to violent criminal behavior have not been substantiated). 

The cri-dii-chat syndrome, which is the result of a terminal deletion of the short arm of chromosome 5 (karyotype 4 ,XY,del[5p]), is a classic example of a chromosome deletion. The name is derived from the French term "cry of the cat," which denotes the characteristic kittenlike cry of babies with this condition. It is rare, affecting approximately 1 in 50,000 newborns. Characteristic features include mental retardation (average IQ of 35), miaocephaly, and wide-set eyes. Most children born with this condition do not survive to adulthood. 

Several other types of structural abnormalities are seen in human karyotypes. In general, their frequency and clinical consequences tend to be less than those of translocations and deletions. 

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