Most of chromosomes, at the prophase and the metaphase, are characterized by a banding pattern. But this banding pattern is more evident and clear in the case of larger chromosomes such as the polytene chromosomes of drosophila melanogaster, or the fruit fly. The banding patterns are the regions rich in heterochromatin, where the histone-DNA interaction is more. These complexes can be stained very easily by the conventional nuclear dyes or chromosomal dyes such as orcein. The regions between the bands are actually the active regions of the chromatin where more genes are present, but the quantity of DNA is very low and therefore the histone proteins. That is why they appear unstained or colored lightly. These interband regions can be detected with immuno-staining by using fluorescently labeled antibodies against the DNA-dependent RNA polymerase, which is usually seen with euchromatin regions required for the process of transcription.
Specialized staining techniques are now available, which enable one to differentiate or precisely identify individual chromosome homologes, chromosome regions, and/ or chromosome bands. A renewed interest in the chromosomal or cytogenetic status of various species has been generated by the advancements of genetic mapping techniques utilizing fluorescence in situ hybridization or FISH. Depending upon the type of dye or fluorochrome or the chromosome pretreatment, there can be different types of banding patterns. They include banding patterns such as G-banding, Q-banding, C-banding, and R-banding. The data generated by multiple chromosome banding techniques can be used for karyotypic analysis.
Q-banding: This banding pattern is obtained by treating with a fluorochrome or the fluorescent dye quinacrin. They can be identified by a yellow fluorescence of different intensity. Most parts of the stained DNA are heterochromatin. Quinacrin binds those regions which are rich in AT and G-C, but fluorescences only A-T-quinacrin regions. A-T regions are seen more in heterochromatin than in euchromatin. Therefore, by this banding method heterochromatin regions are labeled preferentially. The characters of the banding regions and the specificity of the fluorochrome are not exclusively dependent on their affinity to regions rich in A- T, but it depends on the distribution of A- T and its association with other molecules such as histone proteins.
G-banding: This technique is not a fluorochrome-based pretreatment. It is well suited L "lr animal cells. It resembles the C-banding technique without pretreatrne.1.t. During mitosis, the 23 pairs of human chromosomes condense and are visible with a light microscope. A karyotype analysis usually involves blocking cells in mitosis and staining the condensed chromosomes with Giemsa dye. The dye stains regions of chromosomes that are rich in the base pairs Adenine (A) and Thymine (T) producing a dark band. A common misconception is that bands represent single genes, but in fact the thinnest bands contain over a million base pairs and potentially hundreds of genes. For example, the size of one small band is about equal to the entire genetic information for one bacterium.
C-banding: The name C-banding originated from centromeric or constitutive heterochromatin. The centromere appears as a stained band compared to other regions. The technique involves a pretreatment with alkali before staining. The alkaline pretreatment leads to the complete depurination of the DNA. The remaining DNA is again renatured and stained with Giemsa solution consisting of methylene azure, methylene violet, methylene blue, and eosin. In this staining the heterochromatin take a lot of stain but the rest of the chromosomes stain only a little. This banding technique is well suited for the characterization of plant chromosomes.
R-banding:This is known as a reverse banding technique. This technique results in the staining of areas rich in G-C that is typical for euchromatins. G-, Q-, and R-bandings are not observed with plant chromosomes.
Hy-banding:This is a common technique used with plant cells. The technique involves a pretreatment of the cells in which the cells are warmed in the presence of HCl and then stained with acetocarmine. The pattern of Hy-band is different from that of C-bands. The binding of histone protein to DNA and its complete extraction has an impact on the binding ability of acetocarmine and formation of bands.
Further variations in the procedure of the pre-treatment choice of dyes and fluorochromes further enhanced the resolution of the banding techniques. Many of the techniques are well suited for animal chromosomes, but face many difficulties with plant chromosomes. The reason for this is not well understood. The banding pattern of plant chromosomes with any of these techniques never comes to the same degree as that of animal chromosome banding patterns. The consistent banding patterns of the constitutive heterochromatin and the remaining chromatin are exactly constant in many species with an intraspecific variable karyotype.
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