C - Banding/Q - Banding/NOR Staining Flashcards
C-banding stains the constitutive heterochromatin of which chromosomes?
1, 9, 16, Y
What technique is used to identify a small marker?
DAPI/Distamycin A
What does the acronym CBG stand for?
C bands by Barium Hydroxide using Giemsa
Which one of these techniques would be MOST helpful in identifying that a marker chromosome is bisatellited?
DAPI/Distamycin staining
After G-band analysis, a cytogenetic director suspects that chromosome number 1p has a terminal deletion. What banding procedure would be most helpful in deciding if a deletion has occurred?
R-banding
Which one of these abnormalities will produce the brightest Q-band signal?
i(Y)q
What is the function of Mcllvaine’s buffer in Q-banding?
To control the pH level
What are the major clinical applications of C-banding?
1-Identify dicentric chromosomes.
2-Identify centromere region (or centromere region of ring chromosomes).
3-Differentiate active from inactive DNA, especially in marker chromosomes (larger c-band = more inactive DNA).
4-Identify variable regions of chromosomes 1qh, 9qh, 16qh, and Yqh.
What is the major clinical application of R-banding?
Studying small deletions or cryptic translocations, which involve the telomeres of chromosomes.
After C-banding, the chromosome pattern looks like solid staining. How to improve it?
This is due to undertreatment and the timing should be adjusted/increased.
What does QFQ stand for?
Q-banding by fluorescence using quinacrine
What is the preferred pH level of Mcllvaine’s buffer?
pH 5.5
Range - 4.5 to 6.0
Increase the pH to 5.5 increases Q band brightness
Decrease the pH to 4.5 increase band contrast
What is the Basic procedure for Q-banding?
- The short exposure of a chromosome preparation to a weak, buffered quinacrine solution at an acid pH (Mcllvaine’s buffer, pH 4.5-6.0)
- Wet mounting in the buffer with glycerol to slow quenching and cover with coverslip.
- Excitation by UV exposure to observe with the fluorescent microscope
What are the advantages of Q-banding?
- Extremely quick technique; can finish report the same day you harvest
- By software, it can be inverted into G-banding like pattern
- No damage to chromosome morphology and easy removal of stain by fix so easy sequential banding possible
What is the clinical application of Q-Banding?
Identifying the Y chromosome polymorphism
What are the disadvantages of Q-banding?
- Poorer quality of photography
- Expensive fluorescent microscopes required
- Fluorescence fades very fast
- Slide is not permanent
What does RHG stand for?
Reverse banding using heat and Giemsa
What is the procedure for R-Banding?
- Incubate slides in hot phosphate solution for 10 to 15 minutes.
- Cool quickly in tap water. Do not dry
- Stain in 2% Giemsa for 10 to 20 minutes.
- Rinse in xylene
- Rinse in tap water. Air dry
What are the qualities of Heterochromatin?
- Highly condensed
- At centromeres and telomeres
- Gene poor / repetitious sequences
- Replicated in late S phase
What are the qualities of Euchromatin?
- Less condensed
- At chromosome arms
- Gene rich / contains unique sequences
- Replicated throughout S phase
What are the two types of heterochromatin?
Constitutive & Facultative
What does C-banding do?
Stain the constitutive heterochromatin at the centromere.
What are 4 qualities of C-Banding?
1-Gentically inert
2-Late replicating
3-Rich in AT sequences
4- Highly repetitive DNA
No C-bands seen; chromosomes appear ghost-like and pale staining?
Overtreatment which extracted even the heterochromatin
Some other bands still visible or a solid stain?
Undertreatment
What is the most common chromosomal variant in the human race?
Inversion 9
What are ribosomal cistrons?
Ribosomal cistrons are clusters of genes(~400), which code for the ribosomes and are responsible for the formation of the nucleolus in interphase cells.
What does Ag-NOR stain?
AgNOR-banding, stains only those NOR containing genes which were transcribed during the previous cell cycle.
