Chromosome banding

Question 1

What is the most commonly used banding method in North America?

Answer 1

G-banding

Question 2

What are the dark bands created by G-banding called? What do they contain?

Answer 2

G-bands. They have a low G/C content and a low gene density, and are more heterochromatic

Question 3

What are the light bands created by G-banding called? What do they contain?

Answer 3

G-negative bands. They have a high G/C content and a high gene density, and are more euchromatic

Question 4

What is the 400 band resolution?

Answer 4

The number of bands you get if you G-band chromosomes at the height of metaphase, when the chromosomes are the most condensed

Question 5

How do you increase resolution of banding?

Answer 5

Harvest cells and stain earlier to get less condensation

Question 6

Why do we want higher resolution when karyotyping?

Answer 6

Makes it easier to detect translocations

Question 7

What is the 550 band resolution?

Answer 7

The number of bands you get if you G-band chromosomes in early metaphase, when the chromosomes are less condensed

Question 8

What is the 800 band resolution?

Answer 8

The number of bands you get if you G-band chromosomes in late prophase, when the chromosomes are past the spaghetti phase but are less condensed

Question 9

How do you do high resolution banding?

Answer 9

Add a bunch of dTTP to mess with the nucleotide balance and stall the cells in S phase and synchronize them. Then time your harvest to view the chromosomes in late prophase

Question 10

What is the standard notation for nomenclature of chromosome bands?

Answer 10

Chromosome arm (p or q), region, band, period, sub-band, sub-sub-band. Each number is read separately

Question 11

What defines a region on a chromosome? Which direction does the numbering go?

Answer 11

Regions are defined by distinct morphological features like centromeres, constrictions, and telomeres. The numbering starts at 1 near the centromere and goes out towards the telomere

Question 12

Is region p1 more proximal or distal to the centromere than region p2?

Answer 12

Proximal

Question 13

Is band p15 more proximal or distal to the centromere than band p12?

Answer 13

Distal

Question 14

Is sub-band p13.4 more proximal or distal to the centromere than sub-band p13.1?

Answer 14

Distal

Question 15

Is sub-sub-band p11.43 more proximal or distal to the centromere than sub-sub-band p11.45?

Answer 15

Proximal

Question 16

Where does the “G” in G-banding come from?

Answer 16

Giemsa, the stain

Question 17

Why do we do a controlled trypsin digestion before G-banding?

Answer 17

Without it, the entire chromosome stains dark and we don’t get any bands

Question 18

What is the advantage of G-banding over Caspersson’s Q-banding method?

Answer 18

Get the same pattern, but the staining is permanent. No need to worry about quenching

Question 19

What is the name for a chromosome that has undergone a translocation?

Answer 19

A derivative chromosome

Question 20

What is R-banding?

Answer 20

Reverse banding. Staining procedure that creates the opposite pattern as G-banding

Question 21

What is the difference in the procedure between G-banding and R-banding?

Answer 21

R-banding adds one more step, which is heat denaturation in saline before the trypsin digestion and giemsa staining

Question 22

Why does the heat denaturation step in R-banding create the opposite banding pattern as G-banding?

Answer 22

The A/T rich regions of the G bands gets denatured and can’t hold the stain, so they stay light coloured and the G/C rich G-negative bands stay together and get stained instead

Question 23

What is chromomycin A3?

Answer 23

Fluorescent dye that binds to G/C rich regions and produces the same banding pattern as R-banding

Question 24

What is acridine orange staining?

Answer 24

A modified version of the R-banding protocol. Acridine orange is an intercalating agent that fluoresces orange when bound to ssDNA and fluoresces yellow/green when bound to dsDNA

Question 25

What are the two bands we get with acridine orange staining?

Answer 25

Orange bands are the A/T rich G-bands and are from acridine orange binding to ssDNA that was heat denatured
Yellow/green bands are the G/C rich R-bands that are from acridine orange binding to dsDNA

Question 26

What are T-bands?

Answer 26

A subset of highly intense R-bands near the telomere

Question 27

How do you do T-banding?

Answer 27

Use much harsher heat denaturation

Question 28

What are C-bands?

Answer 28

A subset of G-bands localized to constitutive heterochromatin

Question 29

How do you do C-banding?

Answer 29

BaOH treatment instead of heat denaturation before staining

Question 30

What is NOR staining?

Answer 30

Staining rRNA genes in the stalk of acrocentric chromosomes - nucleolar organizing regions

Question 31

What are 2 ways to do NOR staining?

Answer 31

1. Silver nitrate

2. Fluorescent probes for rRNA genes and in situ hybridization

Question 32

What are fragile sites?

Answer 32

Non-staining regions at characteristic sites on several chromosomes

Question 33

How do you view fragile sites?

Answer 33

Treat the cells with something that inhibits DNA synthesis, like having high nucleotide concentrations