Cytogenetics

Question 1

What is cytogenetics?

Answer 1

The study of the structure, inheritance, and behaviour of chromosomes

Question 2

When did we know about the chromosome counts in human?

Answer 2

1956. Counts before then were consistently low

Question 3

Why were we constantly getting chromosome counts in humans wrong in the early days of cytogenetics?

Answer 3

Low resolution. So much so that we thought there might be natural variation in the number of chromosomes between individuals

Question 4

What contribution did Painter make to cytogenetics?

Answer 4

He examined spermatogonial metaphase spreads from paraffin embedded testicular tissue in 1923. Saw 46 chromosomes in his best work, but doubted himself and estimated the counts to be between 45 to 48. Later decides 48

Question 5

What was the problem with using paraffin embedded tissue to count chromosomes?

Answer 5

Multiple focal planes

Question 6

What 3 improvements did Tjio and Levan have over Painter in 1956?

Answer 6

1. Microscopy improvements in lenses and resolution
2. Cell suspensions instead of paraffin embedded tissue
3. Hypotonic solutions

Question 7

Why does having the cells in a hypotonic solution give better images of chromosomes?

Answer 7

Water will enter the cells and cause them to swell, and the pressure allows the cells to lyse easier when pipetted onto a microscope slide. Gives a better spread of the chromosomes

Question 8

What is the best time to view chromosomes?

Answer 8

Metaphase. It’s when they’re the most condensed

Question 9

How do we keep the cells in metaphase long enough to view chromosomes?

Answer 9

Use colchicine or colcemid to block the mitotic spindle from forming and stop the cell cycle from progressing

Question 10

How were chromosomes numbered?

Answer 10

Size. Chromosome 1 is the largest and chromosome 22 is the smallest

Question 11

Did we get the numbering of chromosomes right just by looking at them?

Answer 11

Pretty close. 20 is slightly bigger than 19, and 22 is slightly bigger than 21, but we didn’t know that until sequencing became a thing. Not worth changing it now

Question 12

What are 4 criteria we use to identify which chromosome we’re looking at?

Answer 12

1. Size
2. Centromere position
3. Secondary invaginations
4. Banding

Question 13

What are the 4 centromere positions? How do the lengths of the p and q arms compare in each type?

Answer 13

1. Metacentric - centromere is right in the middle, and p and q arms are about the same length
2. Submetacentric - centromere is slightly off centre, and the p arm is a little shorter than the q arm
3. Acrocentric - centromere is in the subtelomeric region, and the p arm is much shorter than the q arm
4. Telocentric - centromere is right at the telomere and the p arm is essentially non-existent. None of these in humans

Question 14

What is a secondary invagination?

Answer 14

The stalk of acrocentric chromosomes. Less of a constriction than the centromere, but still pretty substantial. The region on the other side is the satellite region

Question 15

What is special about the sequence in stalk regions of acrocentric chromosomes?

Answer 15

Location of rRNA genes

Question 16

What are chromosome groups? How are chromosomes grouped?

Answer 16

Groups A to G. Grouped based on size and centromere position. Larger and more metacentric chromosomes are in earlier groups and smaller and less metacentric chromosomes are in later groups

Question 17

What is a karyotype?

Answer 17

A photomicrograph of the chromosomes of an individual arranged into their groups with sex chromosomes last

Question 18

What was Torbjorn Caspersson’s banding protocol?

Answer 18

Q-banding. He used quinacrine mustard or quinacrine dihydrochloride dyes to produce a distinct dark and light banding pattern along the chromosomes. Deletions, inversions, and translocations were detectable

Question 19

What were 2 problems with Caspersson’s Q-banding method?

Answer 19

Quinacrine mustard and quinacrine dihydrochloride were fluorescent stains

1. Fluorescent microscopes were scarce back in the late 1960s
2. Quenching was an issue

Question 20

What is an ideogram?

Answer 20

A diagram of an individual’s karyotype that isn’t an actual photo

Question 21

What are the 8 steps in generating a karyotype?

Answer 21

1. Draw venous blood from the patient to get actively dividing cells
2. Culture blood cells with a mitogen (phytohemagglutinin) to induce them to divide
3. Add colchicine and hypotonic saline to halt the cells at metaphase
4. Add acetic acid and methanol to fix the chromosomes in place
5. Spread cells onto a slide by dropping them
6. Partial digest with trypsin and stain with Glemsa
7. Analyze the metaphase spread
8. Karyotype