UNIT 7: KARYOTYPING Flashcards by jade doe

Q

is the number and appearance of chromosome in the nucleus of a eukaryotic cell

A

Karyotype

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Q

Information on the karyotype

A

SiSi Po Pre

Size of chromosome
Size of satellites
Position of centromere
Presence of secondary constrictions

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Q

T or F: Karyotyping begins with the culturing of cells

A

T

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Q

Karyotype comes from the Greek word
_______ which means ________.

A

Karyon; nucleus

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Q

the study of whole sets of chromosomes

A

Karyology

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Q

the standard
format of representing chromosomes as
diagram when the haploid set of
chromosomes of an organism are ordered
in a series of decreasing size

A

Idiogram or Karyogram

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Q

show larger differences between smaller and larger chromosomes in a set

have more acrocentric chromosomes and
relatively advanced feature

A

Asymmetric karyotype

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Q

show lesser difference between smaller and larger chromosomes in a set

have more metaphase chromosomes and no advanced feature.

A

Symmetric karyotype

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Q

In 1931 ______________, a Russian scientist suggested that in flowering plants there is a predominant trend towards karyotype asymmetry.

This trend has been carefully studied in
the genus _______ of the family compositae.

A

G.A. Levitzky; Crepis

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Q

T or F: Species showing a greater asymmetry is less advanced.

A

F

It is more advanced.

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Q

Proportion of metacentric, acrocentric chromosomes in a set.

Ratio between size of largest and smallest chromosomes in a set.

A

Degree of asymmetry

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Q

The higher the proportion of __________ chromosomes, the greater the value
of size ratio, more asymmetrical is a karyotype

A

acrocentric

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Q

is the process of pairing and ordering all the chromosomes
of an organism, thus providing a genome-wide snapshot of an
individual’s chromosomes.

A

Karyotyping

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Q

T or F: Karyotypes are prepared using standardized staining procedures that
reveal characteristic structural features for each protein.

A

F

They reveal features of each chromosome

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Q

Banding patterns are a product of what procedure?

A

staining

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Q

T or F: Karyotyping analysis focuses more on aneuploidies rather than aberrations

A

T

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Q

T or F: Telomeres can also be present in ring chromosomes

A

F

Their presence serves as a clue that you are looking at a linear chromosome

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Q

Euchromatin or Heterochromatin?

More proteins

A

Heterochromatin

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Q

Euchromatin or Heterochromatin?

Transcriptionally active

A

Euchromatin

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Q

Euchromatin or Heterochromatin?

Light staining

A

Euchromatin

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Q

Euchromatin or Heterochromatin?

Abundant GC bonds

A

Euchromatin

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Q

Euchromatin or Heterochromatin?

Abundant AT bonds

A

Heterochromatin

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Q

Causes nucleosomes to pack tightly together

A

Methylation of DNA and histones

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Q

T or F: In methylation, transcription factors cannot bind the DNA, so genes are not expressed.

A

T

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Results in loose packing of nucleosomes

Acetylation of histones

T or F: In acetylation, transcription factors can bind the DNA, so genes are expressed.

T

Materials needed for karyotyping

Sterile 5mL syringe 21G syringe needle 15mL conical tubes Green-top vacutube Glass slides Pasteur Pipette Pipettor and Pipette tips Serological Pipettes

Reagents needed for karyotyping

Glacial acetic acid Methanol KCl (hypotonic solution) RPMI Growth Medium Fetal Bovine Serum Phytohemagglutinin Colcemid Giemsa Dye Trypsin

Equipment needed for karyotyping

Centrifuge Incubator at 37°C CO2 Refrigerator Inverted Microscope

5 Major steps in karyotyping

1. Short term lymphocyte culture 2. Harvesting of lymphocytes 3. Fixing the cells 4. Making the chromosome slides 5. Slide analysis

T or F: The collected blood will be grown in vivo by adding cell culture growth medium, fetal bovine serum, antibiotics, and phytohemagglutinin (PHA) – the reagent that induces mitotic activity

F It is grown in vitro

T or F: Hypotonic medium is used for lysing the cells

T

What reagent ensures that only lymphocytes will be affected by the culture process?

RPMI

Mitogen; Reagent that stimulates mitosis

Phytohemagglutinin

Arrests mitosis at metaphase

Colcemid/Colchicine

Reagent used to digest proteins

Trypsin

T or F: In step 1, the cultured blood cells will be grown at 37 °C incubator for 3 days

T

In step 1, the cells must be in ____________ phase because splitting of a cell line 2 days before harvesting, and changing the medium 1 day before harvesting, stimulates cell proliferation significantly.

logarithmic

T or F: In step 2, addition of pre-cooled colcemid arrests the cell cycle at metaphase stage

F It should be pre-warmed

Incubation with colcemid should be done for ____mins

15

Optimal exposure time to colcemid requires a balance between

proliferative activity index of cells and concentration of colcemid

Short proliferative index requires a ______ _______ ________ of colcemid

short high burst (flash)

This refers to how fast or slow the cells grow

proliferative activity index of cells

In step 2, _____ ________ will be formed by centrifuging the tube at 1000 RPM for 10 mins

cell pellets

T or F: Using Actinomycin D or ethidium bromide (added before harvesting), or bromodeoxyuridine (BrdU) before colcemid treatment is just optional

T

can significantly increase the total yield of metaphase chromosomes

Cell synchronization

Cells are arrested at ___ phase by adding an excess amount of BrdU overnight (16 h).

S

In step 2, what is added after arresting the cells at S phase?

Thymidine (for 5.5 h before colcemid treatment)

T or F: In step 3, the cell suspension in hypotonic state will be centrifuged for 1200 RPM for 10 mins.

F Only for 5 mins

ratio of absolute methanol to glacial acetic acid

3:1

the most common method of staining chromosomes for differentiation

GTG-banding

Meaning of GTG in GTG-banding

G-bands by Trypsin using Giemsa

GTG-banding uses trypsin that digests the chromosomes at regions rich in basic amino acids, specifically ____ and ____

Arg; Lys

In step 5, slides that will be chosen for analysis and visualization must be:

Properly trypsinized chromosomes Clearly defined metaphase spreading

is a part of a chromosome which is clearly distinguishable from its adjacent segments by appearing darker or lighter with various banding methods (Paris conference, 1971)

Band

T or F: Classical cytogenetics provide low resolution

T

T or F: Changing the stain would not affect the banding pattern

F Changing the stain would also change the banding pattern

In 1958, _________ et al., published there first paper describing the use of quinacrine mustard to stain chromosome there by ushered in a new era of chromosome banding

Caspersson

the first attempt to provide nomenclature for chromosome banding in any species and thus its recommendations have been adopted to nonhuman species as well

Paris report (1971)

T or F: Studying banding patterns allows you to see smaller pieces of the chromosome, so that you could identify smaller structural chromosome abnormalities not visible on a routine analysis.

T

Classification of Banding Techniques is based on

GC and AT rich regions Constitutive Heterochromatin Region

Classification of Banding Techniques

Quinacrine 1958 by Casperson et al. Giemsa 1971 by Summer et al. NOR 1973 by Matsui & Sasaki Centromeric 1978 by Linde & Laursen

Giemsa stain reacts with

Sulfate structures

T or F: Sulfate structures are found in sugar and phosphate groups of DNA

F They are found in proteins

T or F: Trypsinization precedes staining

T

T or F: Darkness and lightness of Giemsa stain have nothing to do with the number of Hydrogen bonds between bases

T

Banding based on fluorescence

Q-banding

In G-banding, ___-rich regions stain dark due to abundance of proteins

AT

Quinacrine fluotescent dye stains ___-rich regions

AT

Banding pattern is opposite to G-banding

R-Banding

Type of banding that uses phosphate buffer

R-banding

C in C-banding stands for

Centromeric OR Constitutive

Usually stains the entire long arm of the Y chromosome

C-banding

T or F: Human chromosomes do not need a lot of protein encoding regimens

T

Type of bending wherein euchromatin stains dark and heterochromatin stains light

R-banding

Type of banding done by denaturation with alkaline (strong base) followed by Giemsa

C-banding

Type of banding that involves silver staining regions

NOR banding

NOR stands for

Nucleolar Organizing Regions

First stain used in Q banding

Quinacrine Mustard

In Q banding, stained chromosomes need to be subjected to ____ ________ for excitation

UV light

In Q banding, ____ regions quench dye out and do not fluorescence

GC

Type of banding used in study of chromosome heteromorphism

Q banding

T of F: Q banding does not fade during examination

F Its disadvantage is its tendency to fade

In G banding, interaction of the DNA with _______ & _______ components of stain brightens sulfur rich regions

thiazine; eosin

Type of banding used in identification of bands rich in sulfur content

G banding

Type of banding used in the identification of chromosomal abnormalities and gene mapping

G banding

T or F: G banding is also used in plants

F It is not used in plants

T or F: In C banding, repetitive DNA renature but unique DNA do not renature

T

T or F: C banding is used in paternity testing and gene mapping

T

Each chromosome arm is divided into regions, or _________ ________, that can be seen using a microscope and special stains.

cytogenetic bands

T or F: The cytogenetic bands are labeled p1, p2, q1, q2, etc., counting from the centromere out toward the telomeres.

T

T or F: Following ISCN, each area of chromosome has a given number.

T

Each human chromosome has a short arm and long arm, separated by a ___________

centromere