Brainscape's Knowledge GenomeTM


Top Flashcards (Certified)
All Flashcards

Year 2 EMS GiM > Cytogenetics - Techniques of Analysis > Flashcards

Cytogenetics - Techniques of Analysis Flashcards

1
Q

What techniques are used to sequence the whole genome? (3)

A
  • G banding
  • Next generation sequencing
  • Microarrays
How well did you know this?
1
Not at all
2
3
4
5
Perfectly
2
Q

What are the main targeted tests? (3)

A
  • FISH
  • MLPA
  • QF-PCR / qPCR
How well did you know this?
1
Not at all
2
3
4
5
Perfectly
3
Q

What is the purpose of G-banding?

A

Stains condensed chromosomes&raquo_space; visible karyotype.

-heterochromatic (AT-rich) stains dark

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
4
Q

What stains are used for G-banding? (2)

A
  • Trypsin and Leishman’s

- Giemsa stain

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
5
Q

What stage of the cell cycle are cells paused at for G-banding?

A

Metaphase.

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
6
Q

What is the general process of FISH?

A 
Probe
>> fluorescent label
>> denatured
>> hybridisation
>> visualisation (UV).
How well did you know this?
1
Not at all
2
3
4
5
Perfectly
7
Q

What types of probes are used for FISH? (3)

A
  • UNIQUE SEQUENCE (» highlight small region, e.g. dup)
  • CENTROMERIC (» no. of copies of each chromosome e.g. trisomy)
  • PAINTS (» each chromosome different colour e.g. translocation)
How well did you know this?
1
Not at all
2
3
4
5
Perfectly
8
Q

What is FISH used for? (5)

A
  • Copy number imbalance
  • Aneuploidy
  • Confirm G-banding
  • Confirm array CGH
  • Identify specific abnormalities in cancer
How well did you know this?
1
Not at all
2
3
4
5
Perfectly
9
Q

What is copy number variation?

A

DNA segment with variable copy number to reference genome.

  • 12% human genome
  • benign / malignant
How well did you know this?
1
Not at all
2
3
4
5
Perfectly
10
Q

What is a high copy number of CCL3L1 associated with?

A

Decreased susceptibility to HIV.

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
11
Q

What is a low copy number of FCGR3B associated with?

A

Increased susceptibility to inflammatory autoimmune disorders.

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
12
Q

What cytogenetic methods are used to assess copy number?

A
  • FISH
  • MLPA
  • Microarray CGH
  • Next generation sequencing
  • QF-PCR
  • qPCR
How well did you know this?
1
Not at all
2
3
4
5
Perfectly
13
Q

What is Multiplex Ligation-dependent Probe Amplification (MLPA)?

A

Multiplex PCR&raquo_space; detects abnormal copy numbers in up to 50 genomic locations.
-alternative to FISH

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
14
Q

MLPA analysis - what peak areas are normal / duplication / deletion?

A 
Normal = 1.0
Duplication = 1.35
Deletion = 0.65
How well did you know this?
1
Not at all
2
3
4
5
Perfectly
15
Q

What does microarray CGH analyse?

A

Analyses copy number variants (genomic imbalances) compared to reference.

  • highlights imbalances
  • can analyse 1000s of oligonulceotides at a time
How well did you know this?
1
Not at all
2
3
4
5
Perfectly
16
Q

What is replacing karyotyping as a 1st line test?

A

Microarray CGH.

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
17
Q

What samples are required for microarray CGH? (2)

A
  • 3ml blood in EDTA

- Control DNA (same sex)

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
18
Q

How many oligonulceotides are required to determine copy number change in mircroarray CGH?

A

3 oligonucleotides.

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
19
Q

How many genomic disorders can be identified using microarray CGH?

A

> 40 genomic disorders.

-including learning disabilities and dysmorphic infants

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
20
Q

What are the main advantages of microarray CGH? (3)

A
  • Early diagnosis (1st line)
  • High resolution
  • Greater accuracy of imbalance location / size
How well did you know this?
1
Not at all
2
3
4
5
Perfectly
21
Q

What are the main disadvantages of mircoarray CGH? (4)

A
  • Dosage changes only (not balanced rearrangements)
  • Low level mosaics not detected
  • Non-pathogenic changes detected
  • Needs good quality DNA
How well did you know this?
1
Not at all
2
3
4
5
Perfectly
22
Q

What is next generation sequencing (NGS) used for?

A

Generating sequencing libraries by fragmenting genomic DNA.

-divide genome and compare dosage differences to reference DNA

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
23
Q

How many base pairs do NGS beads contain?

A

250 bp.

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
24
Q

What does an increased test:control ratio indicate?

A

Gain.

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
25
What does an decreased test:control ratio indicate?
Loss.
26
What is quantitative fluorescent PCR (QF-PCR)?
PCR amplification of short tandem repeats using fluorescent primers. -products quantified as peak areas >> compared to reference
27
What is prenatal aneuploidy detection carried out on?
DNA extracted from amniotic fluid / chorionic villi.
28
What primers are used for prenatal aneuploidy detection?
Primers from chromosomes 13, 18, 21, X and Y.
29
What happens in prenatal aneuploidy detection when aneuploidy is present?
2 markers with abnormal dosage. | -size of product directly related to ATTT repeats
30
QF-PCR; what is a peak area ratio when trisomy is present?
Either 2:1 or 1:1:1.
31
What is qPCR?
Quantitative / real time PCR. | -confirms small copy number variations
32
What is qPCR used?
When FISH is unsuitable.
33
qPCR; what is relative quantitation (RQ)?
Compares difference in concentration between patient sample and normal control. - 2 different primer sets used - expressed as a ratio relative to 1
34
What are the RQ values for normal DNA, deletion and duplication (qPCR)?
Normal - 1.0 Deletion - 1.5 Duplication - 0.5
35
Which cytogenetic analyses take the least amount of time?
MLPA / QF-PCR. | -10-30 mins per case
36
Which cytogenetic analysis takes the most amount of time?
G-banding. | -30 mins - 4 hours per case
37
What are the timescales for the main cytogenetic analyses?
``` G-banding : 30min - 4hr FISH : 10 min - 1 hr Microarrays : 10 min - 2hr NGS/qPCR : 30 mins - 2hr MLPA/QF-PCR : 10-30 mins ```
38
What are the main pre-natal sample types, and when are they carried out? (3)
- Amniocentesis (16w) - Chorionic villus (12w) - Non-invasive prenatal testing (12w)
39
What type of blood sample is used for G-banding?
2-5 ml unclotted. - T cells stimulated - Culture 2-3 days
40
What are the genetic causes of Down syndrome? (3)
- Trisomy 21 (90-95 %) - Robertsonian Translocation - Mosaic trisomy 21
41
What is the recurrence risk of Down syndrome due to trisomy 21?
~1%. | -less if post-zygotic
42
What is the recurrence risk of Down syndrome due to Robertsonian translocation?
Both parents - 100%. Maternal - 12%. Paternal - 2%. -less in paternal; preselection prevents unbalanced sperm >> egg
43
What is Robertsonian translocation?
Translocation in an acrocentric chromosome. - 13, 14, 15, 21, 22 - normally t(14;21) in Down
44
What testing is done to clarify imbalances?
- Array CGH to analyse imbalance - check if OMIM morbid genes - FISH to confirm
45
What are OMIM morbid genes?
Online Mendelian Inheritance in Man. | -lists relationship between genes and phenotypes
46
What proportion of recurrent miscarriages are due to balanced chromosome changes?
2-3%.
47
What proportion of infertile men have a chromosome change?
10%. | -usually Klinefelter (47, XXY) or Y deletion
48
What chromosome change causes female infertility?
Turner syndrome (45, X).
49
How can t(4;11) cause recurrent miscarriages?
Try to pair the wrong chromosomes.
50
What is the Pachytene cross?
3rd stage of prophase of meiosis. - chromosomes divide into 4 chromatids - 50% chance of unbalanced gamete if translocation
51
What are the main methods of non-invasive prenatal testing (NIPT)?
- Maternal blood | - Extract free foetal DNA
52
What is non-invasive prenatal testing used to assess?
Aneuploidy. | -invasive test to confirm
53
What are the main indicators for prenatal testing? (4)
- Increased maternal age - Serum screen risk - Abnormal ultrasound - FH
54
What proportion of Down's births involve mothers >35 years?
40%.
55
What are the results of a prenatal serum screen (triple test) if the child has Down syndrome?
- Decreased alpha-fp - Increased oestriol - Increased hcg
56
How is amniotic fluid / chorionic villi tested?
DNA extraction >> QF-PCR >> culture cells if abnormal (7-14days) >> G-banding
57
What proportion of trisomy 21 foetuses spontaneously abort?
25%. | -post 16 weeks
58
What proportion of spontaneous abortions have a chromosome abnormality, and what tests are carried out?
50%. - G-banding / QF-PCR / MLPA (aneuploidy) - array CGH (dosage)
59
What cytogenetic analysis is carried out for leukaemia?
1ml unclotted bone marow >> culture overnight >> G-banding / FISH
60
What chromosomal abnormality is associated with leukaemia (CML)?
Philadelphia chromosome. - t(9;22) - bcr/abl fusion gene
61
What cytogenetic analsysis is carried out on solid tumours?
- Fresh tumour >> FISH / G-banding | - Archived tissue (paraffin) >> FISH / genotyping
62
What gene is involved in neuroblastoma?
MYCN gene.

Year 2 EMS GiM (12 decks)

Brainscape helps you reach your goals faster, through stronger study habits.
© 2025 Bold Learning Solutions. Terms and Conditions