5: Molecular Diagnostics for Forensics and Genetics (FINALS) Flashcards by Ma. Clariza Luna

In molecular biology, this refers to the study of the human chromosome

Genetic

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In genetics, this refers to the process of examining complete set of cells in an organism

Karyotyping

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In genetics, what are the characteristics of chromosomes studied

idk but list it ghorl

physical size
structure
main behavior

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Most common application of molecular techniques in genetics

to identify a CHANGE in chromosomes

or specific mutations in the chromosome

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Enumerate the traditional process in studying genes/chromosomes

dapat alam mo na to malaki ka na

Cell culture
Metaphase arrest
Hypotonic swelling
Fixation
Chromosome Spread
Aging
Photomicroscopy
Reporting

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Review

Identify the reagent used in cell culture

Phytohemagglutinin (PHA)

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Review

Reagent/inhibitor used to arrest cell division at metaphase

Colcemid

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Review

Reagent for hypotonic swelling

Hypotonic buffer

eme dapat 0.075M KCl at 37° C for 10-20 mins

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Reagent for Fixation

Carnoy’s fixative

(3:1 ; absolute methanol: glacial acetic acid)

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In terms of the manner of reporting, what parameters are considered?

pairing of the chromosomes
ba(n)ding patterns

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a new technique under the
molecular biology used to match chromosomes

In Situ Hybridization (ISH)

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In its earliest development, ISH used ________ probes as target labels

radiolabeled (radioactive substances)

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due to the harmful effects of radiolabeled probes what was the alternative probe used?

Fluorescence probes

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Type of ISH which uses flurorescence probes

Fluorescence In Situ Hybridization

sis kung mali ka ewan nalang

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➢ a tagging technique
➢ A method on how we add the probe into the
sample or reagent
➢ It is a process wherein the DNA polymerase I is used to replace some nucleotides of
DNA sequences

Nick Translation

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STUDY MO GENERAL PROCEDURE NG FISH

shoo shoo

congrats kung binasa mo, kung hindi magbasa ka na

Nick translation -> denaturation -> required antibodies (labeled) -> visualization (microscope)

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Main advantage of FISH

➢ specifically see the exact location of the gene
➢ less hazardous (compared to using radiolabeled probes

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Enumerate the advanced applications of FISH

INTERPHASE FISH
WCP (Whole Chromosome Painting)
MFISH (Multicolor – FISH)
SKY (Spectral Karyotyping)
ALL – TELOMERE – FISH
Comparative Genomic Hybridization

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Advanced Application of FISH

not arrested in the metaphase stage; instead, what is checked here is the interphase stage.

Interphase FISH

basic btch

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Adv. Appli of FISH

Advantages of Interphase FISH

NO cell culture required, thus
the diagnosis is much rapid
You will proceed immediately to
hybridization (in situ technique is directly
performed)

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Adv. Application of FISH

In this application, different or many types of
probes are used from a SINGLE chromosome

WCP (Whole Chromosome Painting)

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Adv. Application of FISH

Main principle of WCP

On the single chromosome, there are
LOTS of UNIQUE DNA associated with
it.

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Adv. Application of FISH

Main advantage of WCP

for seeing or looking for any rearrangements (i.e.,
interchanged long and short arms) in the
chromosome

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Adv. Application of FISH

Almost the same as WCP but the main difference is that there is a system/software installed

Multicolor FISH (MFISH)

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# Adv. Application of FISH Main advantages of MFISH

* automated (has computer-assisted system) * Fixed profile, which you'll only need to compare to the sample

# Adv. Application of FISH * permits the display and identification of all 24 human chromosomes using painting probes * multiple chromosomal aberrations are detected

Spectral Karyotyping | definition from taken fr both lec and cyto

# Adv. Application of FISH Main advantage of SKY

allows the observation of multiple chromosomal aberratios or changes

# Adv. Application of FISH The main probe/label is a telomere probe

ALL - Telomere FISH

# Adv. Application of FISH Detects six base repeats present at the ends of all chromosome

ALL - Telomere FISH

# Adv. Application of FISH Main advantage of All-Telomere FISH

used as a confirmatory assay (presence/absence of telomeric regions)

# Adv. Application of FISH you simply compare the DNA content of a differentially labeled normal and abnormal cell population

Comparative Genomic Hybridization

Enumerate the 3 Main Applications of FISH

1. DIAGNOSIS 2. PROGNOSIS 3. EVALUATE | 2 sis ate

# 3 Main Appli. of FISH Refers to identifying some of the inherited diseases or those associated with mutation

Diagnosis

# 3 Main Appli. of FISH pertains to certain conditions whether good or bad; or whether there is a presence of a mutated gene

Prognosis

# 3 Main Appli. of FISH Refers to asssesing possible remission of a certain disease | Q asked: Is it possible that a specific condition may recur?

Evaluate

TOF. Anything associated with genetics or genes are applicable for FISH

True

IDK beh study the advantages of FISH ig

1. Much more specific 2. Distinguish one disease from another 3. Detect diseased cells more easily and rapidly, without requiring living cells (since p’wede na yung fixed sa culture) 4. Can be quantitated through automated methods 1. Less labor intensive, which will give a much more rapid time for the turnaround time.

# Match the type of Specimen according to purpose + handling and transport Heparinized peripheral Blood

Purpose: Routine; adults and children H&T: Room temp

# Match the type of Specimen according to purpose + handling and transport Bone marrow aspirates

Purpose: Hematologic disorders H&T: Room temp

# Match the type of Specimen according to purpose + handling and transport Skin biopsies

Purpose: Fibroblast cultures H&T: Ice

# Match the type of Specimen according to purpose + handling and transport Tissues (Kidney, Liver, Muscle, and Lung)

Purpose: obtained after death or fetal loss (not routine) H&T: Ice

# Match the type of Specimen according to purpose + handling and transport Amniotic Fluid

purpose:Prenatal Analysis H&T:Room temp

# Match the type of Specimen according to purpose + handling and transport Chorionic villus

Purpose: prenatal analysis H&T: room temp

# Match the type of Specimen according to purpose + handling and transport Fetal Blood from umbilical cord

Purpose: Rapid karyotyping H&T: room temp

Done so as to detect congenital inherited or genetic diseases or disorders

Prenatal analysis

One of the most powerful tools in the fight against crime and other legal matters is the DNA testing.

Forensic Science

First application that comes to mind with regards to forensic science

Identifying criminal suspects

Familiarize the old methods associated with identifying criminal suspects (old methods)

1. Hair 2. Bone 3. Fingerprints 4. Blood

# Identify the specimen used in identifying criminal suspects (old method) - **common specimen** - gives an indication of age, sex, and race - comparison using this specimen allows police to identify people, with an error rate of 1 in 4500

HAIR

# Identify the specimen used in identifying criminal suspects (old method) - **common specimen** - age, sex race, and sometimes medical history can be determined - **obtained from a disaster site**

Bone

# Identify the specimen used in identifying criminal suspects (old method) - **most common** - introduced in the late 1800s - main principle lies in the fact that this specimen is unique in each indivduals

Fingerprints

Disadvantage of fingerprint as an evidence in identifying criminal suspects

easy to remove | wear gloves kung papatay

# Identify the specimen used in identifying criminal suspects (old method) - **considered as the best sample** - 4 major groups were discovered in the 1900s

Blood

TOF. Blood typing is a powerful tool for resolving criminal matters

F. legal po

Disadvantages of Blood samples in identifying criminal suspects

More blood groups have been discovered over the past 100 years, but even these cannot provide the same level of identification as DNA testing. | tl;dr: ABO system is not the only existing blood group

TOF. In forensics, blood typing is the best method in identification

F. dna testing

Major technique employed in identifying criminal suspects

Molecular Techniques

TOF. Current DNA technology allows any individuals to be distinguished from all other individuals.

Eurt

Enumerate samples from which we can isolate and extract DNA

- blood - semen - saliva - bone - teeth - microscoping flecks of skin - other tishoes

TOF. DNA can be found even in inanimate or non-living objects

T | laway on a table kc madaldal ka, hello?

Study mo tong list kung d mo pa know ang objects where DNA may be found

1. Chewing Gum 2. Stamps and Envelopes 3. Penile Swabs 4. Washed Stains 5. Door Knobs 6. Tooth Brushes 7. Sanitary Pads 8. Tooth Pulp 9. Sweaty Clothing 10. Telephones 11. Bone Marrow 12. Hair Brushes

Main challenge in DNA testing

different samples may contain different concentrations of DNA | some samples have a lot of DNA some may konti ## Footnote pa pcr mo nalang (- *a concerned mls student*)

disadvantage of using objects where DNA may be found for forensics

very low concentration of DNA obtained ## Footnote pa pcr mo ulit

study mo dna content of biological samples picture, yes now na

ayaw mo? edi wag!

The highest amount of DNA can be obtained in?

Male semen | yuck boyz ## Footnote 250,000 ng/mL

best specimen if one wants to investigate crime associated with sexual assault

Semen | kasalanan to ng mga lalaki

TOF. blood samples contain a minimal amount of DNA

F (high amounts; 30,000 ng/mL)

TOF. DNA can also be obtained in hair samples | tru

True | sabi sayo eh (lower amounts; need plucked from the hair root) ## Footnote pa pcr mo ulit, skill issue

DNA CONTENT NG SALIVA DALI

5,000 ng/mL

Best method for paternity and maternity testing

DNA testing

can help answer questions about biological descendants of a particular family (ancestral lineage) | baka si lord to

DNA Analysis

Familiarize other uses of DNA for forensic identification

1. Identify organ donors with recipients in transplant programs 2. Identify human migration patterns 3. Use of DNA forensic science in military matters (victims of war crimes, war, plane crashes, etc.)

# Recall Human cells contain how many **pairs** of chromosomes?

working portions in our chromosomes which we refer to as

genes

short sequences of bases that repeats many times.

Short Tander Repeats (STRs)

- theoretically, more than 30% of the human genomes consist of these - Satellite DNA - acts as fillers or spaces between coding portions of the chromosome

Short Tandem Repeats (STRs)

Among all satellite DNAs, the most commonly used is?

chromosome 11 (C11)

How will you know if a certain sequence is a Satellite DNA?

As long as there are two repeating sequences | take note: 2-5 repeating sequences are called micro-satellite DNA ## Footnote mini-satellite: 9-10 repeating sequences

Main advantage of micro-satellite DNA

can survive degraded DNA

TOF: The length of the repeats (STRs) is inherited and differs from person to person

True