Lesson 14: Genetic analysis techniques

What is FISH and what does it detect?

FISH is a cytogenetic technique using fluorescent DNA probes (labelled with a fluorophore) to detect specific DNA sequences, aneuploidies, and structural chromosome abnormalities in metaphase or interphase cells.

What types of probes are used in FISH?

• Locus-specific probes

• Centromeric probes

• Telomeric probes

• Whole-chromosome probes (spectral karyotype)

What is hybridization in genetics?

Hybridization is the specific binding of two complementary nucleic acid strands (DNA–DNA or DNA–RNA) after denaturation, based on base pairing (A–T, G–C). It is used to detect specific DNA or RNA sequences in techniques such as PCR, FISH, and microarrays.

What is PCR?

It is a Hybridization technique which allows the amplification of a DNA.

It is done in thermocyclers which constantly switch between 3 temperatures.

What are the components of PCR?

-DNA to amplify

-Primers: Indicate region for the synthesis of ne chain

-specific buffer, that neutralizes things that can harm the dna

-dNTPs: deoxy, nucelotide, triphosphate

-Thermostable DNA polymerase enzyme: taq polymerase

-Water

What are the three steps of PCR?

1. Denaturation: separation of the two strands of DNA from the sample at 94-96 degrees

2. Annealing: Hybridization of DNA strands with primers at 68 degrees

3. Polymerization: primers are extended by adding different free nucleotides in the order that is dictated by the nucleotide sequence of the chain that we use as a template at 72 degrees

RT-PCR

Use of mRNA as a template.

Retrotranscription is required to obtain complementary DNA

QPCR or Real-time PCR

technique in which DNA amplification and detection occur simultaneously. Using fluorescence, it allows real-time quantification of DNA, since the fluorescence signal is proportional to the amount of DNA produced.

What is PCR used for in dentistry

-detect periodontal bacteria

-detetct genetic predispositions for periodontitis

-for more complex cases to diagnose

What are the Omics?

Genomics (System biology), Transcriptomic (all messengers of body), Proteomics (Proteins), Metabolomics (all chemical context sugars, fats…) Pharmacogenomics (study of genes involved), Toxicogenomics (reaction to specific molecules)

What is genomics?

Techniques that make it possible to study the genome. Genomics is a field of molecular biology. A Genome is a complete set of DNA within a single cell of an organism, and as such, genomics focuses on the structure, function, evolution and mapping of genomes.

What is the main technique of genomics?

DNA sequencing

DNA sequencing

Can detect all the variation that may exist in a fragment of the genome

Any region can be analyzed

Sanger chain termination method