Advanced Techniques In Molecular Cytogenetics. (Karyotyping, FISH, CGH, SKY)

What is cytogenetic........? 

 

Genetics -  The branch of science which deal with the study of heredity and the variation is called as genetics. 

Cytology - The branch of science which deal with the study of structure and function of cell and cell organelle. 

Cytogenetic :- The branch of genetics which deal with the study of cell and structure and function of the chromosome is called cytogenetic. 

Molecular Cytogenetics:-

 It involves the combination of molecular biology and cytogenetic. 

In general this involves the use of a series of technique in which DNA probe are labelled with different coloured fluorescent tag to visualize one or more specific region of the genome. 

Which are the Molecular Cytogenetics technique........? 

1) karyotyping.

2) Fluroscent in situ hybridization (FISH) . 

3) Comparative Genome Hybridization (CGH) 

4) Spectral karyotyping (SKY) 

1) Karyotyping :- 

The study of whole set of chromosome is sometimes known as karyology. 

The chromosome are depicted in a standard format known as a karyogram, or idiogram. 

Karyotyping  is a test to examine chromosome in a sample of cell this test can help identify genetics problem as the cause of disorder or disease. 

How to do karyotyping.....? 

Application of karyotyping.... 

- karyotyping show the chromosomal makeup of an individual. 

- Knowing the number of chromosome is essential for identifing chomosomal variations that cause genetics disorder. 

- Correct number of chromosome and chromosomal abnormality in numbers can be detect. 

- Correct size and shape of chromosome is visualize in this technique. 

- We can identify gender of unknown sample by this technique. 

2) Fluroscent in situ hybridization (FISH) 

What is FISH :-

              It is cytogenetic technique that uses fluroscent probe that bind to only those parts of the chromosome with a high degree of sequence complementary. 

- It is used to detect and localize the presence or absence of specific DNA sequence on chromosome. 

- FISH id often used for finding specific features in DNA for use in genetics counseling, medicine, and species identification. 

Step of FISH -

 

1) Denaturation - Conversation of double stranded DNA 

2) Hybridization - Application of probe DNA to slide and overnight incubation at 37°c 

Binding of probe DNA to target DNA. 

3) Post hybridization washing and detection -

 Washing of unbound probe DNA. 

Application of counter stain. 

4) Counter stain - Application of counter stain. 

5) Visualization - Visualization using fluorescence Microscopy. 

Type of FISH - 

 

1) Interphase FISH 

2) Telometric FISH 

3) RNA in Situ hybridization

4) prime in Situ labelling. 

5) Cobra FISH 

6) SKY FISH 

7) M- FISH

8) Fibre FISH. 

Application:-

 • FISH generally used compliment classical staining technique. 

• Subtle for chomosome identification at metaphase 

• useful in several clinical setting to determine prognosis

• Discrete information is obtained from each cell 

• FISH is powerful technique use in detection of chromosomal abnormality. 

3)  Comparative genomic hybridization (CGH) 

- It is molecular Cytogenetic method for analyzing copy number variations with the help of hybridization technique. 

Step :-

1) Metaphase slide preparation 

2) Isolation of DNA from test tissue and reference tissue

3) Selection and application of probe. 

4) Hybridization 

5) Fluorescence Dectecter image analysis 

6) Software analysis. 

Application of CGH -

1) It is used as a tool for investigation of genomic alterations in cancer . 

 2) It allows high resolution evolution of DNA copy number alterations. 

3) It increase  sophisticated diagnostic capabilities. 

4) Screening for prenatal gene mutation in fetas. 

5) It detect balanced rearrangement of chromosome. 

4) Spectral karyotyping (SKY) 

It is a cytogenetical technique used to simultaneously visualize all the pair of chromosome in. An. Organisms in different colors. 

- This technique is used for identification of chromosomal abnormality. 

- We can arrange the chromosome according to their number just by visualization of different colors aquire by the chromosome. 

How to do SKY.....? 

 

Application of SKY :-

  

1) This technique allow the isolation of structural abnormality. 

2) It allow to determine of the precise molecular address of chromosome break point associated with deletion translocation and insertion. 

3) SKY can discern the aberration that can't be detect very well by conventional bonding technique. 

4) It allows visualization of chromosome in different colors on same platform which is very easy to detect chromosomal abnormality.