Chromosome Tracking using FISH and GISH

Modern cytogenetics employs molecular hybridization techniques to study the structure, function, and evolution of chromosomes at a high resolution.
Two important methods — Fluorescence In Situ Hybridization (FISH) and Genomic In Situ Hybridization (GISH) — enable visual tracking of specific DNA sequences or entire genomes directly on chromosomes using fluorescently labeled probes.

2. Fluorescence In Situ Hybridization (FISH)

Principle

FISH is based on the hybridization of fluorescently labeled DNA probes to their complementary DNA sequences on denatured chromosomal DNA.
When viewed under a fluorescence microscope, the hybridized regions emit light, marking the location of specific genes or sequences.

Steps in FISH Procedure

1. Preparation of Chromosome Spread: Metaphase chromosomes are fixed on a slide.
2. Denaturation: Both chromosomal DNA and probe DNA are denatured (to single strands).
3. Hybridization: The fluorescently labeled probe binds to its complementary sequence.
4. Washing: Removes unbound probes.
5. Detection: Visualization under a fluorescence microscope using filters matching fluorophore emission.

Types of FISH

• Single-color FISH: Detects one specific DNA sequence.
• Multicolor FISH (mFISH): Uses multiple fluorescent dyes to label several chromosomes simultaneously (useful in karyotyping).
• Chromosome painting: Whole chromosomes or regions are labeled to study translocations or rearrangements.

Applications of FISH

• Detection of chromosomal abnormalities (deletions, duplications, translocations).
• Gene mapping and marker localization.
• Cancer cytogenetics – detection of oncogene amplifications.
• Species identification and evolutionary relationships.
• Studying chromosomal rearrangements and structural variations.

3. Genomic In Situ Hybridization (GISH)

Principle

GISH is an extension of FISH, where total genomic DNA from one species is labeled and hybridized to chromosomes of another species.
It differentiates parental genomes in hybrid or polyploid organisms based on DNA sequence homology.

Procedure

1. Chromosome preparation from hybrid or polyploid plant material.
2. Labeling the total genomic DNA of one species with a fluorescent dye.
3. Hybridization with unlabeled competitor DNA (to block repetitive sequences).
4. Detection under a fluorescence microscope – labeled genome regions appear as distinct signals.

Applications of GISH

• Identification of parental genomes in hybrids and allopolyploids.
• Studying introgression (gene flow) between species.
• Chromosome pairing analysis in meiosis of hybrids.
• Phylogenetic and evolutionary studies in plants.
• Tracking genomic composition in breeding programs.

4. Comparison: FISH vs. GISH

Feature	FISH	GISH
Probe used	Specific DNA sequence	Total genomic DNA
Purpose	Locate genes or chromosomal regions	Differentiate genomes in hybrids/polyploids
Specificity	High (sequence-level)	Broader (genome-level)
Common use	Gene mapping, clinical diagnostics	Evolutionary, hybridization, plant cytogenetics
Blocking DNA	Not required	Required (to prevent non-specific binding)

5. Significance

FISH and GISH provide powerful molecular cytogenetic tools for:

• Visualizing genome organization directly on chromosomes,
• Detecting evolutionary rearrangements,
• Understanding hybrid genome composition, and
• Linking cytology with genomics in both plants and animals.

Chromosome tracking using FISH and GISH bridges the gap between classical karyology and molecular genetics.
FISH enables precise gene localization, while GISH distinguishes genomic origins in hybrids — together, they have revolutionized evolutionary cytogenetics and genome analysis.

🧬 Localization and Mapping of Genes or Genomic Segments

Gene localization and mapping are essential techniques in cytogenetics and genomics that determine the physical position and arrangement of genes or DNA segments on chromosomes.
This helps in understanding genome organization, inheritance patterns, linkage relationships, and in identifying genes responsible for genetic traits or diseases.

2. Types of Gene Mapping

Type	Basis	Description / Use
Genetic (Linkage) Mapping	Recombination frequency	Determines relative positions of genes on a chromosome using genetic crosses.
Physical Mapping	DNA sequence distance	Uses molecular methods to measure actual physical distance (in base pairs) between genes.
Cytogenetic Mapping	Chromosomal landmarks	Locates genes on specific chromosomal bands or regions using microscopy-based techniques.

3. Techniques for Localization and Mapping

A. Classical Cytogenetic Techniques

1. Chromosome Banding (G-, R-, and C-banding):
   Identifies chromosomal regions and structural changes linked with specific genes.
2. Deletion Mapping:
   Correlates the loss of chromosomal segments with loss of function of specific genes.

B. Molecular Cytogenetic Techniques

1. FISH (Fluorescence In Situ Hybridization):

• Uses fluorescently labeled DNA probes to detect the precise chromosomal location of a gene or DNA sequence.
• Enables gene mapping, translocation detection, and genome comparison.
• Multicolor FISH (mFISH) and chromosome painting can localize multiple genes simultaneously.

2. GISH (Genomic In Situ Hybridization):

• Uses total genomic DNA as a probe to distinguish genomic segments or parental origins in hybrids and polyploids.
• Useful for evolutionary and comparative genomics.

C. Molecular Mapping Techniques

1. Restriction Fragment Length Polymorphism (RFLP):

• Detects DNA sequence variations using restriction enzymes and hybridization probes.
• Used for constructing early linkage maps in plants and animals.

2. Microsatellite (SSR) and SNP Markers:

• Highly polymorphic DNA markers used for fine-scale mapping and marker-assisted selection.

3. Genome Sequencing and Bioinformatics:

• High-throughput sequencing allows whole-genome mapping and annotation of gene locations in base-pair resolution.

4. Chromosome Landmarks Used in Mapping

• Centromere: Used as a positional reference for mapping arms (p and q).
• Telomere: Marks terminal regions of chromosomes.
• Banding Patterns: Serve as cytological coordinates for mapping loci.
• Nucleolar Organizer Regions (NORs): Identify genes for rRNA synthesis.

5. Applications of Gene Localization and Mapping

1. Identification of disease genes and chromosomal abnormalities.
2. Comparative genomics — tracing evolutionary relationships among species.
3. Marker-assisted breeding and QTL (Quantitative Trait Loci) mapping in plants.
4. Construction of genome maps for sequencing and annotation projects.
5. Detection of chromosomal rearrangements (duplications, translocations, deletions).

Localization and mapping of genes integrate cytogenetic, molecular, and computational approaches to reveal the organization and evolution of genomes.
Through techniques like FISH, GISH, and molecular marker analysis, scientists can visualize, locate, and compare genes, providing a foundation for genetic improvement, medical diagnostics, and evolutionary studies.

https://biologywala.com/chromosome-banding-patterns

Join SACHIN’S BIOLOGY on Instagram or Facebook to receive timely updates and important notes about exams directly on your mobile device. Connect with Mr. Sachin Chavan, the founder of Sachin’s Biology and author of biologywala.com, who holds an M.Sc., NET JRF (AIR 21), and GATE qualifications. With SACHIN’S BIOLOGY, you can have a direct conversation with a knowledgeable and experienced professional in the field of biology. Don’t miss out on this opportunity to enhance your exam preparation!