gene mapping

GENE MAPPING

  • Definition: Method used to identify the locus of a gene and the distance between genes.

  • Gene: A unit of heredity from parent to offspring influencing characteristics.

  • Genome: Complete set of genes or genetic material in an organism.

  • Purpose of Mapping: Identifies which chromosomes contain specific genes and their precise locations.

TYPES OF GENOME MAPPING

  • Gene mapping

  • Physical mapping

  • Genetic mapping

  • Restriction mapping

  • Fluorescent in Situ Hybridization (FISH)

  • Sequence Tagged Sites (STS) mapping

  • Fingerprint mapping

  • Optical mapping

GENETIC MAPPING

  • Process: Determines the order and relative distances between genetic markers on a chromosome.

  • Representation: Linkage of genes can be illustrated through genetic maps or linkage maps.

  • Historical Context: First chromosomal map created by Alfred Sturtevant in Drosophila using gene recombination frequencies.

RECOMBINATION FREQUENCY

  • Formula:

    • Recombination Frequency = (Total No. Recombinants) x 100 / (Total No. Progenies)

  • Interpretation: Genes with recombination frequency less than 50% are linked and on the same chromosome.

FEATURES OF GENETIC RECOMBINATION

  • Determines gene order and distances measured in map units (1 map unit = 1 cM).

  • Uses recombination frequencies to assess relative distances between alleles (inversely proportional).

MARKERS

  • Definition: Genes or DNA with known locations on chromosomes for identification or relationship determination.

  • Types of Genetic Markers:

    • Gene markers

    • DNA markers:

      1. RELPs (Restriction Fragment Length Polymorphism)

      2. SNPs (Single Nucleotide Polymorphism)

      3. AELPs (Amplified Fragment Length Polymorphism)

      4. RAPD (Random Amplification of Polymorphic DNA)

      5. SSLPs (Simple Sequence Length Polymorphisms)

      6. Microsatellites/SSRs (Simple Sequence Repeats)

      7. Minisatellites/VNTRs (Variable Number of Tandem Repeats)

GENETIC MAPPING TECHNIQUES

  • Constructs a map showing gene positions and features using genetic techniques.

  • Includes cross-breeding and pedigree analysis; quality depends on genetic markers and mapping population size.

STEPS INVOLVED IN GENETIC MAPPING

  1. Determination of linkage groups (Chromosome count)

  2. Determination of map distance (two-point and three-point test crosses)

  3. Determination of gene order

  4. Combining map segments

SAMPLE USAGE IN GENE MAPPING

  • Common Sample: Saliva.

  • Process: Isolate DNA and identify unique patterns (polymorphisms) distinguishing affected from non-affected individuals.

PHYSICAL MAPPING

  • Provides actual DNA base pair distances with high accuracy.

  • Supplies nucleotide numbers and physical distances between genetic markers.

TECHNIQUES IN PHYSICAL MAPPING

  • Linkage analysis

  • In situ hybridization

  • PFGE (Pulsed Field Gel Electrophoresis)

  • Molecular markers

RESTRICTION MAPPING

  • Defines relative positions of DNA recognition sequences for restriction enzymes.

  • Build by comparing fragment sizes upon restriction enzyme digestion.

FLUORESCENT IN SITU HYBRIDIZATION (FISH)

  • Allows visualization of marker positions on chromosomes via fluorescent probes.

  • In optical mapping, markers noted by gaps in DNA fibers.

SEQUENCE TAGGED SITES (STS)

  • Powerful physical mapping technique utilizing short, recognizable DNA sequences (100bp - 500bp) that occur once per chromosome or genome.

MAPPING OF STS

  • Needs overlapping DNA fragments for creating maps from STS data, which can be obtained through hybridization techniques or PCR.

CROSSING OVER AND GENE MAPPING

  • Exchange of genes occurs between homologous chromosomes.

  • Crossing events outside gene regions do not alter their arrangement.

  • Double crossovers distinguish gene order.

GENETIC MAP VS PHYSICAL MAP

  • Genetic Map: Constructed from recombination frequencies (indirect) with units in cM.

  • Physical Map: Locates DNA sequence positions (direct) with units in base pairs.

IMPORTANCE OF GENE MAPPING

  • Anatomy of the human genome for understanding genetic diseases.

  • Develops methods for gene therapy, providing clinically useful linkage information.

GENE SEQUENCING

  • Definition: Process to determine the nucleotide order in DNA.

  • Essential in biological research and applied fields (medical diagnosis, biotechnology).

METHODS OF GENE SEQUENCING

  1. Basic DNA Sequencing

  • Maxam-Gilbert Method

  • Sanger Method

  1. Advanced DNA Sequencing

  • Shotgun Sequencing

  1. Next Generation Sequencing

  • SOLID Sequencing

  • Illumina Sequencing

  • Pyrosequencing

ADVANTAGES AND DISADVANTAGES OF SEQUENCING

  • Advantages: Identifies genome sequences and genes causing diseases.

  • Disadvantages: Concerns over privacy with genetic data (e.g., police databases).

SHOTGUN SEQUENCING

  • Randomly cuts DNA segments into manageable pieces for sequencing and reassembly using overlapping sequences.

PYROSEQUENCING

  • Based on "sequencing by synthesis," detecting pyrophosphate release during nucleotide incorporation.

  • Employs enzyme activity for detection, yielding real-time sequencing data.

APPLICATIONS OF GENE SEQUENCING

  1. Sequencing individual genes, clusters, chromosomes, or entire genomes.

  2. Determining disease causes through DNA sequences.

  3. Significant in research and forensic science.

  4. Evaluation of sequencing for accuracy and reliability.

MOLECULAR BIOLOGY AND EVOLUTIONARY STUDIES

  • Molecular Biology: Sequencing informs on genome changes and disease associations.

  • Evolutionary Biology: Analyzes relationships and evolution through DNA sequence comparisons.

REFERENCES

  • Dr. M Vasudevan, S Sreekumari, Textbook of Biochemistry, 3rd Ed, Jaypee Brothers Medical Publishers.

  • Dr. U. Satyanarayanan, Dr. U. Chakrapani, Biochemistry, 3rd Ed.