Skip to main content

Technique for detection and isolation of somaclonal variants, Factors affecting somoclonal variation


SOMACLONAL Variation


Detection & Isolation of Somaclonal Variants
Factors Affecting Somaclonal Variation


INTRODUCTION


Somaclonal variation refers to genetic and phenotypic variations observed among plants regenerated from in vitro cultured somatic tissues. These variations arise due to chromosomal rearrangements, gene mutations, epigenetic changes, and transposable element activation during tissue culture. Somaclonal variation can be undesirable in clonal propagation or beneficial for crop improvement.


I. TECHNIQUES FOR DETECTION AND ISOLATION OF SOMACLONAL VARIANTS


Detection involves identifying genetic or phenotypic differences between mother plants and regenerated plants, while isolation refers to selecting and stabilizing useful variants.

A. MORPHOLOGICAL AND PHYSIOLOGICAL METHODS


1. Morphological Evaluation
Visual screening of regenerated plants.
Traits observed:
Plant height
Leaf shape, size, and colour
Flower colour and structure
Fruit size, shape, and yield
Advantage: Simple and low cost
Limitation: Influenced by environment


2. Growth and Developmental Analysis
Variation in:
Growth rate
Time to flowering
Maturity period
Useful for detecting early or late maturing variants

B. CYTOLOGICAL METHODS


3. Chromosome Analysis (Karyotyping)
Detection of:
Aneuploidy
Polyploidy
Chromosomal deletions and translocations
Technique:
Root tip squash
Mitotic and meiotic analysis


4. Flow Cytometry
Rapid estimation of:
Nuclear DNA content
Ploidy level
Widely used in sugarcane, banana, potato


C. BIOCHEMICAL AND PHYSIOLOGICAL MARKERS


5. Isozyme Analysis
Electrophoretic separation of enzyme variants
Common enzymes:
Peroxidase
Esterase
Malate dehydrogenase
Detects gene expression changes
Limitation: Low genome coverage


6. Secondary Metabolite Profiling
Used in medicinal plants
Detection by:
HPLC
TLC
GC–MS
Example: Alkaloid or flavonoid variation
D. MOLECULAR MARKER TECHNIQUES


E. SELECTION AND ISOLATION TECHNIQUES
8. In Vitro Selection
Culture cells under selective agents
Select resistant variants
Examples:
Salt stress → salt tolerant plants
Herbicide → herbicide resistant plants
Pathogen toxin → disease resistant plants

9. Somatic Embryo / Cell Line Selection
Isolation of stable cell lines showing desired traits
Regeneration into whole plants


F. FIELD EVALUATION AND STABILIZATION
10. Field Trials
Confirm:
Stability
Heritability
Multi-location trials needed


11. Genetic Stabilization
Through:
Sexual reproduction
Backcrossing
Selfing


II. FACTORS AFFECTING SOMACLONAL VARIATION
Somaclonal variation depends on biological, chemical, physical, and cultural factors.


A. GENOTYPE
Different species and cultivars show varying levels
Highly heterozygous plants show more variation
Example: Sugarcane, banana – high variation
B. EXPLANT SOURCE AND TYPE
Callus-derived plants show more variation than:
Axillary bud culture
Mature tissues > juvenile tissues
Meristem cultures show minimal variation


C. CULTURE MEDIUM COMPOSITION
1. Plant Growth Regulators
High auxin (2,4-D) increases variation
Cytokinin imbalance induces mutations
2. Nutrient Stress
High salts, sugar concentration
Nitrogen source imbalance
D. DURATION AND NUMBER OF SUBCULTURES
Prolonged culture increases:
Chromosomal instability
DNA methylation changes
More subcultures → more variation
E. MODE OF REGENERATION
Regeneration Pathway
Variation
Organogenesis
Moderate
Somatic embryogenesis
High
Direct shoot regeneration
Low
F. PHYSICAL CULTURE CONDITIONS
Light intensity
Temperature
pH of medium
Osmotic stress
G. EPIGENETIC CHANGES
DNA methylation
Histone modification
Often reversible
Responsible for phenotypic plasticity
H. ACTIVATION OF TRANSPOSABLE ELEMENTS
Stress activates mobile genetic elements
Causes insertions and deletions
SIGNIFICANCE OF SOMACLONAL VARIATION
Source of novel genetic variation
Crop improvement without genetic engineering
Development of:
Disease resistant plants
Abiotic stress tolerant plants
Improved quality traits
CONCLUSION
Somaclonal variation is a double-edged sword in plant tissue culture. Proper detection using morphological, cytological, biochemical, and molecular techniques, along with controlled culture conditions, allows effective isolation and utilization of beneficial variants for plant improvement.

Comments

Post a Comment

Popular Posts

••CLASSIFICATION OF ALGAE - FRITSCH

      MODULE -1       PHYCOLOGY  CLASSIFICATION OF ALGAE - FRITSCH  ❖F.E. Fritsch (1935, 1945) in his book“The Structure and  Reproduction of the Algae”proposed a system of classification of  algae. He treated algae giving rank of division and divided it into 11  classes. His classification of algae is mainly based upon characters of  pigments, flagella and reserve food material.     Classification of Fritsch was based on the following criteria o Pigmentation. o Types of flagella  o Assimilatory products  o Thallus structure  o Method of reproduction          Fritsch divided algae into the following 11 classes  1. Chlorophyceae  2. Xanthophyceae  3. Chrysophyceae  4. Bacillariophyceae  5. Cryptophyceae  6. Dinophyceae  7. Chloromonadineae  8. Euglenineae    9. Phaeophyceae  10. Rhodophyceae  11. Myxophyce...
Post a Comment
Read more

Southern Blotting

Southern Blotting  Introduction Southern blotting is a molecular biology technique used for the detection of specific DNA sequences in a complex mixture of DNA. It was developed by Edwin M. Southern in 1975. The method involves restriction digestion of DNA, separation by gel electrophoresis, transfer (blotting) onto a membrane, and hybridization with a labeled DNA probe. Principle of Southern Blotting The technique is based on the principle of complementary base pairing. A single-stranded labeled DNA probe hybridizes specifically with its complementary DNA sequence immobilized on a membrane. Detection of the label confirms the presence and size of the target DNA fragment. Steps Involved in Southern Blotting. 1. Isolation of DNA Genomic DNA is extracted from cells or tissues. DNA must be pure and intact to ensure accurate results. 2. Restriction Enzyme  Digestion DNA is digested using specific restriction endonucleases. Produces DNA fragments of varying lengths. Choice of enzym...
Post a Comment
Read more

Plaque Blotting Technique

Plaque Blotting Technique Introduction Plaque blotting is a molecular biology screening technique used to identify specific DNA or RNA sequences present in bacteriophage plaques formed on a bacterial lawn. It is especially useful in the screening of recombinant phage libraries such as λ (lambda) phage genomic or cDNA libraries. This technique combines: Plaque assay (to isolate individual phage clones) Blotting technique (to transfer nucleic acids onto a membrane) Hybridization (to detect specific sequences using labeled probes) Principle of Plaque Blotting The principle of plaque blotting is based on nucleic acid hybridization. Each plaque represents a clone of phage particles containing identical DNA. DNA from phage particles in plaques is: Released Denatured into single strands Transferred onto a nitrocellulose or nylon membrane The membrane is incubated with a labeled DNA/RNA probe complementary to the target sequence. Hybridization between probe and target DNA identifies positive p...
Post a Comment
Read more

Molecular Marker Techniques

Molecular Marker Techniques (30-Mark Detailed Notes) Introduction Molecular markers are DNA sequences with known locations on chromosomes that can be used to identify individuals, genotypes, or genetic differences. They reveal polymorphism at the DNA level and are not influenced by environmental factors, unlike morphological or biochemical markers. Molecular marker techniques are widely used in genetics, plant breeding, biotechnology, forensics, medical diagnosis, and evolutionary studies. Characteristics of an Ideal Molecular Marker An ideal molecular marker should: Be highly polymorphic Show co-dominant inheritance Be abundant and uniformly distributed in the genome Be environment-independent Have high reproducibility Be easy, rapid, and cost-effective Classification of Molecular Marker    Techniques 1. Hybridization-Based Markers RFLP (Restriction Fragment Length Polymorphism) 2. PCR-Based Markers RAPD AFLP SSR (Microsatellites) ISSR 3. Sequence-Based Markers SNP (Single Nu...
Post a Comment
Read more

DNA FOOTPRINTING

DNA FOOTPRINTING Introduction DNA footprinting is a molecular biology technique used to identify the specific site(s) on DNA where proteins (such as transcription factors) bind. It reveals the exact nucleotide sequences protected by bound proteins against cleavage by nucleases or chemical agents. It is widely used to study DNA-protein interactions, transcription regulation, and gene expression control. Definition DNA footprinting: A technique used to locate the binding site of DNA-binding proteins on DNA by detecting protected regions that are resistant to enzymatic or chemical cleavage. Principle DNA-binding proteins protect the DNA segment they occupy. DNA exposed to nucleases (DNase I) or chemical cleavage agents is cut at accessible regions. Regions bound by protein remain unaffected, leaving a “footprint”. When fragments are separated on a denaturing polyacrylamide gel, the missing bands correspond to protein-binding sites. Key idea: Cleavage occurs everywhere except where the pro...
Post a Comment
Read more

Mapping of DNA

DNA MAPPING   1. Introduction DNA mapping refers to the process of determining the relative positions of genes or DNA sequences on a chromosome. It provides information about the organization, structure, and distance between genetic markers in a genome. DNA mapping is an essential step toward genome sequencing, gene identification, disease diagnosis, and genetic engineering. DNA maps serve as roadmaps that guide researchers to locate specific genes associated with traits or diseases. 2. Objectives of DNA Mapping To locate genes on chromosomes To determine the order of genes To estimate distances between genes or markers To study genome organization To assist in genome sequencing projects. 3. Principles of DNA Mapping DNA mapping is based on: Recombination frequency Physical distance between DNA fragments Hybridization of complementary DNA Restriction enzyme digestion Use of genetic markers The closer two genes are, the less frequently they recombine during meiosis. 4 . Types of DNA...
2 comments
Read more

Gene Transfer Technologies – Detailed Notes

Gene Transfer Technologies – Detailed Notes 1. Definition Gene transfer is the process of introducing foreign DNA or genes into the genome of a target organism or cell. It allows the expression of new traits, study of gene function, and production of therapeutic proteins. Also known as gene delivery or genetic transformation. 2. Principles of Gene Transfer Involves delivery of DNA or RNA into cells or organisms. DNA can be integrated into the host genome or remain episomal (non-integrated). The goal is stable or transient expression of the transferred gene. Key considerations: Vector – vehicle for carrying the gene Target cell – plant, animal, microbial, or human cells Delivery method – physical, chemical, or biological 3. Types of Gene Transfer Gene transfer can be broadly classified into: A. Natural Gene Transfer Occurs in nature between organisms: Transformation: Uptake of naked DNA by bacteria. Transduction: DNA transfer via viruses (bacteriophages). Conjugation: Transfer of plasmi...
Post a Comment
Read more

ANTIGEN

1. Definition of ANTIGEN An antigen is any substance which, when introduced into the body, induces an immune response and specifically reacts with antibodies or sensitized T-cells. 👉 Substances may be foreign or self, but immunogenic antigens are usually foreign molecules. 2. Immunogen vs Antigen Immunogen Substance that induces immune response Antigen Substance that reacts with immune products Hapten Antigenic but not immunogenic alone 👉 All immunogens are antigens, but all antigens are not immunogens. 3. Chemical Nature of Antigens Antigens may be: a) Proteins (Most potent) Enzymes Toxins Structural proteins b) Polysaccharides Bacterial capsules Cell wall components c) Glycoproteins Viral envelope proteins d) Lipids & Nucleic acids Weakly antigenic Become immunogenic when combined with proteins 4. Properties of Antigens An ideal antigen shows: Foreignness High molecular weight (>10,000 Da) Chemical complexity Stability Specificity Degradability (processing by APCs) 5. Types ...
Post a Comment
Read more

NORTHERN BLOTTING

NORTHERN BLOTTING – 30 MARK DETAILED NOTES Northern blotting is a molecular biology technique used to detect specific RNA molecules in a complex mixture. It provides information about gene expression, RNA size, and transcript abundance by hybridizing RNA with a labeled complementary DNA or RNA probe. 📌 Named by analogy to Southern blotting (DNA detection). 2. Principle The principle of Northern blotting is based on: Separation of RNA molecules by size using denaturing agarose gel electrophoresis Transfer (blotting) of separated RNA onto a nylon or nitrocellulose membrane Hybridization of membrane-bound RNA with a labeled complementary probe Detection of RNA–probe hybrids by autoradiography or chemiluminescence ✔ Only RNA sequences complementary to the probe will be detected. 3. Types of RNA Analyzed mRNA (most common) rRNA tRNA miRNA and siRNA (with modified protocols) 4. Requirements / Materials Total RNA or poly(A)+ RNA Denaturing agarose gel (formaldehyde or glyoxal) Electrophoresi...
Post a Comment
Read more

Western Blotting

Western Blotting (Immunoblotting) Introduction Western blotting, also known as immunoblotting, is a widely used analytical technique for the detection, identification, and quantification of specific proteins in a complex biological sample. The technique combines protein separation by gel electrophoresis with specific antigen–antibody interaction. The method was developed by Towbin et al. (1979) and is called “Western” in analogy to Southern blotting (DNA) and Northern blotting (RNA). Principle The principle of Western blotting involves: Separation of proteins based on molecular weight using SDS-PAGE Transfer (blotting) of separated proteins onto a membrane Specific detection of the target protein using primary and secondary antibodies Visualization using enzymatic or fluorescent detection systems 👉 Antigen–antibody specificity is the core principle of Western blotting. Steps Involved in Western Blotting 1. Sample Preparation Protein samples are extracted from cells or tissues Lysis bu...
Post a Comment
Read more