36481 - SOMATIC HYBRIDIZATION Rubina Begum MS PhD1 Jayanti...

Info icon This preview shows page 1. Sign up to view the full content.

View Full Document Right Arrow Icon
This is the end of the preview. Sign up to access the rest of the document.

Unformatted text preview: SOMATIC HYBRIDIZATION Rubina Begum, MS. PhD1; Jayanti Tokas, MS, PhD1; Shalini Jain, MS, PhD2 and Hariom Yadav, MS, PhD2 1 Department of Biotechnology, JMIT, Radaur 2 NIDDK, National Institutes of Health, Bethesda, MD 20892, USA Corresponding: [email protected] Development of hybrid plants through the fusion of somatic protoplasts of two different plant species/varieties is called somatic hybridization Somatic hybridization technique 1. isolation of protoplast 2. Fusion of the protoplasts of desired species/varieties 3. Identification and Selection of somatic hybrid cells 4. Culture of the hybrid cells 5. Regeneration of hybrid plants Isolation of Protoplast (Separartion of protoplasts from plant tissue) 1. Mechanical Method 2. Enzymatic Method 1. Mechanical Method Plant Tissue Cells Plasmolysis Microscope Observation of cells Cutting cell wall with knife Release of protoplasm Collection of protoplasm 1. Mechanical Method Used for vacuolated cells like onion bulb scale, radish and beet root tissues Low yield of protoplast Laborious and tedious process Low protoplast viability Enzymatic Method Leaf sterlization, removal of epidermis Plasmolysed cells Pectinase +cellulase Plasmolysed cells Pectinase Protoplasm released Release of isolated cells cellulase Protoplasm released Isolated Protoplasm Enzymatic Method Used for variety of tissues and organs including leaves, petioles, fruits, roots, coleoptiles, hypocotyls, stem, shoot apices, embryo microspores Mesophyll tissue - most suitable source High yield of protoplast Easy to perform More protoplast viability Protoplast Fusion (Fusion of protoplasts of two different genomes) 1. Spontaneous Fusion 2. Induced Fusion Intraspecific Intergeneric Chemofusion Mechanical Fusion Electrofusion Spontaneous Fusion Protoplast fuse spontaneously during isolation process mainly due to physical contact Intraspecific produce homokaryones Intergeneric have no importance Induced Fusion Chemofusion- fusion induced by chemicals Types of fusogens PEG NaNo3 Ca 2+ ions Polyvinyl alcohal Induced Fusion Mechanical Fusion- Physical fusion of protoplasts under microscope by using micromanipulator and perfusion micropipette Electrofusion- Fusion induced by electrical stimulation Pearl chain of protoplasts is formed by low strength electric field (10kv m-1) Fusion of protoplasts of pearl chain is induced by the application of high strength electric field (100kv m -1) for few microseco Identification and Selection of somatic hybrid cells Hybrid identification- Based on difference between the parental cells and hybrid cell with respect to Pigmentation Cytoplasmic markers Fluorochromes like FITC (fluoroscein isothiocyanate) and RITC (Rhodamine isothiocyanate) are used for labelling of hybrid cells Presence of chloroplast Nuclear staining Heterokaryon is stained by carbol-fuschin, acetocarmine or aceto-orcein stain Hybrid Selection (Several markers are used ) Genetic complementation Phytotoxins Specific amino acid Auxin autotrophy Antibiotics Auxotrophic and metabolic mutants Chromosomal analysis Herbicides Culture of the hybrid cells Hybrid cells are cultured on suitable medium provided with the appropriate culture conditions. Regeneration of hybrid plants Plants are induced to regenerate from hybrid calli These hybrid plants must be at least partially fertile, in addition to having some useful property, to be of any use in breeding schemes. Advantages of somatic hybridization Production of novel intergenic hybrid interspecific and Pomato (Hybrid of potato and tomato) Production of fertile diploids and polypoids from sexually sterile haploids, triploids and aneuploids Transfer gene for disease resistance, abiotic stress resistance, herbicide resistance and many other quality characters Advantages of somatic hybridization Production of heterozygous lines in the single species which cannot be propagated by vegetative means Studies on the fate of plasma genes Production of unique hybrids of nucleus and cytoplasm Limitations of Somatic hybridization Poor regeneration of hybrid plants Non-viability of fused products Not successful in all plants. Production of unfavorable hybrids Lack of an efficient method for selection of hybrids No confirmation of expression of particular trait in somatic hybrids ...
View Full Document

{[ snackBarMessage ]}

What students are saying

  • Left Quote Icon

    As a current student on this bumpy collegiate pathway, I stumbled upon Course Hero, where I can find study resources for nearly all my courses, get online help from tutors 24/7, and even share my old projects, papers, and lecture notes with other students.

    Student Picture

    Kiran Temple University Fox School of Business ‘17, Course Hero Intern

  • Left Quote Icon

    I cannot even describe how much Course Hero helped me this summer. It’s truly become something I can always rely on and help me. In the end, I was not only able to survive summer classes, but I was able to thrive thanks to Course Hero.

    Student Picture

    Dana University of Pennsylvania ‘17, Course Hero Intern

  • Left Quote Icon

    The ability to access any university’s resources through Course Hero proved invaluable in my case. I was behind on Tulane coursework and actually used UCLA’s materials to help me move forward and get everything together on time.

    Student Picture

    Jill Tulane University ‘16, Course Hero Intern