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  Section: General Biotechnology / Plant Biotechnology
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In Vitro Culture Techniques : The Biotechnological Principles

Protoplast Fusion in Fungi
The essence of protoplast fusion in fungi is the improvement of strains to be used for commercial purposes. By this method compatibility barriers between .several species can be overcome] In recent years, several companies have realized to exploit this technique as a breeding tool for strain development in fungi. For example, two titre strains of Cephalosporium acremonium were crossed and cephalosporin C was improved. But attempts to improve citric acid production in A. nigev were unsuccessful (Peberdy, 1989).

Method for isolation of protoplasts is the same as described for plant cells. Generally, PEG is used for inducing protoplast fusion. But many of isolated protoplasts die due to PEG treatment. This results in low rate of survival which facilitates to detect them by using selection techniques.

Electrofusion, a two stage process, is another method for inducing protoplast fusion. In this method, the protoplast membranes are brought into close contact by dielectrophoresis and a field pulse is used to cause the fusion event. For the first time, it was applied in S. cerevisiae (Halfmann et. al, 1982). 

The fungal protoplast techniques are applied in the following four major areas to get the crosses (i) between isogenic strains which may provide an opportunity for genetic mapping, (ii) between different strains or isolates of a species primarily for breeding, (iii) between apparently incompatible strain of the same species, and (iv) between different species and genera.

Intraspecific protoplast fusion
Intraspecific protoplast fusion is the cross between the same species in an individual which involves the isogenic strains or the non-isogenic ones. The true value of protoplast fusion as a mean for establishing parasexual crosses has been realized so far in a few fungi. For example, in C. acremonium this technique offers the only way of carrying out crosses  and genetic analysis. Difficulty arises through the conventional methods because of production of heterokaryons and somatic diploids. However, through this technique the fusion products were not the heterokaryons but the haploid recombinants. Protoplast fusion technique made it possible to produce a preliminary genetic map of 8 linkage groups for C. acremonium. Genes which enhance the production of antibiotics have been identified and allied to specific linkage groups. Another example is the yeast, Candida, of which biotechnological potential would be of great use. The other examples are : Absidia glauca, Candida maltosa, Pleurotus ostreatus, Aspergillus niger, A. sojae, Fusarium graminearum, F. lycopersici, Penicillium verruculosum, Trichoderma, harzianum, T. reesei, etc.

Interspecific protoplast fusion
Interspecific protoplast fusion is the crosses between two different species. Interspecific protoplast fusions are of much importance in the area where new products are to be produced. Due to new genetic set up many noval secondary metabolites such as, antibiotics may be produced. Some of the examples where interspecific hybrids were produced through protoplast fusion are: S. cerevisiae x S. fermentali, S. cerevisiae x S. lipolytica, S. cerevisiae x S. rowcil, P. chrysogenum x P. notatum, P. chrysogenum x P. citrinum, P. chrysogenum x P. baarnense, Asnergillus nidulans x A. rugulosus, Ganoderma applanatum x G. lucidum, Pleurotus ostreatus x P. florida.



Historical background

Requirements for cell and Tissue Cultures


A tissues culture laboratory


Nutrient media



Inorganic chemicals



Growth hormones



Organic constitutents






Amino acids

Culture of plant materials


Explant culture


Callus formation and its culture




Root culture


Shoot culture and micropropagation


Cell culture



Benefits from cell culture


Somatic embryogenesis


Somaclonal variation


Protoplast culture








Protoplast fusion and somatic hybridization



Fusion products



Method of somatic hybridization


Anther and pollen Culture



Culturing techniques


In vitro androgenesis (direct and indirect androgenesis)


Mentor pollen technology


Embryo culture


Embryo rescue


Protoplast fusion in fungi


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