title: Isolation, Regeneration and PEG-Induced Fusion of Protoplasts of Pleurotus pulmonarius and Pleurotus florida.
abstract: Inter-specific hybridization between Pleurotus pulmonarius and P. florida was attempted through PEG-induced protoplast fusion to select a fusant. The protocol for protoplast release, regeneration and fusion in these two Pleurotus species was standardized using the variables controlling the process. The mixture of mycolytic enzymes, i.e. commercial cellulase, crude chitinase and pectinase, KCl (0.6 M) as osmotic stabilizer, pH 6 of the phosphate buffer and an incubation time of 3 hours resulted in the maximum release of protoplasts from 3-day-old mycelia of P. florida (5.3~5.75 × 10(7) protoplasts/g) and P. pulmonarius (5.6~6 × 10(7) protoplasts/g). The isolated protoplasts of P. florida regenerated mycelium with 3.3% regeneration efficiency while P. pulmonarius showed 4.1% efficiency of regeneration. Polyethyleneglycol (PEG) - induced fusion of protoplasts of these two species resulted in 0.28% fusion frequency. The fusant produced fruiting bodies on paddy straw but required a lower temperature of crop running (24 ± 2℃) than its parents which could fruit at 28 ± 2℃. The stable fusant strain was selected by testing for the selected biochemical markers i.e. Carbendazim tolerance and utilization of the lignin degradation product, vanillin.
authors.affiliation: Research Centre in Botany, Thiagarajar College (Autonomous), Madurai-625009, India.
authors.forename: M
authors.initials: M
authors.lastname: Eyini
authors.forename: K
authors.initials: K
authors.lastname: Rajkumar
authors.forename: P
authors.initials: P
authors.lastname: Balaji
keywords: Carbendazim; P. florida; Pleurotus pulmonarius; Polyethyleneglycol (PEG); Protoplast fusion
references.citation: Life Sci. 2000 Feb 25;66(14):1359-67
references.pmid: 10755472
references.citation: World J Microbiol Biotechnol. 1993 Sep;9(5):538-43
references.pmid: 24420196
references.citation: World J Microbiol Biotechnol. 1993 Sep;9(5):503-7
references.pmid: 24420189
references.citation: Arch Microbiol. 1975 Apr 7;103(2):199-203
references.pmid: 1171669
references.citation: World J Microbiol Biotechnol. 1996 Nov;12(6):573-8
references.pmid: 24415417