What plant species are you working on? The protoplast isolation protocol can be (minor) different between different plant species. The attached article includes detail tobacco protoplast isolation protocol.
By the way, if you are using Arabidopsis as your plant material for protoplast isolation, the attached paper provided a fast and an efficient method. According to Google, since published in 2009, the paper has been cited 271 times.
3.6.2 Tobacco BY2 cells
Suspension cultures of tobacco were maintained in LS medium containing 0.2 mg/l 2,
4-dichloro-phenoxacetic acid (2, 4-D) in the dark at 26 °C, and transferred to fresh
medium every 7 days.
file:///C:/Users/alaah/Downloads/Molecular%20studies%20on%20plants%20to%20enhance%20their%20stress%20tolerance.PDF
Thank you Dr. Alaaeldin A Hilali Coud you send me this paper please?
Hi, for citrus you can check the following:
1- Grosser JW, Gmitter FG, Jr. (1990) Protoplast fusion and citrus improvement. Plant Breeding Reviews 8:339-374
2- Omar AA, Song WY, Grosser JW (2007) Introduction of Xa21, a Xanthomonas-resistance gene from rice, into 'Hamlin' sweet orange [Citrus sinensis (L.) Osbeck] using protoplast-GFP co-transformation or single plasmid transformation. J Horticult Sci Biotechnol 82:914-923
3- Omar AA, Dutt M, Gmitter FG, Jr., Grosser JW (2016) Somatic Embryogenesis: Still a Relevant Technique in Citrus Improvement. In: Germanà MA and Lambardi M (eds) In Vitro Embryogenesis in Higher Plants. Springer, New York, pp. 289-327
let me know if you need anything more related to citrus protoplast
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