Isolation, culture and regeneration of protoplasts from potato and several related Solanum species

Haberlach, Geraldine T.; Cohen, Boaz; Reichert, Nancy A.; Baer, M.; Towill, Leigh E.; Helgeson, John P.

doi:10.1016/0168-9452(85)90194-3

Cited by 120 publications

(54 citation statements)

References 12 publications

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“…Protoplasts were isolated from the mesophyll of young leaves that were pretreated according to Haberlach et al [14]. Excised leaves were placed in floatation medium (CaCl 2 and NH 4 NO 3 1 mM each, NAA 10.7 µM, NAA 4.4 µM) at 20ºC in the dark.…”

Section: Protoplast Isolationmentioning

confidence: 99%

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Tetraploid somatic hybrids of potato (Solanum tuberosum L.) obtained from diploid breeding lines

Przetakiewicz

Nadolska‐Orczyk

Kuć

et al. 2007

Cellular and Molecular Biology Letters

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Intraspecific somatic hybrids between 16 different diploid breeding lines of Solanum tuberosum L. were produced by PEG-induced fusion. Manually selected heterokaryons were cultured in a Millicells-CM using a postfusion protoplast mixture. Plants were regenerated from calli derived from heterokaryons obtained from 10 out of 38 combinations of diploid lines. Of the tested putative somatic hybrids, 14.2% were diploid, 72.8% were tetraploid and 13% pentaploid. The DNA amplification pattern obtained with RAPD or semirandom primers confirmed that 6 fusion combinations were hybrids. In most cases, the morphological traits were intermediate to those of the diploid fusion partners. About 23.0% of the tested somatic hybrids showed variation in their morphology. Of the tested somatic hybrids, 78.0% flowered and 86.0% tuberized. The cytoplasm of 9 diploid lines and 6 somatic hybrid combinations was analysed. Two of the diploid lines had W/S chloroplasts and α or ε mitochondria; the remainder contained T chloroplasts and β mitochondria. All the analysed somatic hybrids carried T chloroplasts and β mitochondria.

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Section: Protoplast Isolationmentioning

confidence: 99%

“…At this point of the culture, dividing cells were counted. Protoplastderived colonies were transferred onto C medium [14]. The number of minicalli (counted after two weeks) was expressed as the percentage of the selected putative heterokaryon cells forming minicalli.…”

Section: Protoplast Fusion Selection and Culture Of Heterokaryonsmentioning

confidence: 99%

Tetraploid somatic hybrids of potato (Solanum tuberosum L.) obtained from diploid breeding lines

Przetakiewicz

Nadolska‐Orczyk

Kuć

et al. 2007

Cellular and Molecular Biology Letters

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“…Five previously reported regeneration media were evaluated for the ability to promote shoot induction. Medium R-1 was adopted from Haberlach et al (1985) and Medium R-2 from Shepard and Totten. (1977).…”

Section: Proliferation Of Calli and The Regeneration Of Plantsmentioning

confidence: 99%

Somatic hybridization between Solanum tuberosum and S. cardiophyllum

Shi¹,

Chen²,

Li³

et al. 2006

Can. J. Plant Sci.

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. 2006. Somatic hybridization between Solanum tuberosum and S. cardiophyllum. Can. J. Plant Sci. 86: 539-545. The wild diploid Mexican species, Solanum cardiophyllum Lindl. (2n = 2x = 24), is resistant to important potato diseases. However, introgression of resistance to the tetraploid cultivated potato (S. tuberosum L.) (2n = 4x = 48) by conventional crossing is not feasible due to the difference in their endosperm balance number between these species. Somatic hybrids between S. cardiophyllum and S. tuberosum were produced for the first time by electrofusion of protoplasts isolated from young leaves of each parental line. The hybrid nature of the regenerated plants was confirmed based on morphology, chromosome number and DNA species-specific RAPD markers. All the somatic hybrids produced a violet pigmentation on their stems and petioles, which resembled the wild partner. Most of the hybrid plants had 2n = 72 chromosomes and exhibited a morphology intermediate between the two fusion parents, but with a tendency towards cultivated potato. These plants flowered and set fruit when backcrossed with their S. tuberosum fusion parent. Mots clés: Pomme de terre (Solanum tuberosum), S. cardiophyllum, fusion des protoplastes, marqueur RAPD Potatoes (Solanum tuberosum L.) are plagued by many kinds of diseases. Of these diseases, late blight, caused by Phytophthora infestans (Mont.) de Bary, is the most devastating one worldwide. Under cool wet conditions, the pathogen rapidly attacks the leaves, stems and tubers, causing significant reductions in yield and quality. All of the current commercial potato cultivars are highly susceptible to late blight. Surveys have shown that the disease costs potato growers $3 billion annually worldwide . Control of potato late blight is currently achieved through the extensive use of fungicides. Therefore, it is not surprising that seeking a genetic resource with resistant traits to the disease has long attracted potato breeders' attention. For example, the Mexican wild species Solanum demissum Lindl. was reported to be resistant to late blight (Reddick 1930). Unfortunately, the resistance was only effective against certain races of the pathogen (Black and Gallegly 1957). An accession of another Mexican wild species, S. cardiophyllum Lindl., was recently found to possess a high level of resistance to late blight . However, S. cardiophyllum and the cultivated potato, S. tuberosum are reproductively incompatible due to differences in their endosperm balance numbers although they can be crossed with several other diploid Mexican wild species (Chen et al. 2004). Thus, this prevents the exploitation of this valuable germplasm using conventional breeding methods.Protoplast fusion offers an alternative to conventional crossing as it provides a means of circumventing the sexual reproduction barriers and allows the incorporation of desirable traits from wild species into cultivated potatoes. Using somatic hybridization, several resistance genes have been introduced into cultivated potato from wild s...

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“…Harvested leaves were conditioned in medium 'Con' (Haberlach et al 1985) at 4 • C for 24 h prior to enzymatic digestion and cellulase 'Onozuka' RS (Yakult Pharmaceutical Ind. Co. Ltd., Japan) was used instead of cellulase R-10.…”

Section: Plant Transformationmentioning

confidence: 99%

“…Protoplast culture was carried out in alginate gel (Vieira et al 1990), with a modified version of the initial culture medium of Cardi et al (1990), exchanging 0.1 mg l −1 NAA and 0.5 mg l −1 zeatin for 1 mg l −1 NAA and 0.4 mg l −1 BAP. After 2 weeks, this was followed sequentially by liquid versions of media "Cul" and "Dif" (Haberlach et al 1985) at 4-6 week culture intervals, before finally culturing isolated protoplast-derived calli (pcalli) on the surface of solid shoot regeneration medium (Sidorov et al 1999;"Mon"). Antibiotic selection commenced 2 weeks post-transformation with the application of 5 mg l −1 hygromycin, which was then increased to 10 and 15 mg l −1 following a further 1 and 2 weeks of culture.…”

Section: Plant Transformationmentioning

confidence: 99%

Direct gene transfer in potato: A comparison of particle bombardment of leaf explants and PEG-mediated transformation of protoplasts

Craig¹,

Gargano²,

Scotti³

et al. 2005

Plant Cell Rep

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Direct gene transfer methods in potato would facilitate the transfer of multiple genes and the manipulation of metabolic pathways in this species. In this study, up to 1.8 transformation events per shot (=0.5 per bombarded leaf) and 67.2 events per million protoplasts treated were obtained with particle bombardment and PEG-mediated direct DNA uptake, respectively. Limited disassociation of both HPT and GUS genes appeared to occur during the process of integration in only 19% of transformants. A large number of transformed potato plants with transgene expression at levels comparable to Agrobacterium-mediated transformation was obtained. High levels of GUS expression were only obtained in lines derived from PEG treatment. No correlation between the number of gene insertions and gene expression levels was found, suggesting that multiple insertions may have little or no effect on transgene expression.

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Isolation, culture and regeneration of protoplasts from potato and several related Solanum species

Cited by 120 publications

References 12 publications

Tetraploid somatic hybrids of potato (Solanum tuberosum L.) obtained from diploid breeding lines

Tetraploid somatic hybrids of potato (Solanum tuberosum L.) obtained from diploid breeding lines

Somatic hybridization between Solanum tuberosum and S. cardiophyllum

Direct gene transfer in potato: A comparison of particle bombardment of leaf explants and PEG-mediated transformation of protoplasts

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