Cloning and characterization of a cathepsin D inhibitor gene from Solanum tuberosum L.

A DNA clone encoding a cathepsin D inhibitor CathInh was isolated from a potato genomic library using a CathInh cDNA as hybridization probe. The amino acid sequence of the coding region is nearly identical with a CathInh cDNA and CathInh proteins previously isolated from a tuber-specific cDNA library and from tubers, respectively. Analysis of GUS activity resulting from expression of chimeric CathInh promoter-GUS genes in transgenic potato plants revealed expression exclusively confined to potato tubers. No GUS activity could be detected in any other organ of the transgenic plants either constitutively or after wounding or treatment with abscisic and jasmonic acid (JA). Interestingly, part of the promoter region of the CathInh gene, essential for GUS activity in tubers, shows striking similarity to promoter regions of tuber-specific class I patatin genes.     

Cloning and Characterization of One Member of the Chalcone Synthase Gene Family from Solanum tuberosum L.

We have isolated a chalcone synthase gene 2 (ST-CHS2) from potato by rapid amplification of cDNA ends by PCR. CHS2 cDNA had high homology to tomato LET-CHS2 (98%), petunia PHCHSJ (94%), potato ST-CHS1B (92%), petunia PHCHSA (92%), and LET-CHS1 (90%) at the overall 389-amino acid level. Genomic hybridization analysis indicated that CHS genes of potato comprise a family of at least six individual members.     

A Novel Sesquiterpene,Tuberonone, from Solanum tuberosum L.

The effects of the ovarian hormones progesterone and estrone on the conversion of tryptophan to nicotinamide in rats were investigated. Female rats were fed for 35 days with a 20% casein diet, or with a 20% casein diet containing 0.1% progesterone, or 0.001% estrone, or 0.1% progestrone and 0.001% estrone. The conversion ratio of tryptophan to nicotinamide on the last day of the experiment was 2% in the groups fed with the 20% casein diet and the diet containing 0.1% progesterone, but around 1.2% in the group fed with 0.001% estrone, and 0.7% in the group fed with 0.1% progesterone and 0.001% estrone. These results demonstrated that administration of ovarian hormones significantly decreased the conversion of tryptophan to nicotinamide.     

Salt Tolerance of Solanum tuberosum L. Overexpressing an Heterologous Osmotin-like Protein

Potato (Solanum tuberosum L. cv. Bintje) was transformed with a cDNA clone encoding an osmotin-like protein. Transgenic and non-transgenic in vitro plants were subjected to NaCl for 3 weeks. The shoot and root development was slightly affected by salinity indicating that the salt condition used was a mild stress. The endogenous proline content of the osmotin-like transformed clone only raised slightly as compared to the non-transformed genotype, where a marked increase in proline content could be observed as a result to salt stress. These data provide evidence for the involvement of osmotin-like proteins in the mechanisms of salt tolerance in potato plants. This revised version was published online in July 2006 with corrections to the Cover Date.     

Studies on Plant Regeneration from Mesophyll Protoplasts of Solanum tuberosum L.

Protoplasts from potato mesophyll of two strains (Solannum tuberosum L. cv. Xiao Yie Zi x Duo Zi Bia and Solanum tuberosum L. cv. Wu Meng 601) were induced to callus in culture medium of protoplasts. The callus derived from mesophyll protoplasts were transferred to MS medium with 2 mg/l ZT+0.1 mg/L IAA. Shoots regenerated from the callus were detected after 70 days of culture.The shoots which had grown to a height of 2–3 cm were transferred to MS medium with 0.05 mg/L NAA. Roots were coming out in a few days.Complete plantlets were achieved. Stern segments with 1–2 leaves were then transferred to a mixture of sterilized soil and grown, and produced tuber.     

Hydrolysis of sucrose within isolated vacuoles from Solanum tuberosum L. tubers

The soluble acid invertase (β-D-fructofuranoside fructohydrolase, EC 3.2.1.26) from potato (Solanum tuberosum L. cv. Kennebec) tubers was located in the vacuoles. Although the functionality of this invertase in the vacuoles has been assumed, the activity of the enzyme has never been shown within isolated vacuoles. Vacuoles were prepared by gentle osmotic shock from free protoplasts obtained by enzymic digestion of tuber tissues. The mean volume of these vacuoles, (0.26 ± 0.05) × 10⁻² μl, was estimated by optical microscopy. Sucrose, glucose and fructose concentrations were calculated to be 100 mM, 20 mM and 40 mM, respectively, in the vacuoles. Sucrose hydrolysis and the increase in glucose and fructose concentrations within the vacuoles were measured during vacuolar incubations. An almost identical pattern of sucrose hydrolysis by invertase was found by an in-vitro assay reproducing the vacuolar conditions. In view of the determinations of internal vacuolar pH (5.2), the possibility of spontaneous hydrolysis of sucrose was disregarded. Vacuoles were shown to be free from proteinaceous inhibitors, confirming the extravacuolar location of these inhibitors. The vacuolar hydrolytic pattern of sucrose confirms the regulatory role of the reaction products previously proposed for in-vitro assays. Received: 21 July 1997 / Accepted: 31 August 1997     

Cloning and functional analysis of a cDNA encoding a novel 139 kDa starch synthase from potato (Solanum tuberosum L.)

Three isoforms of starch synthase were shown to be present in soluble potato tuber extracts by activity staining after native gel electrophoresis. An antibody directed against a domain conserved in starch synthases was used to clone a cDNA for one of these isoforms by screening a tuber-specific expression library. A partial cDNA of 2.6 kbp was obtained and used to isolate a full-length cDNA of 4167 bp. The deduced amino acid sequence identifies the protein as a novel type of starch synthase from potato with a molecular mass of 139.2 kDa for the immature enzyme including its transit peptide. This novel isoform was designated SS III. An analysis of the expression pattern of the gene indicates that SS III is equally expressed in tubers of different developmental stages as well as in sink and source leaves. In several independent transgenic potato lines, where the expression of SS III was repressed using the antisense approach, the activity of a specific starch synthase isoform was reduced to non-detectable levels as determined through activity staining after native gel electrophoresis. The reduction of this isoform of starch synthase leads to the synthesis of a structurally modified starch in the transgenic plants: there is a drastic change in granule morphology and an increased level of covalently linked phosphate.     

Changes of gene expression in Solanum tuberosum L. as a result of transgenes

Potato plants of various cultivars, transformed using Agrobacterium tumefaciens with pGV941 plasmid, differed from control plants in glyphosate herbicide tolerance, tryptophane content, intensity of callusogenesis, microtuber formation in vitro and multimolecular forms of peroxidase (EC 1.11.1.7) and superoxide dismutase (EC 1.15.1.11). The results demonstrate the influence of alien DNA on structural gene expression in transgenic plants.     

Comparative genome analysis of Solanum lycopersicum and Solanum tuberosum

Solanum lycopersicum and Solanum tuberosum are agriculturally important crop species as they are rich sources of starch, protein, antioxidants, lycopene, beta-carotene, vitamin C, and fiber. The genomes of S. lycopersicum and S. tuberosum are currently available. However the linear strings of nucleotides that together comprise a genome sequence are of limited significance by themselves. Computational and bioinformatics approaches can be used to exploit the genomes for fundamental research for improving their varieties. The comparative genome analysis, Pfam analysis of predicted reviewed paralogous proteins was performed. It was found that S. lycopersicum proteins belong to more families, domains and clans in comparison with S. tuberosum. It was also found that mostly intergenic regions are conserved in two genomes followed by exons, intron and UTR. This can be exploited to predict regions between genomes that are similar to each other and to study the evolutionary relationship between two genomes, leading towards the development of disease resistance, stress tolerance and improved varieties of tomato.     

麻疯树中ADP核糖基化因子基因的克隆和表达

从麻疯树cDNA文库中筛选到包含完整编码序列的ADP核糖基化因子的cDNA序列,命名为Jc-arf．其长度为887bp,开放阅读框546bp,推测的编码蛋白含181个氨基酸残基,具有典型的GTP结合蛋白家族的特点：P框（GLDAAGKT）、G’框（DVGGQ）和G框（NKQDL）。序列分析显示,Jc-arf矿接近于人类ClassI型arf基因,其蛋白序列与多种植物ARF有很高的同源性。半定量RT-PCR结果显示,Jc-arf的表达具有一定的组织特异性,在花中最丰富。PEG、低温、NaCl胁迫下,Jc-arf的表达受PEG和低温的诱导,而对NaCl胁迫不敏感。     

Growth and photosynthesis of photoautotrophic callus derived from protoplasts of Solanum tuberosum L.

Summary We describe a photoautotrophic culture procedure of potato (cvs Kennebec, Haig, DTO-33) callus derived from mesophyll protoplasts. The protoplast culture was initiated at very low concentration of glucose (down to 0.25 g l^–1). Callus was subcultured under CO₂ enriched air and glucose suppressed by the successive dilutions with glucose free media. Regeneration was successfully obtained under photoautotrophic conditions. The characterization of oxygen exchange and of some enzymes and metabolites of carbon assimilation indicated that chlorophyllous callus, grown on carbohydrate free medium, developed the photosynthetic pathway typical of C3 plants. By comparing the fresh weight of callus cultivated in the light or in non-photosynthetic conditions (in darkness or in the light +3-(3,4-Dichlorophenyl)-1,1-dimethylurea) we concluded that growth depended to about 70 to 88 % on photosynthesis.Abbreviations BAP 6-Benzylaminopurine - cv Cultivar - DCMU 3-(3,4-Dichlorophenyl)-1,1-dimethylurea - 2,4-D (2,4-Dichlorophenoxy) acetic acid - GA3 Gibberellin A₃ - IAA Indole-3-acetic acid - NAA 1-Naphthaleneacetic acid - PePcase Phosphoenolpyruvate carboxylase - RuBPcase Ribulo1,5-bisphosphate carboxylase - ZEA Zeatin     

A study of mitochondrial ribosomes from the higher plant Solanum tuberosum L.

Ribosomal subunits are isolated from potato tuber mitochondria devoid of contaminating organelles. The sedimentation constants of the two mitochondrial ribosomal subunits are 33S and 50S respectively. The apparent sizes of the high molecular weight RNAs are 19S and 25S.The proteins of these ribosomes have been analyzed by two-dimensional electrophoresis in SDS polyacrylamide gels to determine their number and molecular weights. The small subunit contains 35 protein species ranging from 8 to 60 kDa. The 50S large subunit contains 33 protein species ranging from 12 to 46 kDa. These preliminary results are the first analysis made on mitochondrial ribosomes from a higher plant.