1-O-(indole-3-acetyl)-b-D-glucose: myo-inositol indoleacetyl transferase (IA-myo-inositol synthase) is an important enzyme in IAA metabolism. This enzyme catalyses the transfer of the indole acetyl (IA) moiety from 1-O-(indole-3-acetyl)-b-Dglucose to myo-inositol to form IA-myo-inositol and glucose. IA-myo-inositol synthase was purified to an electrophoretically homogenous state from maize liquid endosperm by fractionation with ammonium sulphate, anion-exchange, adsorption on hydroxylapatite, affinity chromatography on ConA-Sepharose, preparative PAGE and isoelectric focusing. We thus obtained two enzyme preparations which differ in their R f on 8% polyacrylamide gel. The preparation of R f 0.36 contained a single 56.4 kDa polypeptide, whereas the preparation of R f 0.39 consisted of two polypeptides of 56.4 and 53.5 kDa. Both purified preparations of IAInos synthase also exhibited the activity of an IAInos hydrolase, showing that the dual activity was associated with a single protein. Results of gel filtration and analytical SDS-PAGE suggest that the native enzyme exists as both a monomeric (65 kDa) and homo-or heterodimeric form (110-130 kDa). Analysis of peptide maps and amino acid sequences of two 21 amino-acid peptides showed that polypeptides of 56.4 and 53.5 kDa have the same primary structure and that the 3 kDa difference in molecular mass is probably caused by different glycosylation levels. Comparison of this partial and internal amino acid sequence with sequences of other plant acyltransferases indicated similarity to several proteins which belonged to the serine carboxypeptidase-like (SCPL) acyltransferase family.

We exploited the potential of cucurbits for ectopic gene expression. Agroinfiltration is a simple and commonly used method to obtain transient expression of foreign genes in plants. In contrast to in vitro transformation techniques, agroinfiltration can be used for genetic modification of mature plant tissues. Although the cucurbits are commonly used as model plants for molecular biology and biotechnology studies, to date there are no literature sources on the possibility of transient gene expression in mature cucurbit tissues. Our research has shown that mature leaves of Luffa cylindrica L. (luffa), in contrast to other cucurbit species, can be successfully transiently transformed with Agrobacterium tumefaciens. We efficiently transformed luffa leaves with a reporter gene encoding β-glucuronidase (GUS). The GUS activity in transiently transformed leaf tissues was detected within 24 h after the infiltration with bacteria. Additionally, we have shown that the activity of a transiently expressed the GUS gene can be monitored directly in the EDTA-exudates collected from the cut petioles of the agroinfiltrated leaves. The results suggest that luffa leaves can be useful as a plant expression system for studies of physiological and biochemical processes in cucurbits.

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The fluorochrome JC-1 is mainly used in mammalian cells to estimate mitochondrial membrane potential (MMP) as a stress marker, with far less data being available for plants or green algae. To address this, we have validated the possibility for changes in MMP to be used as a sensitive stress indicator in the green alga Chlamydomonas reinhardtii. To optimize the method, we analyzed the conditions for applicable MMP determination, including the proper buffer, excitation/emission wavelengths, and solvent (dimethyl sulfoxide, DMSO) influence on the fluorescence signal, and evaluated usefulness of the method for toxicological research. Our results have allowed us to develop a complete protocol for the estimation of MMP changes in C. reinhardtii. Statistical analyses have confirmed the reproducibility, sensitivity, and applicability of this method for toxicological studies.

Conditioned medium (CM) is a general term describing media in which cells have already been cultivated for some time. Such media, usually clarified by filtration, have been used by plant biotechnologists as additives supporting the growth of cell suspensions, organs and whole plants. This study examined the effect of CM obtained from green alga Desmodesmus subspicatus on the growth and functioning of the photosynthetic apparatus of Nicotiana tabacum and Arabidopsis thaliana in culture in vitro. Plants where cultured on CM diluted 1.25-, 2-and 5-fold with MS medium. The increase in fresh and dry weight was highest in tobacco and Arabidopsis cultured on CM/2 and CM/1.25 media. Those two concentrations also increased the amount of chlorophylls in both plants tested. CM improved parameter PI (reflecting the photosynthetic "vitality" of the organism) and electron transport efficiency, and increased the fraction of active reaction centers. Analysis of chlorophyll fluorescence in vivo suggests that the improvement of these plants grown in the presence of algal CM may result from stimulation of photosynthesis. Algal CM offers a convenient, cheap, universal supplement for stimulating the growth of higher plants in vitro.K Ke ey y w wo or rd ds s: : Algal exudates, in vitro culture, growth improvement, photosynthesis.

Valdensia leaf blight on blueberry in Poland was reported in one commercial nursery plantation near Prażmów, Mazovia voivodship, where heavy defoliation was observed on cv. Bluecrop, grown in nursery pots, in August 2011. Older fruiting bushes were only slightly affected by the disease. Initial symptoms of the disease were small, oval to circular zonated necrosis surrounded with dark brown borders that enlarged on the leaves throughout the canopy. Multicellular, hyaline or light brown, star-shaped conidiospores were observed on the necrotic areas. The mean length of 50 conidiospores from the end of head to the end of arm apex was 307 to 348 μm (4). Eight single-spore isolates of the fungus were obtained. Single conidiospores were picked up from necrotic spots on leaves and transferred with sterile needle on potato dextrose agar (PDA) and incubated at 20°C under ambient light. After 10 days of incubation, total DNA was extracted. Amplification of the internal transcribed spacer (ITS) region of rDNA was done using primers ITS1F and ITS4A (1). PCRs were carried out as follows: initial denaturation at 94°C for 2 min, denaturation at 94°C for 1 min, annealing at 57°C for 1 min, extension at 72°C for 1 min, and final extension at 72°C for 5 min for 28 cycles (Applied Biosystems Veriti 96 Wel Thermal Cycler). Amplicons, which were approximately 520 bp, were sequenced and nucleotide sequences were analyzed by Clustal W2EBI. The sequences of all eight isolates showed 100% similarity to each other and were compared with sequences stored in GenBank using BLAST. Sequences were 525 bp long and showed 100% homology to Valdensinia heterodoxa Peyronel, Sclerotiniaceae (anamorph: Valdensia heterodoxa Peyronel) from Japan and Norway (Accession Nos. AB663682 and Z81447, respectively) (3). The sequence from one isolate was submitted to GenBank (Accession No. KF212190). To fulfill Koch's postulates, each of the eight isolates was used to inoculate 20 healthy young leaves of Vaccinium corymbosum L. cv. Bluecrop and bilberry (V. myrtillus L.) (10 leaves per plant). Mycelial plugs 5 mm in diameter were taken from PDA cultures, approximately 20 days old, and used as inoculum and placed in the center of each leaf and moistened with sterile distilled water. Mycelium-free plugs were used as control. Inoculated leaves were placed in plastic box and incubated at 20°C in laboratory for 5 days, at which time small necrotic lesions consistent with initial symptoms of the disease were observed. Isolates obtained from these symptoms were morphologically identical to those used for inoculation. Control leaves did not show any disease symptoms. Valdensia leaf blight occurrence may be attributed to rainy July and August 2011 and long presence of water on soil surface. In Poland, Valdensinia heterodoxa causes heavy defoliation of Vaccinium myrtillus in pine stands and is a common pathogen of some herbaceous plants (2). To our knowledge, this is the first report of Valdensia leaf blight on highbush blueberry in Poland. References: (1) I. Larena et al. 75:187, 1999. (2) W. Mułenko and S. Woodward. Mycologist 10:69, 1996. (3) S. Nekoduka et al. J. Gen. Plant Pathol. 78:151, 2012. (4) S. Zhao and S. F. Shamoun. Mycology 1:113, 2010.

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