May 2012 issue | Plant Omics Journal

In silico motif diversity analysis of the glycon preferentiality of plant secondary metabolic glycosyltransferases

Ritesh Kumar, Rajender S. Sangwan , Smrati Mishra, Farzana Sabir and Neelam S. Sangwan

Central Institute of Medicinal and Aromatic Plants (CIMAP-CSIR), P.O.-CIMAP, Lucknow-226015, India

Abstract
Glucosyltransferases are the class of enzymes which specifically glycosylate various natural and artificial substrate aglycons into their glycosidic linked compounds with enhanced water solubility and transport. In several instances, glucosylation is the last step in the biosynthesis of a number of secondary plant products involving flavonoids, terpenoids, steroidal alkaloids, and saponin biosynthetic pathway. The conjugation reactions catalyzed by UGTs may therefore are critical in regulating the levels of several secondary metabolites including signaling and hormonal compounds. In this work we have analyzed genes from the databases for the presence of GT’s in diverse plant families. Considerable degree of homology was seen in alignment of all available GT sequences in dicot plants as revealed by CLUSTAL-W and other phylogenetic tree constructing tools. Also, a highly conserved motif in their C-terminus, named the PSPG box (Plant secondary product glucosyl transferase signature) was found in all the sequences through motif discovery tools e.g. MEME. The motif discovery tool identified two other distinct motifs in GT sequences, however interestingly P. patens and a putative GT sequence from A. thaliana was found to be deficient in motif 3 at N terminal of the sequence. A wide range of gene sequences were analyzed in a systematic manner to determine the structure, function and evolution of PSPG box motif found at the C-terminal.

Pages 200-210 | Full Text PDF | Supplementary data

Nikita retrotransposon movements in callus cultures of barley (Hordeum vulgare L.)

Emre Bayram, Sibel Yilmaz, Halide Hamat- Mecbur, Gonul Kartal-Alacam, Nermin Gozukirmizi

Istanbul University, Faculty of Science, Department of Molecular Biology and Genetics, 34134, Vezneciler, Istanbul-TURKEY

Abstract
Retrotransposons are dynamic elements of the genome and exist in high percentages in the genome of many organisms. However, the majority of retrotransposons are inactivated during development by different mechanisms such as methylation. Some stress conditions may have a stimulating effect on the activation of retrotransposons. In vitro culture conditions can be considered as one of these stress factors due to nutrients, chemicals, physical factors and photoperiods. In this study, Nikita retrotransposon’s polymorphism was investigated on different calli ages (30-, 60- and 90-day-old) of barley, which developed from the same embryo on MS medium supplemented with 3 mg/L 2,4-D. Mature barley embryos (Hordeum vulgare L. cv. Zafer-160) were cultured for callus formation and sub-cultured every 30 days. Three experiment sets were constructed to determine the polymorphism between individual calli originated from different embryos in the same culture time. Polymorphism was detected using Inter-Retrotransposon Amplified Polymorphism (IRAP) technique with two different Nikita specific forward primers. Three mature embryos were used as control. In total, 20 homomorphic PCR bands were obtained from both reactions in intact embryos. However, some polymorphic bands (~ 550 and 650 bp) were solely observed in calli. Our results showed that tissue culture conditions caused the movement of Nikita retrotransposon at different ages of calli that originated from the same embryo and at the same time. We explained that all individuals did not show the same effect. To our knowledge, this is the first report on the employment of Nikita based IRAP application in barley in terms of callus development.

Pages 211-215 | Full Text PDF

Water relations, nitrogen compounds and enzyme activities in leaf and root of young Yellow Lapacho (Tabebuia serratifolia) plants subjected to flooding

Gustavo Antonio Ruffeil Alves, Benedito Gomes dos Santos Filho, Allan Klynger da Silva Lobato, Daniel Kean Yuen Tan, Cândido Ferreira de Oliveira Neto, Roberto Cezar Lobo da Costa, Fabrício William Ávila, Douglas José Marques, Rosemiro dos Santos Galate

Núcleo de Pesquisa Básica e Aplicada da Amazônia, Universidade Federal Rural da Amazônia, Paragominas, Brazil
Faculty of Agriculture, Food & Natural Resources,University of Sydney, Sydney, NSW 2006, Australia
Center for Agriculture and Health, Cornell University, Ithaca, USA
Departamento de Ciência do Solo, Universidade Federal de Lavras, Lavras, Brazil

Abstract
The aim of this study was to investigate the (i) behavior of young Tabebuia serratifolia (Yellow Lapacho) plants exposed to flooding condition, (ii) measuring the physiological and biochemical parameters in leaf and root, and (iii) to discover if this species can be used in areas where the flooding occurs frequently. A completely randomized block design with two water conditions (control and flooding) was used. The parameters measured were the leaf water potential, stomatal conductance, transpiration rate, nitrate reductase activity, nitrate, free ammonium, glutamine synthetase activity, total soluble amino acids, total soluble proteins, alcohol dehydrogenase activity, and lactate dehydrogenase activity. The data showed significant differences in all parameters. The reductions were observed in water relations and nitrogen compounds. However, increases were determined in alcohol dehydrogenase and lactate dehydrogenase activities after flooding. Regarding with organs evaluated, modifications more intense into root for many parameters such as glutamine synthetase and lactate dehydrogenase enzymes, and leaf was more responsive in nitrate, free ammonium, and alcohol dehydrogenase activity. Therefore, Tabebuia serratifolia plants are sensitive to flooding, and are not suitable for areas prone to flooding.

Pages 216-222 | Full Text PDF

Functional annotation of expressed sequence tags of Papaver somniferum

Avantika Priya, Himanshu Tripathi, Dharmendra K. Yadav, Feroz Khan, Vikrant Gupta, Rakesh K. Shukla, M.P. Darokar

Metabolic and Structural Biology Department, CSIR-Central Institute of Medicinal and Aromatic Plants, P.O.-CIMAP, Kukrail Picnic Spot Road, Lucknow-226015 (U.P.), India
Biotechnology Division, CSIR-Central Institute of Medicinal and Aromatic Plants, P.O.-CIMAP, Kukrail Picnic Spot Road, Lucknow-226015 (U.P.), India

Abstract
Papaver somniferum (opium poppy) is the source for several pharmaceutical benzylisoquinoline alkaloids (BIA) including morphine, codeine and sanguinarine. In an attempt to identify additional biosynthetic steps at the molecular level, a large expressed sequence tag (EST) dataset with a total of 20,532 sequences from opium poppy were assembled and functionally annotated. Sequence cleaning filter showed 20,445 (709 trimmed) valid sequences and 87 trashed sequences at 96% minimum identity parameter for an alignment with a contaminant. Repeat masking filter showed length of valid sequences 1,69,60,151 bp, GC level 41.12% and masked bases 5,01,768 bp (2.96%) by using RepBase (update 8.12) repeats library of Arabidopsis. Elements observed in EST sequences were 1017 retroelements, 55 LINEs, 55 L1/CIN4, 962 LTR elements, 412 Ty1/Copia, 550 Gypsy/DIRS1, 56 DNA transposons, 15 hobo-Activator, 7 Tc1-IS630-Pogo, 7 En-Spm, 11 MuDR-IS905, 6 Tourist/Harbinger, 1 satellite, 920 simple repeats and 1697 low complexity elements. NCBI's vector library UniVec (core) and plastids library of Arabidopsis were used for masking vector and organelle sequences, respectively. For EST sequence assembly, the cutoff for overlap sequence identity was kept 80%. A total of 15,279 assembled unique transcripts were identified, which include 1,408 contigs and 13,871 singletons. Functional annotation was performed through BLASTX using UniProtKB/Swiss-Prot database. EST sequences were retrieved from dbEST (NCBI) and processed, cleaned, clustered and assembled through Gencheck, EGassembler, Phred, RepeatMasker, Cross-Match, Phrap and Cap3 softwares. Mostly hypothetical genes were observed apart from important genes related to secondary metabolism such as enzymes of BIA biosynthesis e.g., stylopine synthase, S-norcoclaurine synthase 1, salutaridine synthase, salutaridine reductase, (S)-coclaurine N-methyltransferase, S-adenosyl-L-methionine:3'-hydroxy-N-methylcoclaurine 4'-O-methyltransferase and salutaridinol 7-O-acetyltransferase. This functionally annotated EST dataset will be a useful resource for further studies such as taxonomy, molecular breeding, genetics, genomics and secondary metabolism.

Pages 223-230 | Full Text PDF

Discrimination among rice varieties based on rapid detection of single nucleotide polymorphisms by a newly developed method, mass spectrometric cleaved amplified polymorphic sequence (MS-CAPS) analysis

Hideyuki Kajiwara, Masayuki Yamaguchi, Hideki Sato, Hiroyuki Shibaike

National Institute of Agrobiological Sciences, 2-1-2 Kannondai, Tsukuba, Ibaraki 305-8602, Japan
National Agricultural Research Center for Tohoku region, 4 Akahira, Shimo-kuriyagawa, Morioka, Iwate 020-0198, Japan
National Institute for Agro-Environmental Sciences, 3-1-3 Kannondai, Tsukuba 305-8604, Japan

Abstract
The method described here discriminates among rice cultivars based on single nucleotide polymorphisms (SNPs). The method is rapid (less than 1 hour), and combines PCR and matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS). We obtained sequence data from genome databases, and identified SNPs that could be used to distinguish among rice cultivars after digestion with restriction enzymes or urasil-DNA glycosylase (UDG). A crude extract was prepared by vortexing rice grains in a tube, and this was used as the template for PCR without further purification of genomic DNA. The PCR primers were designed based on the sequence around the SNP. The PCR was performed with a short extension time (2 s) and the amplicons were treated with restriction enzymes or UDG. Single-stranded DNA (ssDNA) obtained by alkali denaturation was analyzed by MALDI-TOF MS. This method, which we have used previously to identify transgenic genes in plants, successfully discriminated among rice cultivars based on SNPs. We added an asymmetric PCR to obtain greater quantities of ssDNA and a fast PCR step to increase the speed of amplification.

Pages 231-237 | Full Text PDF | Supplementary data

Cloning, heterologous expression and characterization of three thioredoxin h isoforms (OsTrx1, OsTrx20 and OsTrx23) from rice

Zahra Papzan, Azar Shahpiri

Department of Agricultural Biotechnology, College of Agriculture, Isfahan University of Technology, Isfahan-84156-83111, Iran

Abstract
Thioredoxin h (Trx h) is a major cytoplamic and mitochondrial disulfide reductase. In plants, Trx h isoforms are encoded by a multigenic family of genes. The multiplicity of these isoforms raises the question of their functional specificity. In this study, we describe isolation and cloning of three cDNAs encoding different Trx h isoforms, namely OsTrx1, OsTrx20 and OsTrx23. Three Trx h were heterologously expressed in Escherichia coli and their activities were compared using DTT-dependent insulin assay. OsTrx23 and OsTrx1 demonstrated highest (0.05 ?650/min) and lowest (0.016 ?650/min) activity, respectively. In contrast to OsTrx1 and OsTrx20 isoforms, OsTrx23 was efficiently reduced by NADPH-dependent thioredoxin reductase from barley (HvNTR2). The gene expression of three Trx h was analyzed in one, two and three-week old rice seedlings. The transcripts of OsTrx23 with high intensity and OsTrx20 with low intensity were expressed in both root and shoot, whereas OsTrx1 was only expressed in root. All of three isoforms were appeared partially dimerized under non-reducing conditions suggesting that disulfide bridges were responsible for dimerization.

Pages 238-243 | Full Text PDF

The effect of salinity on growth and ion accumulation in six turfgrass species

Md. Kamal Uddin, Abdul Shukor Juraimi, Mohd Razi Ismail and Md. Amirul Alam

Institute Tropical Agriculture, Universiti Putra Malaysia, Serdang, Malaysia
Department of Crop Science, Faculty of Agriculture, Universiti Putra Malaysia, Serdang, Malaysia

Abstract
The objective of this study was to evaluate the effect of salinity on the growth and ion accumulation in six turfgrass species namely Paspalum vaginatum, Zostera japonica, Zostera matrella, Digitaria didcatyla, Cynodon dactyon cv ‘Satiri’, and Cynodon dactylon cv ‘Tifdwarf’. Six salinity levels were applied with electrical conductivity of 0, 88, 176, 264, 352, 440 and 528 mM (sea water). At the highest salinity level (528 mM), the least shoot dry weight reduction was observed in P. vaginatum (40%) compared to control treatment, followed by C. dactylon ‘satiri’ (44%) and Z. japonica (48%). While at this salinity level, maximum shoot reduction was recorded for Z. matrella (55%) followed by C. dactylon ‘Tifdwarf’ (53%). At the highest salinity (528 mM), root dry weight reduction was also least in P. vaginatum (23%) followed by Z. japonica (29%), while the highest recorded in C. dactylon ‘Tifdwarf’ (44%) followed by D. didactyla (39%). Increasing the salinity level decreased the K+, Ca++, Mg++ content and K/Na ratio but increased Na+ content in the shoot and root tissues. P. vaginatum was the less Na+ accumulating species at all salinity levels followed by Z. japonica and Z. matrella, while C. dactylon ‘tifdwarf’ was the highest Na+ accumulating species followed by D. didactyla and C. dactylon ‘satiri’. P. vaginatum was among the least K+ reducing species at all salinity levels followed by Z. japonica and Z. matrella, while the highest K+ reducing species was D. didactyla followed by C. dactylon ‘tifdwarf’ and C. dactylon ‘satiri’. The highest K+/Na+ ratios at all salinity levels were recorded in P. vaginatum followed by Z. japonica and Z. matrella.

Pages 244-252 | Full Text PDF

Optimization of shade condition and harvest time for Dendrobium candidum plants based on leaf gas exchange, alkaloids and polysaccharides contents

Yueping Zheng, Wu Jiang, Evandro Nascimento Silva, Lingzhi Mao, David B. Hannaway, Hongfei Lu

College of Chemistry and Life Science, Zhejiang Normal University, Jinhua, Zhejiang, 321004, China
Biochemistry and Biology Molecular Department, Laboratory of Plant Metabolism, Federal University of Ceará, CP 6004, CEP 60451-970, Fortaleza, Ceará, Brazil
Department of Crop and Soil Science, Oregon State University, Corvallis, OR 97331-3002, United States

Abstract
To meet the increasing demand for Dendrobium candidum plants and its bioactive products, the optimal shade condition and the best harvest time was determined. Gas exchange of D. candidum plants under various shade treatments (93%, 88%, 85% and 75% shading) was measured in four seasons to determine the optimal light intensity for plant growth. Diurnal variations of the net CO2 assimilation rate showed that D. candidum plants under the 85% and 88% shading had the highest value of PN. Also, the 85% and 88% shading treatments resulted in higher values of areas enclosed by diurnal various curves of net CO2 assimilation rate and x axis (AECX) than other treatments in the first two seasons (spring and summer) and the last two seasons (autumn and winter), respectively. To obtain highest levels of bioactive products, the optimal harvest time of D. candidum was determined to be of two-year-old stems grown under 88% shading treatments which had the highest contents of polysaccharides and alkaloids. The relationships among AECX, increased stem volume, leaf area, leaf perimeter, polysaccharide content and alkaloid content showed that biomass and bioactive substance contents were directly proportional to photosynthesis. Therefore, approximately 88% shading and harvesting two-year-old stems in the autumn season can be recommended for this species as photosynthesis, biomass and bioactive products were comprehensively considered.

Pages 253-260 | Full Text PDF

Effects of plant growth substances on callus re-differentiation of medicinal plant Achyranthes bidentata

Hongying Duan, Xiaosheng Ding, Jingjing Xing, Jianying Song, Liu Yang, Longdou Lu

College of Life Science, Henan Normal University, Xinxiang 453007, China

Abstract
In this study, callus from stem could easily differentiate to form bud and root, while re-differentiation potential of callus from leaf and petiole was lower, especially bud was not discovered in callus from petiole. Compared with other plant growth substances, effect of 2,4-D on callus re-differentiation of various explants was extremely significant, simultaneously the interaction among plant growth substances was also discovered, in which the optimum combination of plant growth substances for bud and root formation from callus of stem was respectively as follows, combination of 0.5mg/L 2,4-D, 1.0mg/L NAA, 0.1mg/L IBA and 0.1mg/L ZT, and combination of 0.5mg/L 2,4-D, 1.0mg/L KT, 0.5mg/L NAA and 0.5mg/L IBA. Besides, combination of plant growth substances for root formation from callus of leaf and petiole was also obtained. In brief, the re-differentiation capacity of callus from various explants was different, and roles of plant growth substances in re-differentiation were also significantly diverse, thus these results would provide theory basis for tissue culture and plant regeneration of Achyranthes bidentata.

Pages 261-265 | Full Text PDF

Research Note

Plant regeneration from alginate-encapsulated shoot tips of Momordica dioica for short term storage and germplasm exchange and distribution

Muthu Thiruvengadam, Nagella Praveen, Ill-Min Chung

Department of Applied Life Science, Konkuk University, Seoul 143-701, Republic of Korea

Abstract
An efficient protocol for regeneration of encapsulated shoot tip explants of spine gourd (Momordica dioica Roxb. ex. Willd) has been developed. Shoot tip explants excised from in vivo proliferated shoots were encapsulated in calcium alginate beads. A gelling matrix of 3% sodium alginate and CaCl2.2H2O was found most suitable for formation of ideal calcium alginate beads. Maximum response (100%) for conversion of encapsulated shoot tip explants into plantlets was obtained on 0.7% agar solidified full-strength MS medium containing 0.5 µM BAP. Encapsulated shoot tips could be stored at low temperature (4°C) up to 10 weeks with a survival frequency of 50%. Well developed regenerated plantlets were hardened, acclimatized and established in field with 90% survival frequency, where they grew well without any detectable variations. The present synthetic seed technology could be useful in large-scale propagation as well as short-term conservation, germplasm distribution and exchange of spine gourd.

Pages 266-270 | Full Text PDF

Salinity effects on macro and micro nutrients uptake in medicinal plant King of Bitters (Andrographis paniculata Nees.)

Daryush Talei, Mihdzar Abdul Kadir, Mohd Khanif Yusop, Mohd Puad Abdullah and Alireza Valdiani

Department of Cell and Molecular Biology, Faculty of Biotechnology & Biomolecular, Universiti Putra Malaysia, 43400 UPM, Serdang, Selangor, Malaysia
Medicinal Plant Research Centre, Shahed University, Tehran, Iran
Department of Agriculture Technology, Faculty of Agriculture, Universiti Putra Malaysia, 43400 UPM, Serdang, Selangor, Malaysia
Department of Land Management, Faculty of Agriculture, Universiti Putra Malaysia, 43400 UPM, Serdang, Selangor, Malaysia

Abstract
Plants species require macro and micro nutrients to complete the growth cycles such as photosynthesis, enzyme activities and secondary metabolite production. The objective of the present study was to investigate the effects of salinity on biomass as well as macro and micro nutrients uptake in the medicinal plant of Andrographis paniculata (AP). In this regard, a split plot experiment was carried out based on Randomized Complete Block Design (RCBD) with two factors; five salinity levels (control, 4, 8, 12 and 16 dsm-1) in main plots and 12 different AP accessions in sub-main plot with three replicates. Seventy-day-plants (before flowering stage) were exposed to different salinity levels on a Hoagland medium. The results indicated that salinity levels had a significant effect on the measured traits. Salinity caused a reduction in dry material and salt tolerance index (STI). The sodium (Na+), iron (Fe), zinc (Zn) and copper (Cu) contents significantly increased, while nitrogen (N), phosphorus (P), potassium (K+), calcium (Ca2+), magnesium (Mg2+) and manganese (Mn) content decreased in high salinity levels. Furthermore, under salinity stress, tolerant accessions could produce higher K+, P, N, Mg2+ and Ca2+ and lower Na+, Fe, Zn and Cu than sensitive accessions. Interestingly, Na+ concentration was highly correlated to STI (r = -0.79), while the lowest correlation was observed between P and Na+ concentration (r = -0.15). Salt-stress caused changes in macro and micro nutrients uptake, which may lead to decline in photosynthesis capacity and respiration. This type of abiotic stress can also disturb metabolism of several cellular components such as protein synthesis.

Pages 271-278 | Full Text PDF

The induced physiological changes by foliar application of amino acids in Aloe vera L. plants

Zahra Oraghi Ardebili, Ali Reza Ladan Moghadam, Narges Oraghi Ardebili, Amir Reza Pashaie

Department of Biology, Garmsar Branch, Islamic Azad University, Garmsar, Iran
Department of Agriculture, Garmsar Branch, Islamic Azad University, Garmsar, Iran
Garmsar Branch, Islamic Azad University, Garmsar, Iran

Abstract
Amino acids are involved in plant metabolism. This research was conducted to evaluate the physiological changes induced by foliar applied active formulations of amino acids based on the commercial formulation; Aminol-Forte, on the Aloe vera plants. The foliar applications of four different concentrations of Aminol-Forte (0, 0.05, 0.1 and 0.15% v/v) were applied on plants. The activities of antioxidant enzymes, peroxidase and poly phenol oxidase, significantly increased at the applied concentrations of 0.15 and 0.1%. Foliar applied amino acid treatments, 0.1 and 0.15%, resulted in significant induction of phenylalanine ammonia lyase activities and improved phenol contents. The applied treatments of 0.05, 0.1 and 0.15% enhanced alkaloid contents. The foliar used amino acids of 0.15% led to a significant increase in the contents of total soluble carbohydrates. The contents of antioxidant compounds, ascorbate and reduced glutathione, were significantly affected by foliar applied amino acid treatment, more specifically the concentrations of 0.1 and 0.15%. Antioxidant activities, free radical scavenging capacity, of all amino acid treated plants were more than control plants. The obtained results from the present research indicated that the foliar application of amino acid, especially at suitable concentrations, had stimulating effects on the content of secondary metabolites, antioxidants and antioxidant activity in Aloe vera.

Pages 279-284 | Full Text PDF

Ribulose-1, 5-biphosphate carboxylase (rbcL) gene sequence and random amplification of polymorphic DNA (RAPD) profile of regionally endangered tree species Coptosperma graveolens subsp. arabicum (S. Moore) Degreef

Sameera O. Bafeel, Abdullah Alaklabi, Ibrahim A. Arif, Haseeb A. Khan, Ahmad H. Alfarhan, Anis Ahamed, Jacob Thomas and Mohammad A. Bakir

Department of Biology, College of Science, King Abdulaziz University (KAU), Jeddah, Saudi Arabia
Department of Biology, College of Arts and Science, Al-Baha University (BU), Baljurashi, Saudi Arabia
Department of Botany and Microbiology, College of Sciences, King Saud University (KSU), Riyadh, Saudi Arabia
Molecular Fingerprinting and Biodiversity Unit, Prince Sultan Research Chair for Environment and Wildlife, College of Sciences, King Saud University, Riyadh, Saudi Arabia

Abstract
Coptosperma graveolens subsp. arabicum (Rubiaceae) is a shrub or small tree and currently endangered in Saudi Arabia. In this study, ribulose-1, 5-biphosphate carboxylase (rbcL) gene sequence and random amplification of polymorphic DNA (RAPD) profile of C. graveolens were determined. Our study showed that this species is phylogenetically related with Tarenna supra axilaris, Coptosperma sp. and Paracephaelis sp. group on the basis of rbcL gene sequences. The studied specimen and related taxa inferred from rbcL gene sequence, demonstrated a distinct lineage; therefore, could distinguish the species as C. graveolens unequivocally. The specimen (6R1) demonstrated the highest level of sequence similarities (99.5%) with three close species in the group. RAPD profile of C. graveolens generated amplified products ranging from 218 to 1338 bp. A total of 18 bands were observed for plant species using 5 commercially available RAPD primers (P). The number of major bands for the plant species for a single primer ranged from 3 to 5. The maximum number of major bands was observed for the P1 (5 bands), 4 bands for P4 and the equal number of bands (3) for P2, P3 and P5. The determined rbcL gene sequence and RAPD profile of the endangered plant species C. graveolens will improve the identification process with morphology based taxonomic methods.

Pages 285-290 | Full Text PDF | Supplementary data

Genotype-dependent response of St. John`s wort (Hypericum perforatum L.) shoot tips to cryogenic treatment: Effect of pre-culture conditions on post-thaw recovery

Linda Petijová, Matúš Skyba, Eva Cellárová

Institute of Biology and Ecology, Faculty of Science, P. J. Šafárik University in Košice, Mánesova 23, 041 54 Košice, Slovakia

Abstract
Cryopreservation of medicinal plants is an important tool for restoration of their potential to synthesize valuable secondary metabolites. Establishment of an efficient protocol requires detailed understanding of species/genotype-specific responses to changing external variables. The aim of this work was to study the genotype-dependent response of Hypericum perforatum L. shoot tips to different pre-cryogenic conditions. Shoot apices isolated from plants growing on the hormone-free medium and from clusters induced by addition of benzylaminopurine, were subjected to pre-culture using two agents (0.3M sucrose and 0.076µM abscisic acid) prior to cryopreservation. Although the genotypes represented the seed progeny of one mother plant, the results showed their different capability to withstand cryogenic treatment. The differences were apparent especially in case of plants cultivated on the hormone-free medium. The ability to recover depended also on the pre-culture agent, duration of the pre-culture period and under certain conditions also on interaction between benzylaminopurine and abscisic acid in the medium. The results are illustrated by anatomical alterations of the meristematic zones and adjacent leaf primordia. Long term exposure of cultures to benzylaminopurine resulted in the delay of abscisic acid effect during pre-culture step and thus impaired regeneration capability of shoot apices after cryopreservation.

Pages 291-297 | Full Text PDF

Characterization of expression patterns of small RNAs among various organs in Arabidopsis and rice based on 454 platform-generated high-throughput sequencing data

Chaogang Shao, Xiaoxia Ma, Ming Chen, Yijun Meng

College of Life Sciences, Huzhou Teachers College, Huzhou 313000, P. R. China
College of Life and Environmental Sciences, Hangzhou Normal University, Hangzhou 310036, P. R. China
Department of Bioinformatics, College of Life Sciences, Zhejiang University, Hangzhou 310058, P. R. China

Abstract
The advent of high-throughput sequencing (HTS) strengthened our capacity for small RNA (sRNA) discovery. Here, we did a transcriptome-wide survey of sRNAs, mainly focusing on the microRNAs, with organ-specific expression patterns in both Arabidopsis (Arabidopsis thaliana) and rice (Oryza sativa). By using sRNA HTS data generated by 454 sequencing technology, four organs in Arabidopsis (i.e. six-day-old seedling, rosette leaf, flower and silique), and four in rice (shoot apex, leaf, root apex and inflorescence) were investigated. Chromosome-wide distribution patterns of the organ-specific sRNAs were obtained. We found that, in rice, the 21-nt (nucleotide) sRNAs occupy a large portion of the sRNA population highly expressed in inflorescences. In contrast, the sRNAs not expressed in rice floral organ are predominantly 24 nt in length. Through literature mining, ath-miR156d, ath-miR400, ath-miR822 and ath-miR824 were suggested to be involved in seedling development in Arabidopsis, and osa-miR169 induced by drought and high salinity was indicated to regulate leaf growth in rice.

Pages 298-304 | Full Text PDF | Supplementary data

Transcriptional analysis of hydroxycinnamoyl transferase (HCT) in various tissues of Hibiscus cannabinus in response to abiotic stress conditions

Emran Md Chowdhury, Bo Sung Choi, Sang Un Park, Hyoun-Sub Lim, and Hanhong Bae

School of Biotechnology, Yeungnam University, Gyeongsan 712-749, Republic of Korea
Department of Crop Science, Chungnam National University, Daejeon 305-754, Republic of Korea
Department of Applied Microbiology, Chungnam National University, Daejeon 305-764, Republic of Korea

Abstract
We cloned a full-length gene from the kenaf plant putatively encoding hydroxycinnamoyl CoA:shikimate/quinate hydroxycinnamoyl transferase enzyme (HcHCT), which is involved in the lignin biosynthesis pathway. We examined the tissue and organ specific expression of an HcHCT ortholog during developmental stages and in response to abiotic stresses. The full-length of the HcHCT ortholog consisted of a 1,296 bp open reading frame (ORF) encoding 431 peptides. The molecular weight of deduced amino acids was 47.71 kDa, with an isoelectric point (pI) of 5.79. The deduced amino acid sequence showed 80-86% identities with HCTs of other plants. The deduced amino acid sequence of the HcHCT ortholog has a histidine containing motif (HHAAD), characteristic for acyl transfer catalysis. A second consensus sequence, a DFGWG block, is another acyl transferase of the BAHD family. Phylogenetic analysis showed the closest relationship (86%) with HCT of Populus trichodcarpa (ACC63882). According to quantitative real-time reverse transcription PCR (QPCR) analysis, HcHCT transcript was expressed in all the tissues and organs, but the highest expression was observed in roots and mature flowers. The expression of HcHCT transcript was also examined in stem tissues of 3-week-old kenaf plants in response to various abiotic stresses. The expression of HcHCT transcript was highly induced by all treatments, including wound, SA, NaCl, cold, H2O2, ABA, and drought. HcHCT was highly expressed in response to cold, SA, and H2O2 at 24 h, 6 h, and 6 h after treatment, respectively. Our results suggest that we have cloned the full-length gene putatively encoding for HCT, which is responsive to various abiotic stresses.

Pages 305-313 | Full Text PDF

Micropropagation of lisianthus (Eustoma grandiflorum), an ornamental plant

Sara Ghaffari Esizad, Behzad Kaviani, Alireza Tarang and Sahar Bohlooli Zanjani

Department of Horticultural Science, Rasht Branch, Islamic Azad University, Rasht, Iran
Department of Horticultural Science, Rasht Branch, Islamic Azad University, Rasht, Iran
Department of Tissue Culture, Branch of North Region of Iran (Rasht), Agricultural Biotechnology Research Institute of Iran (ABRII), Iran

Abstract
Lisianthus (Eustoma grandiflorum) is an ornamental plant with beautiful flowers. Micropropagation is a powerful tool for large-scale propagation of ornamental plants. The shoot tips explants from Lisianthus were cultured on MS medium supplemented with concentrations of 0, 0.5, 1 and 2 mg/L of NAA and KIN. Here, we present a simple and reliable strategy for micropropagation of Eustoma grandiflorum in presence of the single growth regulator, KIN, which enables the production of stock plants. Multiple shoots containing roots can be obtained simultaneously on MS basal medium only supplemented with 0.5-1 mg/L KIN. Shoot tips media supplemented with 1 mg/L KIN without NAA resulted in the best shoot length per explant (2.058 cm) and shoot number per explant (2.62). Also, the most number of nodes per explant (8.86) was obtained in medium containing 0.5 mg/L KIN without NAA. The highest root number per shoot (2.40) was seen in medium supplemented with 2 mg/L KIN + 0.5 mg/L NAA. Shoot tips grown in medium containing 2 mg/L NAA without KIN showed the most callus formation. The results of this study revealed that the best shoot proliferation was achieved in MS medium supplemented with 0.5 or 1 mg/L KIN without NAA. Regenerated plants were transferred to peat and perlite (1:1) after hardening and they showed 100% survival.

Pages 314-319 | Full Text PDF