6(2) 2013 issue
March 2013 issue
Southern Cross Publishing Group©2013

Plant Omics Journal | March 2013 issue
Genome organization and non-colinear distribution of the knob-associated sequences in maize

Fei Yang1, Yanhao Xu2, Jing Huang1, Lijia Li1*

1State Key Laboratory of Hybrid Rice, College of Life Sciences, Wuhan University, Wuhan, 430072, Hubei, China
2Engineering Research Center of Wetland Agriculture in the Middle Reaches of the Yangtze River, Ministry of Education, College of Agriculture, Yangtze University, Jingzhou 434025, Hubei, China

The maize (Zea mays L.) genome contains abundant of repetitive elements. In this study, fluorescence in situ hybridization (FISH) was used to reveal the distribution of the knob-associated 180-bp repeats and TR-1 elements among eleven maize inbred lines. Non-colinear distribution of the knob-associated sequences was detected, and a distinct FISH pattern for each maize inbred line was constructed. This result confirmed that FISH using the knob-associated sequences as probes may be a rapid method to identify maize germplasm resources. The 180-bp repeats were more numerous and widely distributed than the TR-1 elements in the maize genome, which provided cytogenetical evidence for the hypothesis that the TR-1 elements were derived from the 180-bp repeats. The knob-associated sequences were concentrated in the satellite of chromosome 6. The signals of the TR-1 elements were readily detected in the satellites of all the maize inbred lines, and the signals of 180-bp repeats were detected in the satellites of six maize inbred lines. This finding indicated that the TR-1 elements tend to accumulate in regions of exceedingly low recombination.

Pages 95-99 | Full Text PDF
Cloning and molecular characterization of selenocysteine methyltransferase (AchSMT) cDNA from Astragalus chrysochlorus

Özgür Çakir1* and Sule Ari1,2 

Department of Molecular Biology and Genetic, Faculty of Science, Istanbul University, 34118 Vezneciler, Istanbul, Turkey
2Research and Application Center for Biotechnology and Genetic Engineering, 34118 Vezneciler, Istanbul, Turkey

Selenium (Se) is a dietary essential trace element in human nutrition and has been implicated in health, including cancer preventation. Methylselenocysteine (MeSeCys) is an effective chemopreventative compound. Astragalus sp. are known to accumulate Se with the majority of the selenoamino acids in the form of MeSeCys. The aim of this study was the cloning and charactherization of a cDNA encoding selenocysteine methyltransferase (SMT), the key enzyme responsible for MeSeCys formation, from Astragalus chrysochlorus using specific primers. Our results showed that Astragalus chrysochlorus SMT (AchSMT) is a 1020 bp (base pair) cDNA (GQ844862) with an open reading frame predicted to encode a 339 amino acid, 36.94 kDa protein. The predicted amino acid sequence of AchSMT (AEI53593) revealed 97% identity with A. bisulcatus selenocysteine methyltransferase (AbSMT). Bioinformatic analysis revealed that AchSMT lacks chloroplast and mitochondrial targeting sequences. The AchSMT possess a possible zinc-binding motif (GGCC) and a conserved cysteine residue upstream of this motif. AchSMT was expressed in Escherichia coli, then confirmed by DNA sequence analysis, western blotting and enzyme activity. Expression of AchSMT correlated with the presence of SMT enzyme activity in cell extracts. In A. chrysochlrous plants, analysis of AchSMT gene expression in response to selenate treatments showed that the AchSMT was constitutively expressed. The isolation of SMT gene will result in further studies for overproduction of valuable metabolite, methylselenocysteine.  

Pages 100-106 | Full Text PDF
Identification and expression analysis of TLPs as candidate genes promoting the responses to both biotic and abiotic stresses in wheat

Tahereh Deihimi1, Ali Niazi1*, Esmaeil Ebrahimie2,3*

1Institute of Biotechnology, College of Agriculture, Shiraz University, Shiraz, Iran
2Department of Crop Production & Plant Breeding, College of Agriculture, Shiraz University, Shiraz, Iran
3School of Molecular & Biomedical Science, The University of Adelaide, SA, Australia

Thaumatin like protein (TLPs) gene family have the ability to respond to both biotic and abiotic stresses. In this research, gene expression and bioinformatics analysis were applied to reveal the function of TLPs more evidently. Real Time PCR technique was used to compare TLP gene expression between susceptible and resistant cultivars of wheat under the Mycosphaerella graminicola inoculation. Our results indicated the early resistant-related response by TLP up-regulation within 3h after inoculation. In abiotic stress we observed induction of TLP gene expression during salt stress in susceptible cultivar. 50mM NaCl induced rapid induction of TLP within 3h, while 100mM NaCl conferred TLP up-regulation later at 72h. In silico chromosome walking analysis revealed that four loci can be the candidates for conferring tolerance to both biotic and abiotic stresses. In this study, a combination of Real Time PCR technique and bioinformatics tools suggested the possible role of TLP homologs in response to both biotic and abiotic stresses.

Pages 107-115 | Full Text PDF
Methyl jasmonate stimulates 20-hydroxyecdysone production in cell suspension cultures of Achyranthes bidentata

Qiu Jun Wang1, Li Ping Zheng2, Yang Hu Sima3, Hui Yan Yuan2, Jian Wen Wang1

1College of Pharmaceutical Sciences, Soochow University, Suzhou 215123, China
2College of Architecture and Urban Environment, Soochow University, Suzhou 215123, China
3School of Biology and Basic Medical Sciences, Soochow University, Suzhou 215123, China

20-Hydroxyecdysone (20E), one of major phytoecdysteroids (i.e. analogues of insect steroid hormones) in plants, has important agrochemical, medicinal and pharmaceutical uses. In order to develop a sustainable source of 20E, cell suspension cultures were established from shoot cultures of Achyranthes bidentata
. When cultivated in Murashige & Skoog medium supplemented with 1.5 mg/l 1-naphthlcetic acid (NAA) and 1.5 mg/l 6-benzyladenine (6-BA), A. bidentata cells in suspension culture grew rapidly, yielding 20E (5.4 mg/l) after 24 days. The increase of 20E was dependent on the growth stage of cell cultures as well as on the dose of methyl jasmonate (MeJA) applied. When cells of 18-day-old cultures were exposed to 0.6 mM MeJA for 6 days, it was found that total (intracellular and extracellular) 20E production reached the maximum yield 7.5 mg/l, a 2.6-fold increase over the control. This is the first report of 20E production in cell suspension cultures of A. bidentata.

Pages 116-120 | Full Text PDF
Wild rices of Eastern Indo-Gangetic plains of India constitute two sub-populations harbouring rich genetic diversity

Aparajita Singh1, Balwant Singh2, Kabita Panda2, Ved Prakash Rai1, Anil Kumar Singh1, Sheo Pratap Singh1, Sandeep Kumar Chouhan1, Vandna Rai2, Pawan Kumar Singh1*, Nagendra K. Singh2

Department of Genetics and Plant Breeding, Institute of Agricultural Sciences, Banaras Hindu University, Varanasi221005, India
2National Research Centre on Plant Biotechnology, Indian Agricultural Research Institute, New Delhi 110 012, India

Analysis of variability and genetic structure of wild rice populations is important for the management and conservation of the valuable genetic resources. To better understand the relationships among wild rice accessions, we analyzed a subset of germplasm collected from the eastern Indo-Gangetic plains of Uttar Pradesh and Bihar. Thirty-five wild rice accessions were characterized for fourteen morphological traits and genotyped using 25 genome wide SSR markers. The accessions showed significant phenotypic variation for all the traits analyzed. Analysis of SSR markers revealed average 2.4 alleles per locus with PIC values ranging from 0.51 to 0.90 with an average of 0.79. Cluster analysis and principal component analysis clearly demarcated the wild rice accessions into two main groups representing Oryza rufipogon and Oryza nivara. The high level of genetic diversity found in wild rices of this region suggests that it is a valuable resource that should be conserved for utilization in rice breeding programs.

Pages 121-127 | Full Text PDF
Intron targeted amplified polymorphism (ITAP), a new sequence related amplified polymorphism-based technique for generating molecular markers in higher plant species

Faqian Xiong1,2**, Junxian Liu3*, Ruichun Zhong1, Jing Jiang1, Zhuqiang Han1,2, Liangqiong He1, Zhong Li1, Xiumei Tang1, Ronghua Tang1,2*

1Cash Crops Research Institute, Guangxi Academy of Agricultural Sciences, Nanning, Guangxi 530007, China
2Guangxi Crop Genetic Improvement and Biotechnology Laboratory, Guangxi Academy of Agricultural Sciences, Nanning, Guangxi 530007, China
3Guangxi Sugarcane Research Institute, Guangxi Academy of Agricultural Sciences, Nanning, Guangxi 530007, China

Based on the principles of sequence related amplified polymorphism (SRAP), target region amplification polymorphism (TRAP) and conserved region amplification polymorphism (CoRAP), we developed a new molecular marker technique called intron targeted amplified polymorphism (ITAP). For ITAP, primer pairs were exploited to conduct the polymerase chain reaction (PCR). One primer targeting intron was retrieved from the SRAP technique. The other was designed from the 3’ widely distributed conserved intron-exon splice junction sequences. Amplification was carried out according to the procedure of the SRAP technique. The PCR products were resolved through standard agarose gel electrophoresis. To test this technique, we have applied it to fingerprint three plant species including banana (Musa spp.), longan (Dimocarpus longan Lour.), and cultivated peanut (Arachis hypogaea L.). Depending upon the different primer pairs, 3-11 bands were detected among these three plant species. Of these bands, 80%, 79.25%, and 45.30% were polymorphic in banana, longan, and cultivated peanut, respectively. Cluster analysis of ITAP markers was largely consistent with previous studies. Since primers were designed from the 3’ widely distributed intron-exon splice junction sequences in higher plant species assuming these primers should be universal across other plant species. This technique provides an alternative way to produce molecular markers for plant genotyping and fingerprinting.

Pages 128-134 | Full Text PDF
Enhanced expression of a thaumatin-like gene, involved in Pseudoperonospora cubensis and abiotic stresses, induced by DNA introgression from a wild relative, Cucumis hystrix

Hongjian Wan1, 2, Jinfeng Chen1*

State Key Laboratory of Crop Genetics and Germplasm Enhancement, College of Horticulture, Nanjing Agricultural University, Nanjing 210095, People’s Republic of China
2Institute of Vegetables, Zhejiang Academy of Agricultural Sciences, Hangzhou 310021, People’s Republic of China

Introgression of alien DNA from wild relatives into cultivated plant genomes by means of interspecific hybridization is a method which has been widely used in breeding. However, the effects of introgression on the patterns of gene transcription of the host genome have rarely been studied. In the present study, in order to understand the molecular defense response of an introgressed line (IL5211S) with introgression from a wild cucumber, Cucumis hystrix, a thaumatin-like gene, referred to as CsPR5
, was successfully isolated from IL5211S. The gene was 1504 bp in length with a putative open reading frame of 726 bp, encoding 241 amino acid residues. Open reading frame sequences of CsPR5 from IL5211S and backcross parents were identified. It was also observed that the expression of the CsPR5 gene was enhanced remarkably by DNA introgression of C. hystrix using real-time quantitative PCR (RT-qPCR). In addition, the expressions of CsPR5 in response to Pseudoperonospora cubensis and four different abiotic stresses stimuli (namely salicylic acid, methyl jasmonate, abscisic acid, and hydrogen peroxide) were further analyzed at different time points. These stimuli triggered a significant induction of CsPR5 within 72 h after treatment. These findings indicate that the expression of the CsPR5 gene was enhanced by alien DNA introgression from C. hystrix. This may play a role in the molecular defense of IL5211S against pathogen invasion, and aid in protecting against environmental stresses.

Pages 135-143 | Full Text PDF
Review article

Sex-linked markers in dioecious plants

Monika Milewicz, Jakub Sawicki

Department of Botany and Nature Protection, University of Warmia and Mazury in Olsztyn, Plac Lódzki 1, 10-727 Olsztyn, Poland

Dioecy is generally associated with sexual dimorphism, which can be noticed with the naked eye, but in plants the sex of an individual, at the early stages of development before flowering, is difficult to diagnose. The situation is more complicated in bryophytes, where  determination of sex in adult individuals can be problematic or even impossible. Here, molecular tools are helpful. This paper focuses on sex-linked molecular markers in plants. It presents plant species, for which sex-linked markers have been developed, indicates the most popular marker systems used in the research and tries to emphasise that some kind of  information about sex determination can be provided during development of sex-linked markers.

Pages 144-149 | Full Text PDF
In-silico prediction of an uncharacterized protein generated from heat responsive SSH library in wheat (Triticum aestivum L.)

Jasdeep Chatrath Padaria*, Deepesh Bhatt, Koushik Biswas, Gagandeep Singh, Rajkumar Raipuria

Biotechnology and Climate Change Laboratory, National Research Centre on Plant Biotechnology, PUSA Campus, New Delhi, 110012, India

Wheat is exposed to various abiotic stresses at different stages of its life cycle leading to severe decline in productivity. With rapid climate changes, high temperature stress is a major limitation to wheat production. Certain cultivars of wheat display a tolerant response to heat stress. Studies on differential expression in response to heat stress leads to identification of genes involved in molecular mechanism of thermo tolerance. Large-scale differential display analysis generates a large number of transcripts, of which a few are stress responsive whereas, many are of unknown or uncharacterized functional identity. The present study was done to identify a transcript of uncharacterized function obtained from heat responsive subtractive library generated from anthesis stage of thermo-tolerant wheat cv. Raj3765. Real time PCR analysis showed a four-fold increase in expression of the identified transcript at a stress of 37°C at the anthesis stage, indicating its role in facilitating the plant to cope the deleterious effects of high temperature at anthesis stage. Protparam tool analysis revealed that the leucine (Leu) is dominant amino acid present in the sequence, involving 15.5% of total amino acids. In-silico analysis revealed the existence of conserved domain region similar to leucine rich repeat (LRR) motif, an important DNA-binding domain. The presence of LRR motif in the protein predicted from the transcript under study indicates that this protein has a role as a signaling molecule involved in stress responses. Functional validation of the identified transcript in a model plant system shall confirm its role in heat stress tolerance.

Pages 150-156 | Full Text PDF | Supplementary data