Introducing sorghum DREB2 gene in maize (Zea mays L.) to improve drought and salinity tolerance.

Salinity and drought are significant abiotic stresses causing a considerable loss of seed and biomass yield in most commercial crops. Some of the most critical players in the abscisic acid pathway are drought responsive element binding (DREB) proteins that are a part of AP2/ethylene response factor transcription factors that bind to promoters of some family genes needed to be expressed under abiotic stresses. In this study, salt- and drought-tolerant maize plants were produced from immature maize embryos bombarded by the sorghum (Sorghum bicolor L.) DREB2 gene that is linked to hygromycin resistance (hpt) genes. The putative transgenic calli were transferred to an N6 medium containing 1 mg/L benzylaminopurine and 50 mg/L hygromycin. Regeneration was completed after 4 weeks on selective media under a 16/8 h light/dark condition at 25°C. Polymerase chain reaction (PCR) and reverse transcription-PCR approved the existence of upstream promoter (rd29a), hpt gene, and the expression of the DREB2 in transgenes up to the third generation (T2). It was found that the K+/Na+ ratio and the amount of proline as a screening indicator were higher in transgenic plants compared to their wild types. This result is a promising model to enhance maize tolerance to abiotic stressors.

Role of hydroxymethylglutaryl-coenzyme A (HMG-CoA) reductase 1 in nodule development of soybean.

Autoregulation of nodulation (AON) plays a central role in nodulation by inhibiting the formation of excess number of legume root nodules. In this study, the effect of hydroxymethylglutaryl-coenzyme A reductase 1 (GmHMGR1) gene expression on nodulation and the AON system in Glycine max (L.) Merr was investigated. Wild-type soybean (cultivar Bragg) and its near-isogenic supernodulating mutant (nitrate tolerant symbiotic) nts1007 were selected to identify the expression pattern of this gene in rootlets after inoculation by its microsymbiont Bradyrhizobium. For further analysis, the full length of GmHMGR1 and its promoter were cloned after amplification by inverse-PCR and BAC library screening. Also, we constructed an intron hairpin RNA interference (ihpRNAi) and a GmHMGR1 promoter: ß-glucuronidase fusion constructs, consequently for suppression of GmHMGR1 and histochemical analysis in transgenic soybean hairy roots induced by Agrobacterium rhizogenes strain K599. The GmHMGR1 gene was functional during the early stages of nodulation with the AON system having a negative effect on GmHMGR1 expression and nodule formation in wild-type rootlets. GmHMGR1 was particularly expressed in the developing phloem within the root, nodules and nodule lenticels. Expression of GmHMGR1 in transgenic hairy roots was suppressed by RNAi silencing approximately 85% as compared to empty vector controls. This suggests that the GmHMGR1 gene has an important role in triggering nodule formation as its suppression caused a reduction of nodule formation in nts mutant lines with a deficient AON system.

Metabolically engineered rice biomass and grain using genes associated with lipid pathway show high level of oil content.

Increasing lipid content using metabolic engineering methods in different parts of plant, including, leaves and stem can be considered as an innovative platform for achieving more energy and biofuel in more green habits. Two key enzymes, including, diacylglycerol acyltransferase (DGAT) and phospholipid:diacylglycerol acyltransferase (PDAT) catalyze the final step of TAG assembly. WRINKLED1 (WRI1) is one of the important transcription factors which regulate the fatty acid biosynthesis network and TAG accumulation by balancing carbon flux between carbohydrates and lipids. In addition, oleosin encoding gene (OLE) can protect TAGs from degradation by packing into oil bodies. In the current study, four important genes involved in TAG assembly and protection (i.e., AtDGAT1 and AtPDAT, AtWRI1, and AtOle) were overexpressed under a constitutive promoter in rice crop. TAG content of transgenic seeds increased significantly (P ≤ 0.05) by 26% in compared with those of control plants. Oleic and palmitic acid contents were significantly increased by 28% (from 32 to 41) and 27% (11 to 14) in seeds of transgenic plants in compared with controls, respectively. Our results showed an increase in the total grain and leaf oil contents by 70% (from 1.1 to 1.87%) and 22.5% (from 1.88 to 2.3%) in the metabolically engineered lines, respectively. This is the first report of transformation in rice for enhancing oil content and energy density in its seeds and vegetative parts. Such metabolically engineered crops would be cultivated for production much more oils in seeds and straw for food and biodiesel consequently.

Characterization of specific DNA markers at VRN-H1 and VRN-H2 loci for growth habit of barley genotypes.

Requirement of vernalization is an important factor which plays a crucial role in cereals to transit from vegetative to reproductive phase. There are three types of growth habit in barley: winter, spring and facultative types; in which spring type does not require vernalization but winter and facultative genotypes require full and partial vernalization, respectively. Combination of two loci, Vrn-h1 and Vrn-h2, regulates vernalization in barley genotypes. Specific DNA markers have been identified for growth habit regulator genes in barley. In this study, we examined 24 barley genotypes using specific primers for detecting Vrn-h1and Vrn-h2 loci. Results showed that among all differently suggested primer combinations, a few markers were precisely correlated with seasonal growth habit in barley. The specific markers of 600, 600 and 200 bps were verified for ZCCT-Ha, ZCCT-Hb and ZCCT-Hc loci, respectively. Our field growth habit test showed that cultivar Bahman as a winter growth habit, where all the others genotypes exhibited spring growth habit. By using specific primers for Vrn-h1, only Bahman cultivar produced 616 bp and 830 bp fragments and spring genotypes showed 574 bp or 616 bp alleles without any amplification for 830 bp fragments. Therefore, presence of 616 bp and 830 bp alleles together in each genotype can be considered as an informative marker for winter growth habit in barley. These informative markers can be used easily in barley breeding programmes for detection of growth habit types in the seedling stage.

Chemical Compositions, Somatic Embryogenesis, and Somaclonal Variation in Cumin.

This is the first report evaluating the relationship between the chemical compositions of cumin seeds (based on the analysis of the content of catalase, ascorbate peroxidase, proline, protein, terpenic compounds, alcohol/phenols, aldehydes, and epoxides) and the induction efficiency of somatic embryogenesis in two Iranian superior cumin landraces (Golestan and North Khorasan). Cotyledons isolated from Golestan landrace seeds cultivated on MS medium supplemented with 0.1 mg/L kinetin proved to be the best primary explant for the induction of somatic embryogenesis as well as the regeneration of the whole plantlet. Results indicated that different developmental stages of somatic embryos were simultaneously observed on a callus with embryogenic potential. The high content of catalase, ascorbate peroxidase, proline, and terpenic hydrocarbons and low content of alcoholic and phenolic compositions had a stimulatory effect on somatic embryogenesis. Band patterns of RAPD markers in regenerated plants were different from those of the mother plants. This may be related to somaclonal variations or pollination system of cumin. Generally, measurement of chemical compositions can be used as a marker for evaluating the occurrence of somatic embryogenesis in cumin. Also, somaclonal variations of regenerated plants can be applied by the plant breeders in breeding programs.

Marker-assisted selection of high molecular weight glutenin alleles related to bread-making quality in Iranian common wheat (Triticum aestivum L.).

Bread-making quality in hexaploid wheats is a complex trait. It has been shown that the amount and composition of protein can influence dough rheological properties. The high-molecular-weight (HMW) glutenins are encoded by a complex locus, Glu-1, on the long arm of group-1 homoeologus chromosome of the A, B and D genomes. In this work we used PCR-based DNA markers as a substitution tool to distinguish wheat bread-making quality. We detected PCR-based DNA markers for coding sequence of Glu-A1x, Glu-B1x and Glu-D1x to be 2300 bp, 2400 bp and 2500 bp respectively. DNA markers related to coding sequence of Glu-A1y, Glu-B1y and Glu-D1y were; 1800 bp, 2100 bp and 1950 bp, however, the repetitive region of their coding sequence were shown to be about 1300 bp, 1500 bp and 1600 bp. The results demonstrate that the size variation was due to different lengths of the central repetitive domain. Good or poor bread-making quality in wheat is associated with two allelic pairs of Glu-D1, designated 1Dx5-1Dy10 and 1Dx2-1Dy12. The 1Bx7 allele has moderate-to-good quality score. The specific DNA markers, of 450 bp, 576 bp, 612 bp and 2400 bp respectively were characterized for 1Dx5, 1Dy10, 1Dy12 and 1Bx7 alleles. These markers are very important in screening of wheat for bread-making quality.

Bioinformatics study of the 3-hydroxy-3-methylglotaryl-coenzyme A reductase (HMGR) gene in Gramineae.

Isoprenoids or terpenoids are synthesized by two important units' including dimethylallyl diphosphate and isopentenyl diphosphate (IPP). Plants use two different methods for formation of IPP, which is a cytosolic and a plastidial method. The 3-hydroxy-3-methylglutaryl-CoA reductase (HMGR, EC 1.1.1.34) catalyzes the conversion of HMG-CoA to mevalonate, which is the first stage in the cytosolic pathway for biosynthesis of isoprenoid in plants. In this study, a total of fifty HMGR protein sequences from Gramineae and three animal samples including human, mouse and fruit fly were aligned and analyzed by computational tools to predict the protein properties, such as molecular mass, pI, signal peptide, transmembrane and conserved domains, secondary and spatial structures. Sequence comparison analysis revealed that there is high identity between plants and animals. Three catalytic regions including L domain, N domain and S domain were detected by structural modeling of HMGR. The tertiary structure model of Oryza sativa HMGR (Accession Number: NP_001063541) was further checked by PROCHECK algorithm, and showed that 90.3 % of the amino acid residues were located in the most favored regions in Ramachandran plot, indicating that the simulated three-dimensional structure was reliable. Phylogenetic analysis indicated that there is a relationship among species of Gramineae and other organisms. According to these results, HMGRs should be derived from a common ancestor.

Allelic variations in Glu-1 and Glu-3 loci of historical and modern Iranian bread wheat (Triticum aestivum L.) cultivars.

Proline and glutamine-rich wheat seed endosperm proteins are collectively referred to as prolamins. They are comprised of HMW-GSs, LMW-GSs and gliadins. HMW-GSs are major determinants of gluten elasticity and LMW-GSs considerably affect dough extensibility and maximum dough resistance. The inheritance of glutenin subunits follows Mendelian genetics with multiple alleles in each locus. Identification of the banding patterns of glutenin subunits could be used as an estimate for screening high quality wheat germplasm. Here, by means of a two-step 1D-SDS-PAGE procedure, we identified the allelic variations in high and low-molecular-weight glutenin subunits in 65 hexaploid wheat (Triticum aestivum L.) cultivars representing a historical trend in the cultivars introduced or released in Iran from the years 1940 to 1990. Distinct alleles 17 and 19 were detected for Glu-1 and Glu-3 loci, respectively. The allelic frequencies at the Glu-1 loci demonstrated unimodal distributions. At Glu-A1, Glu-B1 and Glu-D1, we found that the most frequent alleles were the null, 7 + 8, 2 + 12 alleles, respectively, in Iranian wheat cultivars. In contrast, Glu-3 loci showed bimodal or trimodal distributions. At Glu-A3, the most frequent alleles were c and e. At Glu-B3 the most frequent alleles were a, b and c. At Glu-D3 locus, the alleles b and a, were the most and the second most frequent alleles in Iranian wheat cultivars. This led to a significantly higher Nei coefficient of genetic variations in Glu-3 loci (0.756) as compared to Glu-1 loci (0.547). At Glu-3 loci, we observed relatively high quality alleles in Glu-A3 and Glu-D3 loci and low quality alleles at Glu-B3 locus.