Transcriptomic analysis of salt stress induced chlorophyll biosynthesis-related genes in photoheterotrophic Arabidopsis thaliana calli.

In order to investigate the salt stress induced chlorophyll biosynthesis-related genes in photoheterotrophic cultures, we performed RNA-Seq analysis on A. thaliana calli exposed to 100 mM NaCl on MS medium containing 0.5 mg/L 2,4-D 30 days. Four different conditions of samples were sequenced on Illumina HiSeq Platform in total and generated about 4.49 Gb per sample. The average genome and gene mapping rates were 93.52% and 90.78%, respectively. According to expression profile analysis, some DEGs demonstrated altered related to chlorophyll pigment metabolism. According to analysis, green callus color of photoheterotrophic calli were mainly connected with the induction of LHCB4.3 light harvesting complex photosystem II (Gene ID:818599), AT1G49975 photosystem I reaction center subunit N (Gene ID: 841421), PAM68 PAM68-like protein (DUF3464) (Gene ID: 2745715) and AT3G63540 thylakoid lumenal protein (Mog1/PsbP/DUF1795-like photosystem II reaction center PsbP family protein)(Gene ID: 7922413) genes. Furthermore, 8 DEGs were randomly selected to validate the transcriptome profiles via qPCR. These results will provide a foundation for further studies aimed at giving photosynthetic properties to in vitro plant cultures.

Genetic, Morphological, and Biochemical Diversity of Argan Tree (Argania spinosa L.) (Sapotaceae) in Tunisia.

Argan trees are normally endemic to Morocco and Algeria, but hundreds of argan trees exist in Tunisia, some introduced from Morocco and some from unknown origins. The aim of the present study was to evaluate the genetic, morphological, and biochemical diversity of the argan trees in Tunisia. In this study, we used morphometric data collected from vegetative tissue, as well as pomological characteristics related to fruits, stones, and kernels. Genetic variation in 60 trees of Tunisian Argania spinosa L. was estimated using inter-simple sequence repeats (ISSRs). Mutation screening and genotyping by high-resolution melting (HRM) was performed to detect delta-6-desaturase (D6D) variants in the tested individuals, and finally fatty acid analysis of argan leaves with gas chromatography (GC) was performed. The plant materials used in this study originated from four different sites in Tunisia. Analysis of morphological characteristics showed large variability both within and between the studied collections. The analysis of ISSR polymorphisms gave information about the diversity within and between populations. HRM analysis showed that all 60 argan individuals were grouped into 10 different categories. The results of the gas chromatography analysis showed that the presence of omega-3 fatty acids EPA and DHA was noticeable in some argan leaves.

Barley Genes as Tools to Confer Abiotic Stress Tolerance in Crops.

Barley is one of the oldest cultivated crops in the world with a high adaptive capacity. The natural tolerance of barley to stress has led to increasing interest in identification of stress responsive genes through small/large-scale omics studies, comparative genomics, and overexpression of some of these genes by genetic transformation. Two major categories of proteins involved in stress tolerance are transcription factors (TFs) responsible from the re-programming of the metabolism in stress environment, and genes encoding Late Embryogenesis Abundant (LEA) proteins, antioxidant enzymes, osmolytes, and transporters. Constitutive overexpression of several barley TFs, such as C-repeat binding factors (HvCBF4), dehydration-responsive element-binding factors (HvDREB1), and WRKYs (HvWRKY38), in transgenic plants resulted in higher tolerance to drought and salinity, possibly by effectively altering the expression levels of stress tolerance genes due to their higher DNA binding affinity. Na(+)/H(+) antiporters, channel proteins, and lipid transporters can also be the strong candidates for engineering plants for tolerance to salinity and low temperatures.

Genetic diversity at the Dhn3 locus in Turkish Hordeum spontaneum populations with comparative structural analyses.

We analysed Hordeum spontaneum accessions from 21 different locations to understand the genetic diversity of HsDhn3 alleles and effects of single base mutations on the intrinsically disordered structure of the resulting polypeptide (HsDHN3). HsDHN3 was found to be YSK2-type with a low-frequency 6-aa deletion in the beginning of Exon 1. There is relatively high diversity in the intron region of HsDhn3 compared to the two exon regions. We have found subtle differences in K segments led to changes in amino acids chemical properties. Predictions for protein interaction profiles suggest the presence of a protein-binding site in HsDHN3 that coincides with the K1 segment. Comparison of DHN3 to closely related cereals showed that all of them contain a nuclear localization signal sequence flanking to the K1 segment and a novel conserved region located between the S and K1 segments [E(D/T)DGMGGR]. We found that H. vulgare, H. spontaneum, and Triticum urartu DHN3s have a greater number of phosphorylation sites for protein kinase C than other cereal species, which may be related to stress adaptation. Our results show that the nature and extent of mutations in the conserved segments of K1 and K2 are likely to be key factors in protection of cells.

Enhancing T-DNA Transfer Efficiency in Barley (Hordeum vulgare L.) Cells Using Extracellular Cellulose and Lectin.

A major limitation of transforming barley tissues by Agrobacterium tumefaciens is the low frequency of T-DNA transfer due to recalcitrance of barley as a host. The effect of extracellular cellulose and lectin on Agrobacterium transformation efficiency was investigated in this study. Barley callus cultures were transformed with the AGL1 strain containing the vector pBI121 in the presence of 10 mg mL(-1) cellulose or 0.001, 0.05 and 0.1 mg mL(-1) lectin. Addition of cellulose significantly (P ≤ 0.05) increased the number of GUS spots by 50 % compared to standard conditions in the presence of only 200 µM acetosyringone (AS). Frequency of G418-resistant aggregates on the surfaces of callus cultures was 29 and 71.5 %, following AS and AS + cellulose treatments, respectively, after 4 weeks of selection. Presence of 0.05 or 0.1 mg mL(-1) lectin also increased the number of GUS spots and frequency of G418-resistant cells in the selection period, but the increase in blue spots was not significant. We examined the effect of lectin and cellulose on bacterial attachment to callus tissues. Both cellulose and lectin were found to have a significant positive effect on the numbers of bacteria attached to barley callus. Epifluorescence microscopy revealed that Agrobacterium cells had accumulated in the scaffolds of irregular fibrous cellulose with a mean particle size of 200 µm. Expression of nptII in transformed callus lines confirmed the stable transformation of the gene. Our study showed for the first time the binding of Agrobacterium cells to fibrous cellulose and also demonstrated how polysaccharides and glycoproteins can be used to improve T-DNA transfer in monocotyledon transformation procedures.

Analysis of expressed sequence tags from cDNA library of Fusarium culmorum infected barley (Hordeum vulgare L.) roots.

Fusarium culmorum is one of the most common and globally important causal agent of root and crown rot diseases of cereals. These diseases cause grain yield loss and reduced grain quality in barley. In this study, we have analyzed an expressed sequence tag (EST) database derived from F. culmorum infected barley root tissues available at the National Center for Biotechnology Information (NCBI). The 2294 sequences were assembled into 1619 non-redundant sequences consisting of 359 contigs and 1260 singletons using the program CAP3. BLASTX analysis for these sequences was conducted in order to find similar sequences in all databases. Gene Ontology search, enzyme search, KEGG mapping and InterProScan search were done using Blast2GO 3.0.7 tool. By BLASTX analysis, 41.7%, 7.7%, 3.2% and 47.4% of ESTs were categorized as annotated, unannotated, not mapping and without blast hits, respectively. BLASTX analysis revealed that the majority of top hits were barley proteins (43.5%). Based on Gene Ontology classification, 38.3%, 31.3%, and 16% of ESTs were assigned to molecular function, biological process, and cellular component GO terms, respectively. Most abundant GO terms were as follows: 157 sequences were related to response to stress (biological process), 207 sequences were related to ion binding (molecular function), and 160 sequences were related to plastid (cellular component). Furthermore, based on KEGG mapping, 369 sequences could be assigned to 264 enzymes and 83 different KEGG pathways. According to Enzyme Commission (EC) distribution; 94 sequences were transferases (EC2) while 70 sequences were hydrolases (EC3).