Genome-wide identification and expression profile analysis of nuclear factor Y family genes in Sorghum bicolor L. (Moench)

Members of the plant Heme Activator Protein (HAP) or NUCLEAR FACTOR Y (NF-Y) are trimeric transcription factor complexes composed of the NF-YA, NF-YB and NF-YC subfamilies. They bind to the CCAAT box in the promoter regions of the target genes and regulate gene expressions. Plant NF-Ys were reported to be involved in adaptation to several abiotic stresses as well as in development. In silico analysis of Sorghum bicolor genome resulted in the identification of a total of 42 NF-Y genes, among which 8 code for the SbNF-YA, 19 for SbNF-YB and 15 for the SbNF-YC subunits. Analysis was also performed to characterize gene structures, chromosomal distribution, duplication status, protein subcellular localizations, conserved motifs, ancestral protein sequences, miRNAs and phylogenetic tree construction. Phylogenetic relationships and ortholog predictions displayed that sorghum has additional NF-YB genes with unknown functions in comparison with Arabidopsis. Analysis of promoters revealed that they harbour many stress-related cis-elements like ABRE and HSE, but surprisingly, DRE and MYB elements were not detected in any of the subfamilies. SbNF-YA1, 2, and 6 were found upregulated under 200 mM salt and 200 mM mannitol stresses. While NF-YA7 appeared associated with high temperature (40˚C) stress, NF-YA8 was triggered by both cold (4˚C) and high temperature stresses. Among NF-YB genes, 7, 12, 15, and 16 were induced under multiple stress conditions such as salt, mannitol, ABA, cold and high temperatures. Likewise, NF-YC 6, 11, 12, 14, and 15 were enhanced significantly in a tissue specific manner under multiple abiotic stress conditions. Majority of the mannitol (drought)-inducible genes were also induced by salt, high temperature stresses and ABA. Few of the high temperature stress-induced genes are also induced by cold stress (NF-YA2, 4, 6, 8, NF-YB2, 7, 10, 11, 12, 14, 16, 17, NF-YC4, 6, 12, and 13) thus suggesting a cross talk among them. This work paves the way for investigating the roles of diverse sorghum NF-Y proteins during abiotic stress responses and provides an insight into the evolution of diverse NF-Y members

An update and perspectives on the use of promoters in plant genetic engineering

Genetically engineered plants have varied applications in agriculture for enhancing the values of food and feed. Genetic engineering aims to introduce selected genetic regions with desirable traits into target plants for both spatial and temporal expressions. Promoters are the key elements responsible for regulating gene expressions by modulating the transcription factors (TFs) through recognition of RNA polymerases. Based on their recognition and expression, RNA polymerases were categorized into RNA pol II and pol III promoters. Promoter activity and specificity are the two prime parameters in regulating the transgene expression. Since the use of constitutive promoters like Cauliflower mosaic virus (CaMV) 35S may lead to adverse effects on nontarget organisms or ecosystem, inducible/tissue specific promoters and/or the RNA pol III promoters provide myriad opportunities for gene expressions with controlled regulation and with minimum adverse effects. Besides their role in transgene expression, their influence in synthetic biology and genome editing are also discussed. This review provides an update on the importance, current prospects, and insight into the advantages and disadvantages of promoters reported thus far would help to utilize them in the endeavour to develop nutritionally and agronomically improved transgenic crops for commercialization

Root and Leaf Anatomy, Ion Accumulation, and Transcriptome Pattern under Salt Stress Conditions in Contrasting Genotypes of Sorghum bicolor

Roots from salt-susceptible ICSR-56 (SS) sorghum plants display metaxylem elements with thin cell walls and large diameter. On the other hand, roots with thick, lignified cell walls in the hypodermis and endodermis were noticed in salt-tolerant CSV-15 (ST) sorghum plants. The secondary wall thickness and number of lignified cells in the hypodermis have increased with the treatment of sodium chloride stress to the plants (STN). Lignin distribution in the secondary cell wall of sclerenchymatous cells beneath the lower epidermis was higher in ST leaves compared to the SS genotype. Casparian thickenings with homogenous lignin distribution were observed in STN roots, but inhomogeneous distribution was evident in SS seedlings treated with sodium chloride (SSN). Higher accumulation of K+ and lower Na+ levels were noticed in ST compared to the SS genotype. To identify the differentially expressed genes among SS and ST genotypes, transcriptomic analysis was carried out. Both the genotypes were exposed to 200 mM sodium chloride stress for 24 h and used for analysis. We obtained 70 and 162 differentially expressed genes (DEGs) exclusive to SS and SSN and 112 and 26 DEGs exclusive to ST and STN, respectively. Kyoto Encyclopaedia of Genes and Genomes (KEGG) and Gene Ontology (GO) enrichment analysis unlocked the changes in metabolic pathways in response to salt stress. qRT-PCR was performed to validate 20 DEGs in each SSN and STN sample, which confirms the transcriptomic results. These results surmise that anatomical changes and higher K+/Na+ ratios are essential for mitigating salt stress in sorghum apart from the genes that are differentially up- and downregulated in contrasting genotypes

Functional characterization of the promoter of pearl millet heat shock protein 10 (PgHsp10) in response to abiotic stresses in transgenic tobacco plants

In the present study, the promoter region of the pearlmillet heat shock protein 10 (PgHsp10) genewas cloned and characterized. The PgHsp10 promoter (PgHsp10pro) sequence region has all the cis-motifs required for tissue and abiotic stress inducibility. The complete PgHsp10pro (PgHsp10PC) region and a series of 5′ truncations of PgHsp10 (PgHsp10D1 and PgHsp10D2) and an antisense formof PgHsp10pro (PgHsp10AS)were cloned into a plant expression vector (pMDC164) through gateway cloning. All four constructs were separately transformed into tobacco through Agrobacterium-mediated genetic transformation, and PCR-confirmed transgenic plants progressed to T1 and T2 generations. The T2 transgenic tobacco plants comprising all PgHsp10pro fragments were used for GUS histochemical and qRT-PCR assays in different tissues under control and abiotic stresses. The PgHsp10PC pro expression was specific to stemand seedlings under control conditions. Under different abiotic stresses, particularly heat stress, PgHsp10PCpro had relatively higher activity than PgHsp10D1pro, PgHsp10D2pro and PgHsp10ASpro. PgHsp10pro from a stress resilient crop like pearl millet responds positively to a range of abiotic stresses, in particular heat, when expressed in heterologous plant systems such as tobacco. Hence, PgHsp10pro appears to be a potential promoter candidate for developing heat and drought stress-tolerant crop plants

Investigation of beam transmission in A 9SDH-2 3.0 MV NEC pelletron tandem accelerator

Electrostatic tandem accelerators are widely used to accelerate ions for experiments in materials science such as high energy ion implantation, materials modification, and analyses. Many applications require high beam current as well as high beam brightness at the target; thus, maximizing the beam transmission through such electrostatic accelerators becomes important. The Ion Beam Modification and Analysis Laboratory (IBMAL) at University of North Texas is equipped with four accelerators, one of which is a 9SDH-2 3.0 MV National Electrostatic Corporation (NEC) Pelletron® tandem accelerator. The tandem accelerator is equipped with three ion sources: one radio frequency-He ion source (Alphatross) and two ion sources of Cs-sputter type, the SNICS II (Source of Negative Ions by Cesium Sputtering) and a Cs-sputter source for trace-element accelerator based mass spectrometry. This work presents a detailed study of the beam transmission of hydrogen, silicon, and silver ions through the accelerator using the SNICS ion source with injection energies ranging from 20 keV to 70 keV. The beam transmission is quantified for three different terminal voltages: 1.5 MV, 2.0 MV and 2.5 MV. For a given terminal voltage, it has been found that beam transmission is strongly dependent on the ion source injector potential. Details of experiments and data analysis are presented

Overexpression of RNA ‐binding bacterial chaperones in rice leads to stay‐green phenotype, improved yield and tolerance to salt and drought stresses

Genes encoding bacterial cold shock proteins A (CspA, 213 bp) and B (CspB, 216 bp) were isolated from Escherichia coli strain K12, which showed 100% homology with gene sequences isolated from other bacterial species. In silico domain, analysis showed eukaryotic conserved cold shock domain (CSD) and ribonuclease-binding domain (RBD) indicating that they bind to RNA and are involved in temperature stress tolerance. Overexpression of these two genes in E. coli resulted in higher growth in presence of 200 mM NaCl and 300 mM mannitol. Western blot confirmed the translational products of the two genes. Seedlings of indica rice were transformed with Agrobacterium tumefaciens containing pCAMBIA1301 CspA and CspB genes. Transgene integration was confirmed by β-glucuronidase (GUS) histochemical assay, polymerase chain reaction (PCR) amplification, and gene copy number by Southern blotting. Chlorophyll, proline, Na+, and K+ contents were higher in transgenics exposed to 150 mM NaCl and drought (imposed by withholding water) stresses during floral initiation stage. Catalase (CAT), superoxide dismutase (SOD), and guaiacol peroxidase (GPX) activities increased, while malondialdehyde (MDA) content was low in transgenics. Transgenics displayed increased root, shoot, and panicle lengths, root dry mass, and a distinct stay-green (SGR) phenotype. Higher transcript levels of CspA, CspB, SGR, chlorophyllase, isopentenyl adenine transferase 1 (IPT1), 9-cis-epoxycarotenoid dioxygenase (NCED), SOD, and sirtuin 1 (SIRT1) genes were observed in transgenics compared to wild type plants (WT) under multiple stresses. Present work indicates that bacterial chaperone proteins are capable of imparting SGR phenotype, salt and drought stress tolerance alongside grain improvement

An overview of the facilities, activities, and developments at the University of North Texas Ion Beam Modification and Analysis Laboratory (IBMAL)

The Ion Beam Modification and Analysis Laboratory (IBMAL) at the University of North Texas includes several accelerator facilities with capabilities of producing a variety of ion beams from tens of keV to several MeV in energy. The four accelerators are used for research, graduate and undergraduate education, and industrial applications. The NEC 3MV Pelletron tandem accelerator has three ion sources for negative ions: He Alphatross and two different SNICS-type sputter ion sources. Presently, the tandem accelerator has four high-energy beam transport lines and one low-energy beam transport line directly taken from the negative ion sources for different research experiments. For the low-energy beam line, the ion energy can be varied from ∼20 to 80 keV for ion implantation/modification of materials. The four post-acceleration beam lines include a heavy-ion nuclear microprobe; multi-purpose PIXE, RBS, ERD, NRA, and broad-beam single-event upset; high-energy ion implantation line; and trace-element accelerator mass spectrometry. The NEC 3MV single-ended Pelletron accelerator has an RF ion source mainly for hydrogen, helium and heavier inert gases. We recently installed a capacitive liner to the terminal potential stabilization system for high terminal voltage stability and high-resolution microprobe analysis. The accelerator serves a beam line for standard RBS and RBS/C. Another beamline for high energy focused ion beam application using a magnetic quadrupole lens system is currently under construction. This beam line will also serve for developmental work on an electrostatic lens system. The third accelerator is a 200 kV Cockcroft-Walton accelerator with an RF ion source. The fourth accelerator is a 2.5 MV Van de Graaff accelerator, which was in operation for last several decades is currently planned to be used mainly for educational purpose. Research projects that will be briefly discussed include materials synthesis/modification for photonic, electronic, and magnetic applications, surface sputtering and micro-fabrication of materials, development of high-energy ion microprobe systems, and educational and outreach activities