Genotyping by sequencing-based linkage map construction and identification of quantitative trait loci for yield-related traits and oil content in Jatropha (Jatropha curcas L.).

BACKGROUND: Jatropha (Jatropha curcas L.) has been considered as a potential bioenergy crop and its genetic improvement is essential for higher seed yield and oil content which has been hampered due to lack of desirable molecular markers. METHODS AND RESULTS: An F2 population was created using an intraspecific cross involving a Central American line RJCA9 and an Asiatic species RJCS-9 to develop a dense genetic map and for Quantitative trait loci (QTL) identification. The genotyping-by-sequencing (GBS) approach was used to genotype the mapping population of 136 F2 individuals along with the two parental lines for classification of the genotypes based on single nucleotide polymorphism (SNPs). NextSeq 2500 sequencing technology provided a total of 517.23 million clean reads, with an average of ~ 3.8 million reads per sample. We analysed 411 SNP markers and developed 11 linkage groups. The total length of the genetic map was 4092.3 cM with an average marker interval of 10.04 cM. We have identified a total of 83 QTLs for various yield and oil content governing traits. The percentage of phenotypic variation (PV) was found to be in the range of 8.81 to 65.31%, and a QTL showed the maximum PV of 65.3% for a total seed number on the 6th linkage group (LG). CONCLUSIONS: The QTLs detected in this study for various phenotypic traits will lay down the path for marker-assisted breeding in the future and cloning of genes that are responsible for phenotypic variation.

Development of genome wide transposable elements based repeat junction markers in Jatropha (Jatropha curcas L.).

Jatropha curcas is a potential biodiesel crop and a highly adaptable species to various agro-climatic conditions. In this study, we have utilized transposable elements' (TE) repeat junctions (RJs) which are an important constituent of the genome, used to form a genome-wide molecular marker platform owing to its use in genomic studies of plants. We screened our previously generated Jatropha hybrid genome assembly of size 265 Mbp using RJPrimers pipeline software and identified a total of 1274 TE junctions. For the predicted RJs, we designed 2868 polymerase chain reaction (PCR) based RJ markers (RJMs) flanking the junction regions. In addition to marker design, the identified RJs were utilized to detect 225,517 TEs across the genome. The different types of transposable repeat elements mainly were scattered into Retro, LTR, Copia and Gypsy categories. The efficacy of the designed markers was tested by utilizing a subset of RJMs selected randomly. We have validated 96 randomly selected RJ primers in a group of 32 J. curcas genotypes and more than 90% of the markers effectively intensified as amplicons. Of these, 10 primers were shown to be polymorphic in estimating genetic diversity among the 32 Jatropha lines. UPGMA cluster analysis revealed the formation of two clusters such as A and B exhibiting 85.5% and 87% similarity coefficient respectively. The various RJMs identified in this study could be utilized as a significant asset in Jatropha functional genomics including genome determination, mapping and marker-assisted selection.

Synthesis, crystal structure, thermal stability and biological study of bis{(2-methoxy-6-[(E)-(propylimino)methyl]phenolato}nickel(II) complex.

A Schiff base complex of nickel, bis{(2-methoxy-6-[(E)-(propylimino)methyl]phenolato}nickel(II) was synthesised by condensing bis(2-hydroxy-3-methoxybenzaldehyde) nickel (II) and n-propylamine in methanolic medium. Single crystal X-ray diffraction analysis of the complex revealed it to possess planar geometry with a monoclinic crystal system. The non-isothermal TG/DTA runs on this complex in a high purity (99.99 %) nitrogen environment at atmospheric pressure confirmed the absence of any coordinated water. A sharp endotherm in its DTA shows a melting temperature range of 168-171 °C. It is thermally stable up to 243 °C and decomposes in two steps, yielding NiO and carbon as residue. In addition to the methoxy group (-OCH3), infrared analysis (IR) confirmed the presence of the characteristic azomethine group (-C[bond, double bond]N-) which is also responsible for the biological action. It was further analysed by elemental analyser (C, H, N), 1H and 13C NMR as well as mass spectrometry. It showed considerable antibacterial activity towards Escherichia coli and Staphylococcus aureus when the concentration exceeds 200 µg/ml. The antifungal study shows significant inhibition with the antifungal drug imidazole as a positive control (PC). Small values of MIC, MBC/MIC indicate a lesser quantity of complex is required to inhibit the growth of micro-organisms.

Efficient protocols for in vitro regeneration of Pennisetum glaucum (L) Br.

A system was developed for in vitro regeneration of Pennisetum glaucum through organogenesis and somatic embryogenesis. Mature embryo and leaf base explants of Pennisetum glaucum (L) Br. cv HH B60 (Poaceae) were cultured on Murashige and Skoog agar medium supplemented with 11.3 microM of 2,4-D for callus induction. Embryogenic calli were induced within eight weeks. Percentage of callus induction and somatic embryogenesis was significantly higher in mature embryo than leaf base explants. Maximum shoot regeneration was obtained via organogenesis on MS medium supplemented with 4.43 microM of BAP and 4.64 microM of kinetin from the calli of both the explants. The frequency of plant regeneration through somatic embryogenesis was comparatively lower than organogenesis. Regeneration frequency was higher in mature embryo explants than leaf base explants. The shoots regenerated via organogenesis were elongated and rooted efficiently on MS medium supplemented with IBA (0.49 microM). The rooted plantlets were hardened and transferred to soil.

Influence of growth regulators and explant type on in vitro shoot propagation and rooting of red sandal wood (Pterocarpus santalinus L.).

In vitro shoot regeneration in Pterocarpus santalinus L. was achieved when detached cotyledons from in vitro germinated seedlings were cultured on MS medium containing NAA (0.1 mg/L), BA (1 mg/L) and kinetin (1 mg/L). The regenerated shoots rooted on 1/4 strength MS medium with IAA (1 mg/L) and the fully developed plantlets were successfully established in the soil.

Regeneration of transgenic plants from two indica rice (Oryza sativa L.) cultivars using shoot apex explants.

We have established a reproducible procedure for transformation of shoot apices and regeneration of transgenic plants for two indica rice cultivars, white ponni (WP) and Pusa Basmathi 1 (PB 1). Four-day-old shoot apex explants were transformed by cocultivation with Agrobacterium tumefaciens strain EHA 101 harbouring a binary plasmid pRIT1. The vector contained an improved hygromycin phosphotransferase (hpt) gene for hygromycin resistance driven by actin 1 promoter and the reporter gene beta-glucuronidase intron (INT-GUS) controlled by CaMV 35S promoter. Rice shoots were induced on media containing 0.1 mg/l napthalene acetic acid (NAA), 1.0 mg/l kinetin (kn), 1.0 mg/l N(6)-benzyleaminopurin (BAP), 300 mg/l casaminoacid, 500 mg/l proline, 50 mg/l hygromycin and 500 mg/l cefotaxime. Transgenic plants were raised in pots and seeds were collected. Histochemical and polymerase chain reaction (PCR) analyses of field established transgenic rice plants and their offsprings confirmed the presence of GUS gene. Integration of T-DNA into the genome of putative transgenics was further confirmed by southern analysis. The transformation efficiency of WP was found to be ranging from 5.6 to 6.2% whereas in the case of PB1, it was from 7 to 8%. Progeny analysis of these plants showed a pattern of classical Mendelian inheritance for both hpt and GUS gene.