Explore the vast range of titles available.

Sesamum indicum is an important oilseed crop with beneficiary nutrients such as essential fatty acids, proteins, and lignan. S. indicum crop improvement using conventional breeding methods was disastrous. Moreover, no reproducible regeneration and transformation protocol for large-scale generation of independent primary transformed plant lines of S. indicum virtually exists. Therefore, in the present study, a reproducible regeneration and transformation method was developed for S. indicum. Direct shoot regeneration (94.44%) was achieved using plumule tips as an explant source and when maintained on regeneration medium (BM11) an amalgamation of half the strength of MS macronutrients and B5 macronutrients together with MS micronutrients and vitamins. Maximum number of shoots (~ 16–19 shoots/explant) was obtained on BM11 medium supplemented with TDZ (1 mg/L) and BAP (0.1 mg/L). High root induction efficiency (~ 97%) was observed on RM4 medium consisted of half MS macronutrients, SH micronutrients and vitamins. The developed protocol was found effective for other commercially valuable genotypes such as Co 1, Phule til and Tilottama as well. Successful regeneration was subsequently extended to transformation of reporter gene (gfp: green fluorescent protein) as revealed by molecular analyses includes Southern hybridization, northern hybridization, real time PCR and histological study of the transformants developed in the present study. Based on the Southern analysis the calculated transformation frequency was found to be 1.33%. The presently developed regeneration and transformation system might facilitate any desired improvement of sesame crop.Key messageThe present study demonstrated an improved regeneration and transformation method for S. indicum, which is genotype independent and can undergo desired modifications.

Background The application of hybrid rice technology has significantly increased global rice production during the last three decades. Approximately 90% of the commercially cultivated rice hybrids have been derived through three-line breeding involving the use of WA-CMS lines. It is believed that during the 21 st century, hybrid rice technology will make significant contributions to ensure global food security. This study examined the poorly understood molecular basis of the WA-CMS system in rice. Results RFLPs were detected for atp6 and orfB genes in sterile and fertile rice lines, with one copy of each in the mt-genome. The RNA profile was identical in both lines for atp6 , but an additional longer orfB transcript was identified in sterile lines. 5' RACE analysis of the long orfB transcript revealed it was 370 bp longer than the normal transcript, with no indication it was chimeric when compared to the genomic DNA sequence. cDNA clones of the longer orfB transcript in sterile lines were sequenced and the transcript was determined unedited. Sterile lines were crossed with the restorer and maintainer lines, and fertile and sterile F 1 hybrids were respectively generated. Both hybrids contained two types of orfB transcripts. However, the long transcript underwent editing in the fertile F 1 hybrids and remained unedited in the sterile lines. Additionally, the editing of the 1.1 kb orfB transcript co-segregated with fertility restoring alleles in a segregating population of F 2 progeny; and the presence of unedited long orfB transcripts was detected in the sterile plants from the F 2 segregating population. Conclusion This study helped to assign plausible operative factors responsible for male-sterility in the WA cytoplasm of rice. A new point of departure to dissect the mechanisms governing the CMS-WA system in rice has been identified, which can be applied to further harness the opportunities afforded by hybrid vigor in rice.

The endoglucanase Ⅱ of Trichoderma reesei is considered the most effective enzyme for biofinishing cotton fabrics and biostoning denim garments. However, the commercially available preparation of endoglucanase Ⅱ is usually mixed with other cellulase components, especially endoglucanase I, resulting in hydrolysis and weight loss of garments during biofinishing and biostoning. We thus isolated the endoglucanase Ⅱ gene from T. reesei to express this in Pichia pastoris, under the control of a methanol-inducible AOX1 promoter, to avoid the presence of other cellulase components. A highly expressible Mut+ transformant was selected and its expression in BMMH medium was found most suitable for the production of large amounts of the recombinant protein. Recombinant endoglucanase Ⅱ was purified to electrophoretic homogeneity, and functionally characterized by activity staining. The specific activity of recombinant endoglucanase Ⅱ was found to be 220.57 EU/mg of protein. Purified recombinant endoglucanase Ⅱ was estimated to have a molecular mass of 52.8 kDa. The increase in molecular mass was likely due to hyperglycosylation. Hyperglycosylation of recombinant endoglucanase Ⅱ secreted by P. pastoris did not change the temperature or pH optima as compared to the native protein, but did result in increased thermostability. Kinetic analysis showed that recombinant endoglucanase was most active against amorphous cellulose, such as carboxymethyl cellulose, for which it also had a high affinity.

The rice Ubiquitin1 (Ubi1) promoter was tested to evaluate its capacity to express the heterologous gusA gene encoding β-glucuronidase in transgenic rice tissue relative to the commonly used Ubi1 corn promoter and the rice gibberellic acid insensitive (GAI) gene promoter element. Experimental results showed increased expression of gusA gene in rice tissue when driven by the native Ubi1 promoter when compared to the use of corn Ubi1 promoter. Results further indicated that the cis-regulatory elements present in the native promoter element might have been responsible for high expression. However, the gusA gene expression level when driven by the rice GAI promoter was notably lower than both Ubi1 promoters. The present study, thus, for the first time helped to demonstrate that the native Ubi1 promoter is a promising genetic element in transgenic approaches for constitutive expression of any gene in rice tissue.

Generation of insect-resistant, transgenic crop plants by expression of the insecticidal crystal protein (ICP) gene of Bacillus thuringiensis (Bt) is a standard crop improvement approach. In such cases, adequate expression of the most appropriate ICP against the target insect pest of the crop species is desirable. It is also considered advantageous to generate Bt-transgenics with multiple toxin systems to control rapid development of pest resistance to the ICP. Larvae of yellow stem borer (YSB), Scirpophaga incertulas, a major lepidopteran insect pest of rice, cause massive losses of rice yield. Studies on insect feeding and on the binding properties of ICP to brush border membrane receptors in the midgut of YSB larvae revealed that cryIAb and cryIAc are two individually suitable candidate genes for developing YSB-resistant rice. Programs were undertaken to develop Bt-transgenic rice with these ICP genes independently in a single cultivar. A cryIAc gene was reconstructed and placed under control of the maize ubiquitin 1 promoter, along with the first intron of the maize ubiquitin 1 gene, and the nos terminator. The gene construct was delivered to embryogenic calli of IR64, an elite indica rice cultivar, using the particle bombardment method. Six highly expressive independent transgenic ICP lines were identified. Molecular analyses and insect-feeding assays of two such lines revealed that the transferred synthetic cryIAc gene was expressed stably in the T2generation of these lines and that the transgenic rice plants were highly toxic to YSB larvae and lessened the damage caused by their feeding.

Main conclusion The present study documented the predominant role of WRKY transcription factor in controlling genes of different pathways related to fibre formation in jute and could be a candidate gene for the improvement of jute fiber. Understanding of molecular mechanism associated with bast fibre development is of immense significance to achieve desired improvement in jute (Corchorus sp.). Therefore, suppression subtractive hybridization was successfully applied to identify genes involved in fibre developmental process in jute. The subtracted library of normal Corchorus capsularis as tester with respect to its fibre-deficient mutant as driver resulted in 2,685 expressed sequence tags which were assumed to represent the differentially expressed genes between two genotypes. The identified expressed sequence tags were assembled and clustered into 225 contigs and 231 singletons. Among these 456 unigenes, 377 were classified into 15 different functional categories while others were of unknown functional category. Reverse Northern analysis of the unigenes showed distinct variation in hybridization intensity of 11 transcripts between two genotypes tested. The findings were also documented by Northern and real-time PCR analysis. Varied expression level of these transcripts suggested their crucial involvement in fibre development in this species. Among these transcripts, WRKY transcription factor was documented to be a most important transcript which was in agreement with its known role in other plant species in possible regulation related to cell wall biosynthesis, expansion and lignification. This report constitutes first systematic analysis of genes involved in fibre development process in jute. It may be suggested that the information generated in this study would be useful for genetic improvement of fibre traits in this plant species.

An efficient protocol for shoot regeneration was developed for sesame ( Sesamum indicum L.) internodes using the transverse thin cell layer (tTCL) culture method. The frequency of shoot regeneration and the number of adventitious buds produced from regenerated shoots depend significantly on explant age, thickness of the tTCL sections, and the phytohormones supplemented to the culture medium. A combination of 6-benzyladenine (2.0 mg l −1 ) and α-naphthaleneacetic acid (0.5 mg l −1 ) was found to be the best phytohormone combination for shoot bud induction, with the maximum number of shoots obtained when the tTCL sections were 0.5–1.0 mm thick and derived from 4- to 6-week-old seedlings of sesame. Well-developed shoots were rooted on MS medium without phytohormones, and 80% of the regenerated plantlets were successfully established in soil.

Main conclusion The present study documented the predominant role of WRKY transcription factor in controlling genes of different pathways related to fibre formation in jute and could be a candidate gene for the improvement of jute fiber. Understanding of molecular mechanism associated with bast fibre development is of immense significance to achieve desired improvement in jute ( Corchorus sp.). Therefore, suppression subtractive hybridization was successfully applied to identify genes involved in fibre developmental process in jute. The subtracted library of normal Corchorus capsularis as tester with respect to its fibre-deficient mutant as driver resulted in 2,685 expressed sequence tags which were assumed to represent the differentially expressed genes between two genotypes. The identified expressed sequence tags were assembled and clustered into 225 contigs and 231 singletons. Among these 456 unigenes, 377 were classified into 15 different functional categories while others were of unknown functional category. Reverse Northern analysis of the unigenes showed distinct variation in hybridization intensity of 11 transcripts between two genotypes tested. The findings were also documented by Northern and real-time PCR analysis. Varied expression level of these transcripts suggested their crucial involvement in fibre development in this species. Among these transcripts, WRKY transcription factor was documented to be a most important transcript which was in agreement with its known role in other plant species in possible regulation related to cell wall biosynthesis, expansion and lignification. This report constitutes first systematic analysis of genes involved in fibre development process in jute. It may be suggested that the information generated in this study would be useful for genetic improvement of fibre traits in this plant species.

Establishment of a compatible and suitable regeneration and gene transformation protocol is of immense significance to achieve the desired improvement in pigeonpea through biotechnological approach. However, no reproducible protocol for large-scale generation of independent primary transformants plant lines through organogenesis virtually exists. Therefore, an attempt was initiated to develop a simple and straightforward protocol for easy generation of transformed lines through the facilitation of the production of multiple shooting and their lateral branching followed by selection of generated axillary buds. The protocol involves infection of longitudinally dissected cotyledonary node and shoot-tips of germinated seedling with Agrobacterium harboring genetic cassette having both the bialaphos-resistant and green fluorescent protein ( gfp ) genes as a plant selection marker and reporter gene respectively. The infected explants were allowed to grow in medium with 0.25 mg/L BAP and 0.1 mg/L gibberellic acid (GA 3 ) for development of multiple shoots followed by micropropagation of nodal segments and selection under 4 mg/L bialaphos. The selected plants were found to contain integrated genes and carried through successive generations as revealed by dot blot, Southern hybridization, and expression of the reporter gene. In spite of having the scope of the generation of siblings, the resulted transformation frequency was found to be 6.66% irrespective of genotypes used in the present study as revealed by dot blot analysis. Overall, the protocol ensured generation of pigeonpea transgenic plants with considerable easy within a short time irrespective of genotypes/cultivars of the crop.

Jute ( Corchorus sp.) is a commercially important bast fibre-producing crop. Long-awaited industrial demand for improvement of quality of fibre in this crop species is restricted by the lack of knowledge about genetic basis of economically important characters and their assemblage. Therefore, the present study was aimed at to compare the assemblage of single copy expressed sequence tag loci on nucleolar organizing chromosome of C. capsularis and C. olitorius at the molecular level. The expressed sequence tag loci associated with chromosome 5 of C. olitorius were also found to be linked to same chromosome of C. capsularis . However, arrangements of the loci were found to be altered within as well as between the arms of chromosome 5 of Corchorus sp. The present study, for the first time, documented the occurrence of structural alteration at the molecular level between two jute species at least with respect to the chromosome 5. Such cryptic structural alterations resulted from chromosomal rearrangements (intra-chromosomal shift) in either of the species.