Increase in atmospheric CO2 ([CO2]) improves the efficiency of the unsaturated photosynthetic sys... more Increase in atmospheric CO2 ([CO2]) improves the efficiency of the unsaturated photosynthetic system of C3 plants, like rice, which leads to high crop productivity and increased biomass production. Planting geometry using lower than standard planting density has been shown to be an alternative pre-screening technique for phenotypic plasticity as a proxy of [CO2] responsiveness. More than 200 indigenous rice genotypes were tested for several traits, such as plant height, tiller number, panicle number, panicle dry weight, straw dry weight, total dry weight and harvest index. Their relative response for these phenotypic traits at low density planting versus normal density was determined to assess the plasticity of the genotypes. Ten genotypes were identified as significantly [CO2] responsive based on their higher panicle dry weight and panicle number. Even though it was observed that genotypes with higher days to maturity (DM) were more plastic, rice genotypes with low DM had a signifi...
The rice landrace Horkuch, endemic to the southern saline coast of Bangladesh, is known to have s... more The rice landrace Horkuch, endemic to the southern saline coast of Bangladesh, is known to have salt tolerance traits and can therefore contribute to a high yielding recipient for breeding purposes. In this study, we reciprocally crossed Horkuch with high yielding but salt sensitive IR29 to detect the complement of genes that were responsible for conferring salt tolerance versus sensitivity at the seedling developmental stage. We looked at tolerant and sensitive F 3 families from individual F 2 segregating plants and analyzed them for differential gene expressions using RNAseq. In general, we observed higher numbers of genes differentially expressed in leaves compared to root tissues. This included both upregulation and downregulation of gene expression across our experimental factors. Gene expression decreased in sensitive leaf after stress exposure where tolerant plants showed the opposite trend. In root, tolerant plants expression decreased at higher time points of stress exposure. We also observed a strong maternal cytoplasmic effect on gene expression and this was most evident in roots where there was upregulation in functional enrichments related to phosphorylation, electron carriers, transporter and cation transmembrane activities. Stress groups (tolerant and sensitive) response in F 3 families were distinctive in both cytoplasmic backgrounds and involved uniquely upregulated genes in tolerant progenies including membrane sensor proteins, enzymes involved with signaling pathways, such as those producing trehalose and G-protein coupled receptor proteins, photosynthesis-related enzymes and golgi body recycling as well as prolamin precursor proteins involved in refolding of proteins. On the other hand, sensitivity was found to be associated with differential upregulation of only a few redox proteins and higher number of apoptosis related genes compared to the tolerant response. Overall, our highly replicated experimental design was powerful and allowed the detection of relatively subtle differential expression. Our future goal is to correlate these expression differences with QTLs in this population, which would help identify the relative importance of specific genetic loci and provide a direct avenue for combining higher levels of salt tolerance with better agronomic traits in rice. Nearly 1 million ha of coastal soil in Bangladesh or nearly a ninth of the total cultivable area is affected by soil salinity, mostly due to seawater intrusion 1. The inhibitory levels of salinity for rice cultivation (>4 dS/m) are seasonal, except in certain pockets of the South West 2. During the dry season water levels are usually low due to upper riparian withdrawal as well as its overexploitation, causing a build-up of salts. However, these coastal areas are home to many salt tolerant rice landraces, including both indica and aromatic subgroups, which have allelic diversity at several genetic loci associated with tolerance from donor landraces like Pokkali and Nona Bokra 3,4. One of these rice landraces, Horkuch, was previously characterized as salt tolerant at the seedling stage 3,5 and at the reproductive stage 6. These rice landraces from coastal Bangladesh are likely to harbor novel sources of salt tolerance, due to their allelic diversity, that can complement known salt tolerance genes. The introduction of
DNA helicase (PDH45) from the pea plant (Pisum sativum) is a member of the DEAD box protein famil... more DNA helicase (PDH45) from the pea plant (Pisum sativum) is a member of the DEAD box protein family and plays a vital regulatory role in saline stress tolerance in plants. We previously reported that over-expression of PDH45 gene confers both seedling and reproductive stage salinity tolerance to a Bangladeshi rice landrace, Binnatoa (BA). In this study, transgenic BA-containing PDH45 (♂) was crossed with two different farmer-popular BRRI rice varieties (♀), BR28 and BR47, in a contained net house. F1 plants positive for the transgene and having recipient phenotype were advanced from F1 to F5. Expression of the PDH45 gene was detected in all generations. The expression level of PDH45 was 200-fold higher in the donor compared to the two recipient genotypes but without any effect on their salt stress tolerance ability in various assays. Under 120 mM NaCl stress at seedling stage, all rice genotypes showed vigorous growth, higher chlorophyll content, lower electrolyte leakage and lower L...
Genetic variation of Oryza sativa L. landraces (LRs) collected from the saline coastal belt of Ba... more Genetic variation of Oryza sativa L. landraces (LRs) collected from the saline coastal belt of Bangladesh, modern varieties (MVs), as well as Pokkali, Nona Bokra and salt tolerant modern varieties (SMVs) derived from the last two were analyzed with 60 evenly distributed rice microsatellite DNA markers. A total of 196 reproducible polymorphic alleles were identified from the band loci. Heterozygosity among the 31 LRs was found to be 0.57, 0.46 in the 5 MVs and 0.40 in the 8 SMVs. Computation of genetic similarity with this data, using Jaccard's coefficient followed by UPGMA clustering, divided the landraces into 6 distinct groups. Three groups were composed of LRs only from the highly saline southwest. Two groups consisted of LRs from the mild to moderately saline mid-east and northeast coasts. The sixth group was heterogeneous, with LRs from the northeast, LRs from the southwest and Nona Bokra. Pokkali and Gunshi, a LR of the southwest, branched out individually. When all the 46 O. sativa L. cultivars were clustered together, most of the MVs and SMVs were found to be linked within the heterogeneous group. The measure of seedling Na and K concentration, Na/K ratios, affected leaf area as well as survival under salinity stress in hydroponics identified 6 LRs from the highly saline southwest as the most tolerant. These group with Pokkali when UPGMA clustering using the Pearson product-moment correlation coefficient suitable for the quantitative physiological data on seedling saline stress was computed. Morphological observations of plant type and height, days to maturity and yield components in non-saline soil indicated low variability among the different LRs. When yield performance as well as tolerance scores were considered, 7 LRs from the southwest and 1 LR from the mid-northeast show potential as donors for breeding salt tolerant rice. The microsatellite fingerprinting analysis thus revealed that some of the salt tolerant landraces of the coastal region have unique polymorphic loci, quite distinct from the popular salt tolerance donor Pokkali. The similarity matrices between the O. sativa L. cultivars chosen for the study can be used as a valuable tool for the proper choice of parents for mapping or breeding purposes.
S u m m a r y. The addition of 3 mg/1 of nordihydroguaiaretic acid (NDGA) to BAP and tyrosine for... more S u m m a r y. The addition of 3 mg/1 of nordihydroguaiaretic acid (NDGA) to BAP and tyrosine fortified MS medium was essential to obtain organogeni_c callus from the hypocotyl segments of two varieties (D-154 and CVL-1) of Corchorus capsularis-one of the two jute species. When the organogenic callus, which is rich in large starch granules, was transferred to MS basal medium, it differentiated into single or multiple sboots usually in the first subculture and sometimes in the' second. The activity of glyoxalase-I of the organogenic callus was found to be significantly lower than that observed in the nonorganogenic callus initiated on MS medium supplemented with 2,4-D, tyrosine, BAP or just 'BAP and tyrosine. This suggests an inverse relationship between differentiation and the level of glyoxalase-I activity in the two varieties of C. capsularis jute.
Nitric oxide (NO) and glutathione (GSH) regulate a variety of physiological processes and stress ... more Nitric oxide (NO) and glutathione (GSH) regulate a variety of physiological processes and stress responses; however, their involvement in mitigating Cu toxicity in plants has not been extensively studied. This study investigated the interactive effect of exogenous sodium nitroprusside (SNP) and GSH on Cu homeostasis and Cu-induced oxidative damage in rice seedlings. Hydroponically grown 12-day-old seedlings were subjected to 100 μM CuSO4 alone and in combination with 200 μM SNP (an NO donor) and 200 μM GSH. Cu exposure for 48 h resulted in toxicity symptoms such as stunted growth, chlorosis, and rolling in leaves. Cu toxicity was also manifested by a sharp increase in lipoxygenase (LOX) activity, lipid peroxidation (MDA), hydrogen peroxide (H2O2), proline (Pro) content, and rapid reductions in biomass, chlorophyll (Chl), and relative water content (RWC). Cu-caused oxidative stress was evident by overaccumulation of reactive oxygen species (ROS; superoxide (O2 (•-)) and H2O2). Ascorbate (AsA) content decreased while GSH and phytochelatin (PC) content increased significantly in Cu-stressed seedlings. Exogenous SNP, GSH, or SNP + GSH decreased toxicity symptoms and diminished a Cu-induced increase in LOX activity, O2 (•-), H2O2, MDA, and Pro content. They also counteracted a Cu-induced increase in superoxide dismutase (SOD), ascorbate peroxidase (APX), glutathione reductase (GR), monodehydroascorbate reductase (MDHAR), and glyoxalase I and glyoxalase II activities, which paralleled changes in ROS and MDA levels. These seedlings also showed a significant increase in catalase (CAT), glutathione peroxidase (GPX), dehydroascorbate reductase (DHAR), glutathione S-transferase (GST) activities, and AsA and PC content compared with the seedlings stressed with Cu alone. Cu analysis revealed that SNP and GSH restricted the accumulation of Cu in the roots and leaves of Cu-stressed seedlings. Our results suggest that Cu exposure provoked an oxidative burden while reduced Cu uptake and modulating the antioxidant defense and glyoxalase systems by adding SNP and GSH play an important role in alleviating Cu toxicity. Furthermore, the protective action of GSH and SNP + GSH was more efficient than SNP alone.
At the cellular level, the Salt Overly Sensitive (SOS) signaling pathway comprising SOS3, SOS2, a... more At the cellular level, the Salt Overly Sensitive (SOS) signaling pathway comprising SOS3, SOS2, and SOS1 has been proposed to mediate cellular signaling under salt stress to maintain ion (Na +) homeostasis. In this regulatory pathway, both OsSOS1 encoding plasma membrane and OsNHX1 encoding vacuolar Na + /H + antiporters are regulated by SOS3-SOS2 protein kinase complex. In the present study, the rice variety BRRI dhan28-which is popular with farmers and high yielding, but salt sensitive, was transformed with the OsSOS1 gene isolated from salt tolerant Pokkali rice and driven by the constitutive promoter, CaMV35S. The construct was transformed through a tissue culture-independent Agrobacteriummediated in planta transformation method that circumvents the problems associated with tissue culture-based indica rice transformation methods. Integration of the foreign genes (OsSOS1) into the genome of transgenic plants was confirmed by gene-specific PCR and Southern blot analysis. The level of transgene expression (SOS1) was also quantified by semi-quantitative RT PCR and real time PCR. Genetic segregation ratio for T1 progenies was calculated and found to follow the Mendelian law of inheritance in case of positive transformants. The transformants were shown to be salt tolerant compared to wild type in molecular analysis as well as physiological screening. Future work will involve transformation of both the OsSOS1 and OsNHX1 genes together; with the expectation for enhancing the tolerance level compared to currently available transgenic rice.
Soil salinity is one of the most challenging problems that restricts the normal growth and produc... more Soil salinity is one of the most challenging problems that restricts the normal growth and production of rice worldwide. It has therefore become very important to produce more saline tolerant rice varieties. This study shows constitutive over-expression of the vacuolar Na + /H + antiporter gene (OsNHX1) from the rice landrace (Pokkali) and attainment of enhanced level of salinity tolerance in transgenic rice plants. It also shows that inclusion of the complete un-translated regions (UTRs) of the alternatively spliced OsNHX1 gene provides a higher level of tolerance to the transgenic rice. Two separate transformation events of the OsNHX1 gene, one with 1.9 kb region containing the 5 UTR with CDS and the other of 2.3 kb, including 5 UTR, CDS, and the 3 UTR regions were performed. The transgenic plants with these two different constructs were advanced to the T 3 generation and physiological and molecular screening of homozygous plants was conducted at seedling and reproductive stages under salinity (NaCl) stress. Both transgenic lines were observed to be tolerant compared to WT plants at both physiological stages. However, the transgenic lines containing the CDS with both the 5 and 3 UTR were significantly more tolerant compared to the transgenic lines containing OsNHX1 gene without the 3 UTR. At the seedling stage at 12 dS/m stress, the chlorophyll content was significantly higher (P < 0.05) and the electrolyte leakage significantly lower (P < 0.05) in the order 2.3 kb > 1.9 kb > and WT lines. Yield in g/plant in the best line from the 2.3 kb plants was significantly more (P < 0.01) compared, respectively, to the best 1.9 kb line and WT plants at stress of 6 dS/m. Transformation with the complete transcripts rather than the CDS may therefore provide more durable level of tolerance.
The two major environmental factors that currently reduce plant productivity are drought and sali... more The two major environmental factors that currently reduce plant productivity are drought and salinity. It has been reported that the plant‐specific NAC (NAM, ATAF1/2, CUC2) transcription factors play diverse roles in plant development and stress responses. Stress‐specific NAC1 or SNAC1‐overexpressing transgenic rice plants showed significantly improved drought resistance under field conditions and strong tolerance to salt stress. So the purpose of this study was to clone the SNAC1 gene from the salt tolerant rice landrace Pokkali for transformation into a salt sensitive rice variety. SNAC1 cDNA was isolated and successfully cloned in the pENTR vector. Cloning was confirmed by PCR, restriction digestion and sequencing analysis and subsequently mobilized into the Agrobacterium compatible destination vector pH7WG2 for rice transformation. Transgenic status of T0 plants of SNAC1 was confirmed by PCR analysis. T0 plants were advanced to T1 transgenic lines and further confirmed by PCR an...
Tomato (Solanum lycopersicum) is a very popular nutritious vegetable of Bangladesh. But its produ... more Tomato (Solanum lycopersicum) is a very popular nutritious vegetable of Bangladesh. But its production is much lower than demand. To enhance its production, varieties resistant to various environmental stresses and pathogens need to be developed. In the present study, to establish a transformation protocol cotyledonary leaves of 7-10 days old tomato plants were transformed with Agrobacterium tumefaciens strain LBA4404 harboring binary vector pBI121containing nptII gene and uidA gene as selection markers. Integration of the transgene in the explants was confirmed by GUS histochemical assay under kanamycin selection pressure. Effect of transient GUS expression showed that transformation efficiency was modulated by several factors, such as, optical density, infection duration and co-cultivation duration. Transformation with a bacterial solution (OD600 approx. 0.6) followed by 48 hrs of co-culture resulted the highest transient GUS expression tomato varieties collected from both BARI an...
Increase in atmospheric CO2 ([CO2]) improves the efficiency of the unsaturated photosynthetic sys... more Increase in atmospheric CO2 ([CO2]) improves the efficiency of the unsaturated photosynthetic system of C3 plants, like rice, which leads to high crop productivity and increased biomass production. Planting geometry using lower than standard planting density has been shown to be an alternative pre-screening technique for phenotypic plasticity as a proxy of [CO2] responsiveness. More than 200 indigenous rice genotypes were tested for several traits, such as plant height, tiller number, panicle number, panicle dry weight, straw dry weight, total dry weight and harvest index. Their relative response for these phenotypic traits at low density planting versus normal density was determined to assess the plasticity of the genotypes. Ten genotypes were identified as significantly [CO2] responsive based on their higher panicle dry weight and panicle number. Even though it was observed that genotypes with higher days to maturity (DM) were more plastic, rice genotypes with low DM had a signifi...
The rice landrace Horkuch, endemic to the southern saline coast of Bangladesh, is known to have s... more The rice landrace Horkuch, endemic to the southern saline coast of Bangladesh, is known to have salt tolerance traits and can therefore contribute to a high yielding recipient for breeding purposes. In this study, we reciprocally crossed Horkuch with high yielding but salt sensitive IR29 to detect the complement of genes that were responsible for conferring salt tolerance versus sensitivity at the seedling developmental stage. We looked at tolerant and sensitive F 3 families from individual F 2 segregating plants and analyzed them for differential gene expressions using RNAseq. In general, we observed higher numbers of genes differentially expressed in leaves compared to root tissues. This included both upregulation and downregulation of gene expression across our experimental factors. Gene expression decreased in sensitive leaf after stress exposure where tolerant plants showed the opposite trend. In root, tolerant plants expression decreased at higher time points of stress exposure. We also observed a strong maternal cytoplasmic effect on gene expression and this was most evident in roots where there was upregulation in functional enrichments related to phosphorylation, electron carriers, transporter and cation transmembrane activities. Stress groups (tolerant and sensitive) response in F 3 families were distinctive in both cytoplasmic backgrounds and involved uniquely upregulated genes in tolerant progenies including membrane sensor proteins, enzymes involved with signaling pathways, such as those producing trehalose and G-protein coupled receptor proteins, photosynthesis-related enzymes and golgi body recycling as well as prolamin precursor proteins involved in refolding of proteins. On the other hand, sensitivity was found to be associated with differential upregulation of only a few redox proteins and higher number of apoptosis related genes compared to the tolerant response. Overall, our highly replicated experimental design was powerful and allowed the detection of relatively subtle differential expression. Our future goal is to correlate these expression differences with QTLs in this population, which would help identify the relative importance of specific genetic loci and provide a direct avenue for combining higher levels of salt tolerance with better agronomic traits in rice. Nearly 1 million ha of coastal soil in Bangladesh or nearly a ninth of the total cultivable area is affected by soil salinity, mostly due to seawater intrusion 1. The inhibitory levels of salinity for rice cultivation (>4 dS/m) are seasonal, except in certain pockets of the South West 2. During the dry season water levels are usually low due to upper riparian withdrawal as well as its overexploitation, causing a build-up of salts. However, these coastal areas are home to many salt tolerant rice landraces, including both indica and aromatic subgroups, which have allelic diversity at several genetic loci associated with tolerance from donor landraces like Pokkali and Nona Bokra 3,4. One of these rice landraces, Horkuch, was previously characterized as salt tolerant at the seedling stage 3,5 and at the reproductive stage 6. These rice landraces from coastal Bangladesh are likely to harbor novel sources of salt tolerance, due to their allelic diversity, that can complement known salt tolerance genes. The introduction of
DNA helicase (PDH45) from the pea plant (Pisum sativum) is a member of the DEAD box protein famil... more DNA helicase (PDH45) from the pea plant (Pisum sativum) is a member of the DEAD box protein family and plays a vital regulatory role in saline stress tolerance in plants. We previously reported that over-expression of PDH45 gene confers both seedling and reproductive stage salinity tolerance to a Bangladeshi rice landrace, Binnatoa (BA). In this study, transgenic BA-containing PDH45 (♂) was crossed with two different farmer-popular BRRI rice varieties (♀), BR28 and BR47, in a contained net house. F1 plants positive for the transgene and having recipient phenotype were advanced from F1 to F5. Expression of the PDH45 gene was detected in all generations. The expression level of PDH45 was 200-fold higher in the donor compared to the two recipient genotypes but without any effect on their salt stress tolerance ability in various assays. Under 120 mM NaCl stress at seedling stage, all rice genotypes showed vigorous growth, higher chlorophyll content, lower electrolyte leakage and lower L...
Genetic variation of Oryza sativa L. landraces (LRs) collected from the saline coastal belt of Ba... more Genetic variation of Oryza sativa L. landraces (LRs) collected from the saline coastal belt of Bangladesh, modern varieties (MVs), as well as Pokkali, Nona Bokra and salt tolerant modern varieties (SMVs) derived from the last two were analyzed with 60 evenly distributed rice microsatellite DNA markers. A total of 196 reproducible polymorphic alleles were identified from the band loci. Heterozygosity among the 31 LRs was found to be 0.57, 0.46 in the 5 MVs and 0.40 in the 8 SMVs. Computation of genetic similarity with this data, using Jaccard's coefficient followed by UPGMA clustering, divided the landraces into 6 distinct groups. Three groups were composed of LRs only from the highly saline southwest. Two groups consisted of LRs from the mild to moderately saline mid-east and northeast coasts. The sixth group was heterogeneous, with LRs from the northeast, LRs from the southwest and Nona Bokra. Pokkali and Gunshi, a LR of the southwest, branched out individually. When all the 46 O. sativa L. cultivars were clustered together, most of the MVs and SMVs were found to be linked within the heterogeneous group. The measure of seedling Na and K concentration, Na/K ratios, affected leaf area as well as survival under salinity stress in hydroponics identified 6 LRs from the highly saline southwest as the most tolerant. These group with Pokkali when UPGMA clustering using the Pearson product-moment correlation coefficient suitable for the quantitative physiological data on seedling saline stress was computed. Morphological observations of plant type and height, days to maturity and yield components in non-saline soil indicated low variability among the different LRs. When yield performance as well as tolerance scores were considered, 7 LRs from the southwest and 1 LR from the mid-northeast show potential as donors for breeding salt tolerant rice. The microsatellite fingerprinting analysis thus revealed that some of the salt tolerant landraces of the coastal region have unique polymorphic loci, quite distinct from the popular salt tolerance donor Pokkali. The similarity matrices between the O. sativa L. cultivars chosen for the study can be used as a valuable tool for the proper choice of parents for mapping or breeding purposes.
S u m m a r y. The addition of 3 mg/1 of nordihydroguaiaretic acid (NDGA) to BAP and tyrosine for... more S u m m a r y. The addition of 3 mg/1 of nordihydroguaiaretic acid (NDGA) to BAP and tyrosine fortified MS medium was essential to obtain organogeni_c callus from the hypocotyl segments of two varieties (D-154 and CVL-1) of Corchorus capsularis-one of the two jute species. When the organogenic callus, which is rich in large starch granules, was transferred to MS basal medium, it differentiated into single or multiple sboots usually in the first subculture and sometimes in the' second. The activity of glyoxalase-I of the organogenic callus was found to be significantly lower than that observed in the nonorganogenic callus initiated on MS medium supplemented with 2,4-D, tyrosine, BAP or just 'BAP and tyrosine. This suggests an inverse relationship between differentiation and the level of glyoxalase-I activity in the two varieties of C. capsularis jute.
Nitric oxide (NO) and glutathione (GSH) regulate a variety of physiological processes and stress ... more Nitric oxide (NO) and glutathione (GSH) regulate a variety of physiological processes and stress responses; however, their involvement in mitigating Cu toxicity in plants has not been extensively studied. This study investigated the interactive effect of exogenous sodium nitroprusside (SNP) and GSH on Cu homeostasis and Cu-induced oxidative damage in rice seedlings. Hydroponically grown 12-day-old seedlings were subjected to 100 μM CuSO4 alone and in combination with 200 μM SNP (an NO donor) and 200 μM GSH. Cu exposure for 48 h resulted in toxicity symptoms such as stunted growth, chlorosis, and rolling in leaves. Cu toxicity was also manifested by a sharp increase in lipoxygenase (LOX) activity, lipid peroxidation (MDA), hydrogen peroxide (H2O2), proline (Pro) content, and rapid reductions in biomass, chlorophyll (Chl), and relative water content (RWC). Cu-caused oxidative stress was evident by overaccumulation of reactive oxygen species (ROS; superoxide (O2 (•-)) and H2O2). Ascorbate (AsA) content decreased while GSH and phytochelatin (PC) content increased significantly in Cu-stressed seedlings. Exogenous SNP, GSH, or SNP + GSH decreased toxicity symptoms and diminished a Cu-induced increase in LOX activity, O2 (•-), H2O2, MDA, and Pro content. They also counteracted a Cu-induced increase in superoxide dismutase (SOD), ascorbate peroxidase (APX), glutathione reductase (GR), monodehydroascorbate reductase (MDHAR), and glyoxalase I and glyoxalase II activities, which paralleled changes in ROS and MDA levels. These seedlings also showed a significant increase in catalase (CAT), glutathione peroxidase (GPX), dehydroascorbate reductase (DHAR), glutathione S-transferase (GST) activities, and AsA and PC content compared with the seedlings stressed with Cu alone. Cu analysis revealed that SNP and GSH restricted the accumulation of Cu in the roots and leaves of Cu-stressed seedlings. Our results suggest that Cu exposure provoked an oxidative burden while reduced Cu uptake and modulating the antioxidant defense and glyoxalase systems by adding SNP and GSH play an important role in alleviating Cu toxicity. Furthermore, the protective action of GSH and SNP + GSH was more efficient than SNP alone.
At the cellular level, the Salt Overly Sensitive (SOS) signaling pathway comprising SOS3, SOS2, a... more At the cellular level, the Salt Overly Sensitive (SOS) signaling pathway comprising SOS3, SOS2, and SOS1 has been proposed to mediate cellular signaling under salt stress to maintain ion (Na +) homeostasis. In this regulatory pathway, both OsSOS1 encoding plasma membrane and OsNHX1 encoding vacuolar Na + /H + antiporters are regulated by SOS3-SOS2 protein kinase complex. In the present study, the rice variety BRRI dhan28-which is popular with farmers and high yielding, but salt sensitive, was transformed with the OsSOS1 gene isolated from salt tolerant Pokkali rice and driven by the constitutive promoter, CaMV35S. The construct was transformed through a tissue culture-independent Agrobacteriummediated in planta transformation method that circumvents the problems associated with tissue culture-based indica rice transformation methods. Integration of the foreign genes (OsSOS1) into the genome of transgenic plants was confirmed by gene-specific PCR and Southern blot analysis. The level of transgene expression (SOS1) was also quantified by semi-quantitative RT PCR and real time PCR. Genetic segregation ratio for T1 progenies was calculated and found to follow the Mendelian law of inheritance in case of positive transformants. The transformants were shown to be salt tolerant compared to wild type in molecular analysis as well as physiological screening. Future work will involve transformation of both the OsSOS1 and OsNHX1 genes together; with the expectation for enhancing the tolerance level compared to currently available transgenic rice.
Soil salinity is one of the most challenging problems that restricts the normal growth and produc... more Soil salinity is one of the most challenging problems that restricts the normal growth and production of rice worldwide. It has therefore become very important to produce more saline tolerant rice varieties. This study shows constitutive over-expression of the vacuolar Na + /H + antiporter gene (OsNHX1) from the rice landrace (Pokkali) and attainment of enhanced level of salinity tolerance in transgenic rice plants. It also shows that inclusion of the complete un-translated regions (UTRs) of the alternatively spliced OsNHX1 gene provides a higher level of tolerance to the transgenic rice. Two separate transformation events of the OsNHX1 gene, one with 1.9 kb region containing the 5 UTR with CDS and the other of 2.3 kb, including 5 UTR, CDS, and the 3 UTR regions were performed. The transgenic plants with these two different constructs were advanced to the T 3 generation and physiological and molecular screening of homozygous plants was conducted at seedling and reproductive stages under salinity (NaCl) stress. Both transgenic lines were observed to be tolerant compared to WT plants at both physiological stages. However, the transgenic lines containing the CDS with both the 5 and 3 UTR were significantly more tolerant compared to the transgenic lines containing OsNHX1 gene without the 3 UTR. At the seedling stage at 12 dS/m stress, the chlorophyll content was significantly higher (P < 0.05) and the electrolyte leakage significantly lower (P < 0.05) in the order 2.3 kb > 1.9 kb > and WT lines. Yield in g/plant in the best line from the 2.3 kb plants was significantly more (P < 0.01) compared, respectively, to the best 1.9 kb line and WT plants at stress of 6 dS/m. Transformation with the complete transcripts rather than the CDS may therefore provide more durable level of tolerance.
The two major environmental factors that currently reduce plant productivity are drought and sali... more The two major environmental factors that currently reduce plant productivity are drought and salinity. It has been reported that the plant‐specific NAC (NAM, ATAF1/2, CUC2) transcription factors play diverse roles in plant development and stress responses. Stress‐specific NAC1 or SNAC1‐overexpressing transgenic rice plants showed significantly improved drought resistance under field conditions and strong tolerance to salt stress. So the purpose of this study was to clone the SNAC1 gene from the salt tolerant rice landrace Pokkali for transformation into a salt sensitive rice variety. SNAC1 cDNA was isolated and successfully cloned in the pENTR vector. Cloning was confirmed by PCR, restriction digestion and sequencing analysis and subsequently mobilized into the Agrobacterium compatible destination vector pH7WG2 for rice transformation. Transgenic status of T0 plants of SNAC1 was confirmed by PCR analysis. T0 plants were advanced to T1 transgenic lines and further confirmed by PCR an...
Tomato (Solanum lycopersicum) is a very popular nutritious vegetable of Bangladesh. But its produ... more Tomato (Solanum lycopersicum) is a very popular nutritious vegetable of Bangladesh. But its production is much lower than demand. To enhance its production, varieties resistant to various environmental stresses and pathogens need to be developed. In the present study, to establish a transformation protocol cotyledonary leaves of 7-10 days old tomato plants were transformed with Agrobacterium tumefaciens strain LBA4404 harboring binary vector pBI121containing nptII gene and uidA gene as selection markers. Integration of the transgene in the explants was confirmed by GUS histochemical assay under kanamycin selection pressure. Effect of transient GUS expression showed that transformation efficiency was modulated by several factors, such as, optical density, infection duration and co-cultivation duration. Transformation with a bacterial solution (OD600 approx. 0.6) followed by 48 hrs of co-culture resulted the highest transient GUS expression tomato varieties collected from both BARI an...
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Papers by Zeba Seraj