A rapid and efficient Agrobacterium -mediated transformation system in sorghum has been developed... more A rapid and efficient Agrobacterium -mediated transformation system in sorghum has been developed employing standard binary vectors and bar gene as a selectable marker. Sorghum (Sorghum bicolor) is an important food and biofuel crop worldwide, for which improvements in genetic transformation are needed to study its biology and facilitate agronomic and commercial improvement. Here, we report optimization of regeneration and transformation of public sorghum genotype P898012 using standard binary vectors and bar gene as a selectable marker. The tissue culture regeneration time frame has been reduced to 7-12 weeks with a yield of over 18 plants per callus, and the optimized transformation system employing Agrobacterium tumefaciens strain AGL1 and the bar with a MAS promoter achieved an average frequency over 14 %. Of randomly analyzed independent transgenic events, 40-50 % carry single copy of integrated T-DNA. Some independent transgenic events were derived from the same embryogenic callus lines, but a 3:1 Mendelian segregation ratio was found in all transgenic events with single copy as estimated by Southern blots. The system described here should facilitate studies of sorghum biology and agronomic improvement.
In rice (Oryza sativa L.), the root system plays different essential roles, from water and nutrie... more In rice (Oryza sativa L.), the root system plays different essential roles, from water and nutrient uptake to responding to environmental signals. The mechanisms underlying root development are complex and involve many phytohormones, of which auxin is the most important. This study investigates the involvement of OsGER4, a putative Germin-like protein, in auxin-mediated crown root development in rice. The expression study of OsGER4 in the crl1 mutant confirms that OsGER4 is connected to the CRL1 signaling pathway- a master regulator for crown root development. Transgenic rice carrying the ProGER4::GUS reporter gene revealed that OsGER4 is mainly expressed in the initiation and emergence zone of the crown and lateral root, such as epidermal cell, vasculature, and primordial under auxin treatment condition. Moreover, fewer crown roots of osger4 knockout mutant lines than the wild-type under auxin treatment suggests that OsGER4 might function as a regulator limiting auxin flux to root ...
Chrysanthemum is a high commercial value plant, which is accounted for a large proportion of the ... more Chrysanthemum is a high commercial value plant, which is accounted for a large proportion of the total export of cut flowers in Vietnam. Therefore, the demand for seedling plants is very high. As a short-day plant, chrysanthemum requires supplemental lighting during the 2nd propagation stage in the autumn-winter period. This study aimed to evaluate the applicability of energy-saving LED lights to replace traditional lights in the propagation of some chrysanthemum varieties in Hanoi. The results show that the B3R7 660 nm LED (450 nm blue LED and 660 nm red LED combined in the ratio 7:3) was the most suitable for the 2nd propagation process in the greenhouse of 3 varieties (Pha le Chrysanthemum indicumyellow, Kim cuong C. morifolium cv Kim Cuong and Farm C. morifolium Hibiki). Plants grown under this LED light had a higher shoot multiplication efficiency than those under compact lamps (67.91, 18.47, and 26.89% respectively for Pha le, Kim cuong and Farm). Besides, the quality of seedl...
Trong công tác chọn tạo giống cây trồng, chuyển gene là một trong những cách tiếp cận hiệu quả nh... more Trong công tác chọn tạo giống cây trồng, chuyển gene là một trong những cách tiếp cận hiệu quả nhất vì đưa trực tiếp gene mục tiêu vào giống cây nghiên cứu. Cách tiếp cận này cần phải thông qua hệ thống nuôi cấy mô hoàn thiện. Một trong những trở ngại lớn nhất của nuôi cấy mô ở các giống lúa thuộc nhóm indica là quá trình chuyển hóa các mô sẹo thành phôi vô tính và có tỷ lệ tái sinh rất thấp. Chính vì vậy, nghiên cứu được tiến hành nhằm xây dựng và đánh giá hiệu quả quy trình tái sinh cho giống lúa Nàng Thơm Chợ Đào (NTCD) cũng như mở rộng ứng dụng cho các giống lúa khác ở vùng Đồng bằng sông Cửu Long. Trong nghiên cứu này, khả năng tạo mô sẹo trên môi trường N6D (N6+2,4D) với giống NTCD đạt hiệu quả từ 95 tới 97%. Môi trường phù hợp với giai đoạn phát sinh phôi soma và tái sinh chồi lần lượt là MS+NAA+kinetin và MS+B, với tỷ lệ tạo chồi đạt 31,01%. Tỷ lệ tạo cây hoàn chỉnh đạt 30,71%, cây tái sinh được chuyển thành công ra trồng và chăm sóc trong điều kiện vườn ươm. Kết quả này là ...
Objective: This study was performed to modify the coding sequence of a novel Cry2Ab39 gene derive... more Objective: This study was performed to modify the coding sequence of a novel Cry2Ab39 gene derived from B. thuringiensis serovar canadensis strain SP142 in Vietnam and investigate the expression of this codon-optimized gene in Nicotiana benthamiana （N. benthamiana) leaves. Methods: The Cry2Ab39 gene sequence (Genebank No. MN319700.1) was modified based on codon bias of N. benthamiana. A sequence coding the legumin B4 signal peptide and a His-tag coding sequence followed by endoplasmic reticulum (ER) retention signal KDEL were added to the 5' and 3' ends of the codon optimized Cry2Ab39 gene, respectively. The binary vector pFGC5941 harboring optimized Cry2Ab39 under the control of either CaMV 35S or Arabidopsis rbcS1A promoters was constructed and used for transient gene expression in N. benthamiana via agroinfiltration. Results: The native Cry2Ab39 gene were optimized based on codon usage of N. benthamiana along with a preference of G/C-containing codons to increase the over...
This study was performed to investigate the effects of different supplemental light spectra and d... more This study was performed to investigate the effects of different supplemental light spectra and doses (duration and illuminance) on the essential oil of basil (Ocimum basilicum L.) cultivated in the net-house in Vietnam during four months. Ten samples of basil aerial parts were hydrodistilled to obtain essential oils which had the average yields from 0.88 to 1.30% (v/w, dry). The oils analyzed using GC-FID and GC-MS showed that the main component was methyl chavicol (87.4–90.6%) with the highest values found in the oils of basil under lighting conditions of 6 h/day and 150–200 µmol·m−2·s−1. Additional lighting conditions caused the significant differences (p < 0.001) in basil biomass and oil production with the highest values found in the oils of basil under two conditions of (1) 71% Red: 20% Blue: 9.0% UVA in at 120 μmol·m−2·s−1 in 6 h/day and (2) 43.5% Red: 43.5% Blue: 8.0% Green: 5.0% Far-Red at 100 μmol·m−2·s−1 in 6 h/day. The oils of basil in some formulas showed weak inhibi...
BACKGROUND In the alkaloid biosynthetic pathways of Stephania and Rannunculaceae, columbamine O-m... more BACKGROUND In the alkaloid biosynthetic pathways of Stephania and Rannunculaceae, columbamine O-methyltransferase (CoOMT) is an important enzyme that catalyses the formation of the tetrahydropalmatin (rotundin) biosynthesis pathway. In this study, the transgenic construct pBI121-35S-CoOMT-cmyc-Kdel was designed successfully. METHODS AND RESULTS The real-time RT-PCR results proved that the CoOMT transgene was successfully introduced into Nicotiana tabacum L. plants and produced mRNA. Its transcription levels in three transgenic tobacco lines, T0-7, T0-9, and T0-20, in the T0 generation were higher than those in wild-type tobacco plants. By analysing Western blots and ELISAs, three T0 generation transgenic tobacco lines also expressed recombinant CoOMT (rCoOMT) protein with a molecular weight of approximately 40 kDa, and its contents ranged from 0.048 μg mg-1 to 0.177 μg mg-1. These data illustrated that the CoOMT transgene was expressed; thus, the rCoOMT protein synthesis efficiency increased significantly in comparison with that of the wild-type tobacco plants. The total alkaloid contents ranged from 2.12 g 100 g-1 (of dry weight) to 3.88 g 100 g-1 (of dry weight). The T0-20 plant had the highest total alkaloid content (3.88 g 100 g-1 of dry weight), followed by the T0-7 line (2.75 g 100 g-1 of dry weight). The total alkaloid contents of the CoOMT transgenic tobacco lines increased by approximately 1.09-1.83-fold compared to the wild-type tobacco plants. CONCLUSIONS This is the first study on the transformation and expression of the CoOMT gene in N. tabacum plants. Initial results of the analysis of transgenic plants proved that the transgenic structure pBI121- CoOMT-Cmyc-Kdel can be used for transformation into Stephania plants.
Hairy root induction system has been applied in various plant species as an effective method to s... more Hairy root induction system has been applied in various plant species as an effective method to study gene expression and function due to its fast-growing and high genetic stability. Recently, these systems have shown to be an effective tool to evaluate activities of CRISPR/Cas9 systems for genome editing. In this study, Rhizobium rhizogenes mediated hairy root induction was optimized to provide an effective tool for validation of plant transformation vector, CRISPR/Cas9 construct activities as well as selection of targeted gRNAs for gene editing in cucumber (Cucumis sativus L.). Under the optimized conditions including OD650 at 0.4 for infection and 5 days of co-cultivation, the highest hairy root induction frequency reached 100% for the cucumber variety Choka F1. This procedure was successfully utilized to overexpress a reporter gene (gus) and induce mutations in two Lotus japonicus ROOTHAIRLESS1 homolog genes CsbHLH66 and CsbHLH82 using CRISPR/Cas9 system. For induced mutation, a...
Figure S1. Identification of edited GmFAD2 genes of soybean hairy roots using PCR-based genotypin... more Figure S1. Identification of edited GmFAD2 genes of soybean hairy roots using PCR-based genotyping. Figure S2. Partial deletions and insertions detected in GmFAD2 genes. Figure S3. Genotyping of homozygous expected deletions and transgenes in T2 generation of event ND1–11. Figure S4. Inheritance of GmFAD2 mutations in T2 progenies of event ND1–11. Figure S5. Sequencing results of off-target and flanking regions. Table S1. Primer sequences for genotyping GmFAD2 genes. Table S2. Segregation of bar and Cas9 in T2 progenies derived from event ND1–11. Cas9 was detected by PCR while Bar was detected by PCR and leaf painting. Table S3. Potential off-target mutations in transgenic T2 plants derived from event ND1–11. Table S4. Protein and oil content in ND1-11-14 and wild type (Williams 82 and Maverick) seeds. Measurements were performed over two years under greenhouse (2017) and field (2018) conditions. (PDF 862 kb)
Overexpression of GA20 oxidase gene has been a recent trend for improving plant growth and biomas... more Overexpression of GA20 oxidase gene has been a recent trend for improving plant growth and biomass. Constitutive expression of GA20ox has successfully improved plant growth and biomass in several plant species. However, the constitutive expression of this gene causes side-effects, such as reduced leaf size and stem diameter, etc. To avoid these effects, we identified and employed different tissue-specific promoters for GA20ox overexpression. In this study, we examined the utility of At1g promoter to drive the expression of GUS (β-glucuronidase) reporter and AtGA20ox genes in tobacco and Melia azedarach . Histochemical GUS assays and quantitative real-time-PCR results in tobacco showed that At1g was a root-preferential promoter whose expression was particularly strong in root tips. The ectopic expression of AtGA20ox gene under the control of At1g promoter showed improved plant growth and biomass of both tobacco and M. azedarach transgenic plants. Stem length as well as stem and root ...
Abstract Soil is contaminated with salinity, which inhibits plant growth and development and redu... more Abstract Soil is contaminated with salinity, which inhibits plant growth and development and reduces crop yields. The DREB (dehydration responsive element binding protein) gene responds to salt stresses through enhanced transcriptional expression and activation of genes involved in plant salinity resistance. In this study, we present the results of the analysis of the expression of the GmDREB6 transgene, a gene that encodes the soybean DREB6 transcription factor, regulating the transcription of the NtP5CS and NtCLC genes in transgenic tobacco under salt stress conditions. The transcription of GmDREB6, NtP5CS, and NtCLC in transgenic tobacco lines was confirmed by qRT-PCR. Under salt stress conditions, the GmDREB6 gene transcription levels in the transgenic tobacco lines L1 and L9 had increased from 2.40- to 3.22- fold compared with the condition without salinity treatment. Two transgenic lines, L1 and L9, had transcription levels of the P5CS gene that had increased from 1.24- to 3.60- fold compared with WT plants. For the NtCLC gene, under salt stress conditions, the transgenic lines had transcription levels that had increased by 3.65–4.54 (fold) compared with WT plants (P
Background: Overexpression of GA20 oxidase gene has been a recent trend for improving plant growt... more Background: Overexpression of GA20 oxidase gene has been a recent trend for improving plant growth and biomass. Constitutive expression of GA20ox has successfully improved plant growth and biomass in several plant species. However, the constitutive expression of this gene causes side-effects, such as reduced leaf size and stem diameters, etc. To avoid these effects, different tissue-specific promoters had been identified and employed for GA20ox overexpression. In this study, we demonstrate the potential of At1g, a root-preferential promoter, for GA20ox expression to enhance plant biomass in tobacco and Melia azedarach.Results: We examined the utility of At1g promoter to drive the expression of GUS (β-1,4-glucuronidase) reporter and GA20ox genes in tobacco and Melia azedarach. Histochemical GUS assays in tobacco showed that At1g was a root-preferential promoter whose expression was particularly strong in root tips. The ectopic expression of AtGA20ox gene under the control of At1g pro...
Currently, drought stress has been known as a critical abiotic factors limiting crop production a... more Currently, drought stress has been known as a critical abiotic factors limiting crop production around the world. Several genes associated with water stresses, including codA gene encoded for choline oxidase, have been characterized and transferred to several important crops to enhance the tolerance to drought stress. Therefore, in this study, codA gene under the regulation of a drought inducible promoter-rd29A was introduced into soybean using the Agrobacterium-mediated method. The presence of codA gene in transgenic soybeans was confirmed by PCR and Southern hybridization. Under drought stress treatments, transgenic soybeans showed an enhanced seedling growth and plant biomass as compared to nontransformed soybeans. Moreover, the contents of glycine betaine and free proline, two major organic osmolytes as well as peroxidase activity were found increased in codA transgenic soybean. The malondialdehyde content in transformed soybeans decreased by less than 50% relative to control soybean plants under drought stress conditions. This is the first report on the utilization of the codA gene to improve drought tolerance in soybeans.
A rapid and efficient Agrobacterium -mediated transformation system in sorghum has been developed... more A rapid and efficient Agrobacterium -mediated transformation system in sorghum has been developed employing standard binary vectors and bar gene as a selectable marker. Sorghum (Sorghum bicolor) is an important food and biofuel crop worldwide, for which improvements in genetic transformation are needed to study its biology and facilitate agronomic and commercial improvement. Here, we report optimization of regeneration and transformation of public sorghum genotype P898012 using standard binary vectors and bar gene as a selectable marker. The tissue culture regeneration time frame has been reduced to 7-12 weeks with a yield of over 18 plants per callus, and the optimized transformation system employing Agrobacterium tumefaciens strain AGL1 and the bar with a MAS promoter achieved an average frequency over 14 %. Of randomly analyzed independent transgenic events, 40-50 % carry single copy of integrated T-DNA. Some independent transgenic events were derived from the same embryogenic ca...
A rapid and efficient Agrobacterium -mediated transformation system in sorghum has been developed... more A rapid and efficient Agrobacterium -mediated transformation system in sorghum has been developed employing standard binary vectors and bar gene as a selectable marker. Sorghum (Sorghum bicolor) is an important food and biofuel crop worldwide, for which improvements in genetic transformation are needed to study its biology and facilitate agronomic and commercial improvement. Here, we report optimization of regeneration and transformation of public sorghum genotype P898012 using standard binary vectors and bar gene as a selectable marker. The tissue culture regeneration time frame has been reduced to 7-12 weeks with a yield of over 18 plants per callus, and the optimized transformation system employing Agrobacterium tumefaciens strain AGL1 and the bar with a MAS promoter achieved an average frequency over 14 %. Of randomly analyzed independent transgenic events, 40-50 % carry single copy of integrated T-DNA. Some independent transgenic events were derived from the same embryogenic callus lines, but a 3:1 Mendelian segregation ratio was found in all transgenic events with single copy as estimated by Southern blots. The system described here should facilitate studies of sorghum biology and agronomic improvement.
In rice (Oryza sativa L.), the root system plays different essential roles, from water and nutrie... more In rice (Oryza sativa L.), the root system plays different essential roles, from water and nutrient uptake to responding to environmental signals. The mechanisms underlying root development are complex and involve many phytohormones, of which auxin is the most important. This study investigates the involvement of OsGER4, a putative Germin-like protein, in auxin-mediated crown root development in rice. The expression study of OsGER4 in the crl1 mutant confirms that OsGER4 is connected to the CRL1 signaling pathway- a master regulator for crown root development. Transgenic rice carrying the ProGER4::GUS reporter gene revealed that OsGER4 is mainly expressed in the initiation and emergence zone of the crown and lateral root, such as epidermal cell, vasculature, and primordial under auxin treatment condition. Moreover, fewer crown roots of osger4 knockout mutant lines than the wild-type under auxin treatment suggests that OsGER4 might function as a regulator limiting auxin flux to root ...
Chrysanthemum is a high commercial value plant, which is accounted for a large proportion of the ... more Chrysanthemum is a high commercial value plant, which is accounted for a large proportion of the total export of cut flowers in Vietnam. Therefore, the demand for seedling plants is very high. As a short-day plant, chrysanthemum requires supplemental lighting during the 2nd propagation stage in the autumn-winter period. This study aimed to evaluate the applicability of energy-saving LED lights to replace traditional lights in the propagation of some chrysanthemum varieties in Hanoi. The results show that the B3R7 660 nm LED (450 nm blue LED and 660 nm red LED combined in the ratio 7:3) was the most suitable for the 2nd propagation process in the greenhouse of 3 varieties (Pha le Chrysanthemum indicumyellow, Kim cuong C. morifolium cv Kim Cuong and Farm C. morifolium Hibiki). Plants grown under this LED light had a higher shoot multiplication efficiency than those under compact lamps (67.91, 18.47, and 26.89% respectively for Pha le, Kim cuong and Farm). Besides, the quality of seedl...
Trong công tác chọn tạo giống cây trồng, chuyển gene là một trong những cách tiếp cận hiệu quả nh... more Trong công tác chọn tạo giống cây trồng, chuyển gene là một trong những cách tiếp cận hiệu quả nhất vì đưa trực tiếp gene mục tiêu vào giống cây nghiên cứu. Cách tiếp cận này cần phải thông qua hệ thống nuôi cấy mô hoàn thiện. Một trong những trở ngại lớn nhất của nuôi cấy mô ở các giống lúa thuộc nhóm indica là quá trình chuyển hóa các mô sẹo thành phôi vô tính và có tỷ lệ tái sinh rất thấp. Chính vì vậy, nghiên cứu được tiến hành nhằm xây dựng và đánh giá hiệu quả quy trình tái sinh cho giống lúa Nàng Thơm Chợ Đào (NTCD) cũng như mở rộng ứng dụng cho các giống lúa khác ở vùng Đồng bằng sông Cửu Long. Trong nghiên cứu này, khả năng tạo mô sẹo trên môi trường N6D (N6+2,4D) với giống NTCD đạt hiệu quả từ 95 tới 97%. Môi trường phù hợp với giai đoạn phát sinh phôi soma và tái sinh chồi lần lượt là MS+NAA+kinetin và MS+B, với tỷ lệ tạo chồi đạt 31,01%. Tỷ lệ tạo cây hoàn chỉnh đạt 30,71%, cây tái sinh được chuyển thành công ra trồng và chăm sóc trong điều kiện vườn ươm. Kết quả này là ...
Objective: This study was performed to modify the coding sequence of a novel Cry2Ab39 gene derive... more Objective: This study was performed to modify the coding sequence of a novel Cry2Ab39 gene derived from B. thuringiensis serovar canadensis strain SP142 in Vietnam and investigate the expression of this codon-optimized gene in Nicotiana benthamiana （N. benthamiana) leaves. Methods: The Cry2Ab39 gene sequence (Genebank No. MN319700.1) was modified based on codon bias of N. benthamiana. A sequence coding the legumin B4 signal peptide and a His-tag coding sequence followed by endoplasmic reticulum (ER) retention signal KDEL were added to the 5' and 3' ends of the codon optimized Cry2Ab39 gene, respectively. The binary vector pFGC5941 harboring optimized Cry2Ab39 under the control of either CaMV 35S or Arabidopsis rbcS1A promoters was constructed and used for transient gene expression in N. benthamiana via agroinfiltration. Results: The native Cry2Ab39 gene were optimized based on codon usage of N. benthamiana along with a preference of G/C-containing codons to increase the over...
This study was performed to investigate the effects of different supplemental light spectra and d... more This study was performed to investigate the effects of different supplemental light spectra and doses (duration and illuminance) on the essential oil of basil (Ocimum basilicum L.) cultivated in the net-house in Vietnam during four months. Ten samples of basil aerial parts were hydrodistilled to obtain essential oils which had the average yields from 0.88 to 1.30% (v/w, dry). The oils analyzed using GC-FID and GC-MS showed that the main component was methyl chavicol (87.4–90.6%) with the highest values found in the oils of basil under lighting conditions of 6 h/day and 150–200 µmol·m−2·s−1. Additional lighting conditions caused the significant differences (p < 0.001) in basil biomass and oil production with the highest values found in the oils of basil under two conditions of (1) 71% Red: 20% Blue: 9.0% UVA in at 120 μmol·m−2·s−1 in 6 h/day and (2) 43.5% Red: 43.5% Blue: 8.0% Green: 5.0% Far-Red at 100 μmol·m−2·s−1 in 6 h/day. The oils of basil in some formulas showed weak inhibi...
BACKGROUND In the alkaloid biosynthetic pathways of Stephania and Rannunculaceae, columbamine O-m... more BACKGROUND In the alkaloid biosynthetic pathways of Stephania and Rannunculaceae, columbamine O-methyltransferase (CoOMT) is an important enzyme that catalyses the formation of the tetrahydropalmatin (rotundin) biosynthesis pathway. In this study, the transgenic construct pBI121-35S-CoOMT-cmyc-Kdel was designed successfully. METHODS AND RESULTS The real-time RT-PCR results proved that the CoOMT transgene was successfully introduced into Nicotiana tabacum L. plants and produced mRNA. Its transcription levels in three transgenic tobacco lines, T0-7, T0-9, and T0-20, in the T0 generation were higher than those in wild-type tobacco plants. By analysing Western blots and ELISAs, three T0 generation transgenic tobacco lines also expressed recombinant CoOMT (rCoOMT) protein with a molecular weight of approximately 40 kDa, and its contents ranged from 0.048 μg mg-1 to 0.177 μg mg-1. These data illustrated that the CoOMT transgene was expressed; thus, the rCoOMT protein synthesis efficiency increased significantly in comparison with that of the wild-type tobacco plants. The total alkaloid contents ranged from 2.12 g 100 g-1 (of dry weight) to 3.88 g 100 g-1 (of dry weight). The T0-20 plant had the highest total alkaloid content (3.88 g 100 g-1 of dry weight), followed by the T0-7 line (2.75 g 100 g-1 of dry weight). The total alkaloid contents of the CoOMT transgenic tobacco lines increased by approximately 1.09-1.83-fold compared to the wild-type tobacco plants. CONCLUSIONS This is the first study on the transformation and expression of the CoOMT gene in N. tabacum plants. Initial results of the analysis of transgenic plants proved that the transgenic structure pBI121- CoOMT-Cmyc-Kdel can be used for transformation into Stephania plants.
Hairy root induction system has been applied in various plant species as an effective method to s... more Hairy root induction system has been applied in various plant species as an effective method to study gene expression and function due to its fast-growing and high genetic stability. Recently, these systems have shown to be an effective tool to evaluate activities of CRISPR/Cas9 systems for genome editing. In this study, Rhizobium rhizogenes mediated hairy root induction was optimized to provide an effective tool for validation of plant transformation vector, CRISPR/Cas9 construct activities as well as selection of targeted gRNAs for gene editing in cucumber (Cucumis sativus L.). Under the optimized conditions including OD650 at 0.4 for infection and 5 days of co-cultivation, the highest hairy root induction frequency reached 100% for the cucumber variety Choka F1. This procedure was successfully utilized to overexpress a reporter gene (gus) and induce mutations in two Lotus japonicus ROOTHAIRLESS1 homolog genes CsbHLH66 and CsbHLH82 using CRISPR/Cas9 system. For induced mutation, a...
Figure S1. Identification of edited GmFAD2 genes of soybean hairy roots using PCR-based genotypin... more Figure S1. Identification of edited GmFAD2 genes of soybean hairy roots using PCR-based genotyping. Figure S2. Partial deletions and insertions detected in GmFAD2 genes. Figure S3. Genotyping of homozygous expected deletions and transgenes in T2 generation of event ND1–11. Figure S4. Inheritance of GmFAD2 mutations in T2 progenies of event ND1–11. Figure S5. Sequencing results of off-target and flanking regions. Table S1. Primer sequences for genotyping GmFAD2 genes. Table S2. Segregation of bar and Cas9 in T2 progenies derived from event ND1–11. Cas9 was detected by PCR while Bar was detected by PCR and leaf painting. Table S3. Potential off-target mutations in transgenic T2 plants derived from event ND1–11. Table S4. Protein and oil content in ND1-11-14 and wild type (Williams 82 and Maverick) seeds. Measurements were performed over two years under greenhouse (2017) and field (2018) conditions. (PDF 862 kb)
Overexpression of GA20 oxidase gene has been a recent trend for improving plant growth and biomas... more Overexpression of GA20 oxidase gene has been a recent trend for improving plant growth and biomass. Constitutive expression of GA20ox has successfully improved plant growth and biomass in several plant species. However, the constitutive expression of this gene causes side-effects, such as reduced leaf size and stem diameter, etc. To avoid these effects, we identified and employed different tissue-specific promoters for GA20ox overexpression. In this study, we examined the utility of At1g promoter to drive the expression of GUS (β-glucuronidase) reporter and AtGA20ox genes in tobacco and Melia azedarach . Histochemical GUS assays and quantitative real-time-PCR results in tobacco showed that At1g was a root-preferential promoter whose expression was particularly strong in root tips. The ectopic expression of AtGA20ox gene under the control of At1g promoter showed improved plant growth and biomass of both tobacco and M. azedarach transgenic plants. Stem length as well as stem and root ...
Abstract Soil is contaminated with salinity, which inhibits plant growth and development and redu... more Abstract Soil is contaminated with salinity, which inhibits plant growth and development and reduces crop yields. The DREB (dehydration responsive element binding protein) gene responds to salt stresses through enhanced transcriptional expression and activation of genes involved in plant salinity resistance. In this study, we present the results of the analysis of the expression of the GmDREB6 transgene, a gene that encodes the soybean DREB6 transcription factor, regulating the transcription of the NtP5CS and NtCLC genes in transgenic tobacco under salt stress conditions. The transcription of GmDREB6, NtP5CS, and NtCLC in transgenic tobacco lines was confirmed by qRT-PCR. Under salt stress conditions, the GmDREB6 gene transcription levels in the transgenic tobacco lines L1 and L9 had increased from 2.40- to 3.22- fold compared with the condition without salinity treatment. Two transgenic lines, L1 and L9, had transcription levels of the P5CS gene that had increased from 1.24- to 3.60- fold compared with WT plants. For the NtCLC gene, under salt stress conditions, the transgenic lines had transcription levels that had increased by 3.65–4.54 (fold) compared with WT plants (P
Background: Overexpression of GA20 oxidase gene has been a recent trend for improving plant growt... more Background: Overexpression of GA20 oxidase gene has been a recent trend for improving plant growth and biomass. Constitutive expression of GA20ox has successfully improved plant growth and biomass in several plant species. However, the constitutive expression of this gene causes side-effects, such as reduced leaf size and stem diameters, etc. To avoid these effects, different tissue-specific promoters had been identified and employed for GA20ox overexpression. In this study, we demonstrate the potential of At1g, a root-preferential promoter, for GA20ox expression to enhance plant biomass in tobacco and Melia azedarach.Results: We examined the utility of At1g promoter to drive the expression of GUS (β-1,4-glucuronidase) reporter and GA20ox genes in tobacco and Melia azedarach. Histochemical GUS assays in tobacco showed that At1g was a root-preferential promoter whose expression was particularly strong in root tips. The ectopic expression of AtGA20ox gene under the control of At1g pro...
Currently, drought stress has been known as a critical abiotic factors limiting crop production a... more Currently, drought stress has been known as a critical abiotic factors limiting crop production around the world. Several genes associated with water stresses, including codA gene encoded for choline oxidase, have been characterized and transferred to several important crops to enhance the tolerance to drought stress. Therefore, in this study, codA gene under the regulation of a drought inducible promoter-rd29A was introduced into soybean using the Agrobacterium-mediated method. The presence of codA gene in transgenic soybeans was confirmed by PCR and Southern hybridization. Under drought stress treatments, transgenic soybeans showed an enhanced seedling growth and plant biomass as compared to nontransformed soybeans. Moreover, the contents of glycine betaine and free proline, two major organic osmolytes as well as peroxidase activity were found increased in codA transgenic soybean. The malondialdehyde content in transformed soybeans decreased by less than 50% relative to control soybean plants under drought stress conditions. This is the first report on the utilization of the codA gene to improve drought tolerance in soybeans.
A rapid and efficient Agrobacterium -mediated transformation system in sorghum has been developed... more A rapid and efficient Agrobacterium -mediated transformation system in sorghum has been developed employing standard binary vectors and bar gene as a selectable marker. Sorghum (Sorghum bicolor) is an important food and biofuel crop worldwide, for which improvements in genetic transformation are needed to study its biology and facilitate agronomic and commercial improvement. Here, we report optimization of regeneration and transformation of public sorghum genotype P898012 using standard binary vectors and bar gene as a selectable marker. The tissue culture regeneration time frame has been reduced to 7-12 weeks with a yield of over 18 plants per callus, and the optimized transformation system employing Agrobacterium tumefaciens strain AGL1 and the bar with a MAS promoter achieved an average frequency over 14 %. Of randomly analyzed independent transgenic events, 40-50 % carry single copy of integrated T-DNA. Some independent transgenic events were derived from the same embryogenic ca...
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