scholarly journals Expression of thaumatin, a new type of alternative sweetener in rice

2020 ◽  
Vol 48 (3) ◽  
pp. 1276-1291
Author(s):  
Shahina AKTER ◽  
Md. Amdadul HUQ ◽  
Yu-Jin JUNG ◽  
Kwon-Kyoo KANG
Keyword(s):  
Transgenic Rice ◽  
Oryza Sativa L ◽  
Plasmid Vector ◽  
Single Copy ◽  
Cauliflower Mosaic Virus ◽  
Pcr Analysis ◽  
35S Promoter ◽  
Alternative Sweetener ◽  
Sweet Proteins ◽  
Transgenic Lines

  Sweet proteins are the natural alternative to the artificial sweeteners as well as flavor enhancers. Among other sweet protein, thaumatin protein was isolated from Thaumatococcus daniellii Benth plant fruit. In this study, pinII Ti plasmid vector was constructed with thaumatin gene, where thaumatin was placed under the control of the duel cauliflower mosaic virus 35S promoter into rice (Oryza sativa L. var. japonica cv. ‘Dongjinbyeo’) by Agrobacterium-mediated transformation to generate transgenic plants. Thirteen plant lines were regenerated and the transgenic rice lines were confirmed by different molecular analysis. The genomic PCR result revealed that all of the plant lines were transgenic. The single copy and intergenic plant lines were selected by Taqman PCR analysis and FST analysis, respectively. Expression of thaumatin gene in transgenic rice resulted in the accumulation of thaumatin protein in the leave. Thaumatin protein was also accumulated in leave of T1 generation. Sensory analysis result suggested that the thaumatin protein expressing transgenic lines exerted sweet tasting activity. These results demonstrated that thaumatin was expressed in transgenic rice plants.

scholarly journals Constitutive Expression of Pea Defense Gene DRR206 Confers Resistance to Blackleg (Leptosphaeria maculans) Disease in Transgenic Canola (Brassica napus)

1999 ◽  
Vol 12 (5) ◽  
pp. 410-418 ◽  
Author(s):  
Yaping Wang ◽  
Goska Nowak ◽  
David Culley ◽  
Lee A. Hadwiger ◽  
Brian Fristensky
Keyword(s):  
Brassica Napus ◽  
Transgenic Plants ◽  
Leptosphaeria Maculans ◽  
Constitutive Expression ◽  
Single Copy ◽  
Cauliflower Mosaic Virus ◽  
Adult Plant ◽  
35S Promoter ◽  
Transgenic Lines

To identify genes effective against the blackleg fungus Leptosphaeria maculans (Phoma lingam), we have transformed canola (Brassica napus) with four pea (Pisum sativum) genes under constitutive control by the cauliflower mosaic virus 35S promoter: PR10.1, chitinase, DRR206, and defensin. Transgenic lines containing single-copy T-DNA insertions for each gene were screened for both seedling (cotyledonary) and adult plant resistance. Lines for which pea DRR206 mRNA was expressed showed decreased disease scores, compared with non-expressing transgenic lines. Transgenic plants expressing pea defensin showed a slight enhancement of resistance, while for PR10 and chitinase transgenics there was little or no enhancement of resistance. Resistance to L. maculans cosegregated with DRR206 transgenes. Extracts from DRR206 and defensin transgenic plants inhibited fungal germination in vitro. DRR206 transgenic plants also demonstrated decreased hyphal growth at inoculation sites. While the precise function of DRR206 remains to be determined, these results suggest that it does play an important role in defense against fungi.

Aberrant promoter methylation occurred from multicopy transgene and SU(VAR)3 - 9 homolog 9 (SUVH9) gene in transgenic Nicotiana benthamiana

2012 ◽  
Vol 39 (9) ◽  
pp. 764 ◽  
Author(s):  
Gi-Ho Lee ◽  
Seong-Han Sohn ◽  
Eun-Young Park ◽  
Young-Doo Park
Keyword(s):  
Gene Silencing ◽  
Nicotiana Benthamiana ◽  
Fluorescent Protein ◽  
Cauliflower Mosaic Virus ◽  
Camv 35S Promoter ◽  
35S Promoter ◽  
Camv 35S ◽  
Histone Deacetylase 1 ◽  
Transgenic Lines ◽  
Green Fluorescent

The chemical modification of DNA by methylation is a heritable trait and can be subsequently reversed without altering the original DNA sequence. Methylation can reduce or silence gene expression and is a component of a host’s defence response to foreign nucleic acids. In our study, we employed a plant transformation strategy using Nicotiana benthamiana Domin to study the heritable stability of the introduced transgenes. Through the introduction of the cauliflower mosaic virus (CaMV) 35S promoter and the green fluorescent protein (GFP) reporter gene, we demonstrated that this introduced promoter often triggers a homology-dependent gene-silencing (HDGS) response. These spontaneous transgene-silencing phenomena are due to methylation of the CaMV 35S promoter CAAT box during transgenic plant growth. This process is catalysed by SU(VAR)3–9 homologue 9 (SUVH9), histone deacetylase 1 (HDA1) and domains rearranged methylase 2 (DRM2). In particular, we showed from our data that SUVH9 is the key regulator of methylation activity in epigenetically silenced GFP transgenic lines; therefore, our findings demonstrate that an introduced viral promoter and transgene can be subject to a homology-dependent gene-silencing mechanism that can downregulate its expression and negatively influence the heritable stability of the transgene.

Transgenic Rice Expressing Isoflavone Synthase Gene from Soybean Shows Resistance against Blast Fungus (Magnaporthe oryzae)

Plant Disease ◽  
2021 ◽  
Author(s):  
Suresh Pokhrel ◽  
Sathish K Ponniah ◽  
Yulin Jia ◽  
Oliver Yu ◽  
Muthusamy Manoharan
Keyword(s):  
Disease Resistance ◽  
Transgenic Rice ◽  
Magnaporthe Oryzae ◽  
Plant Disease Resistance ◽  
Rice Blast ◽  
Plant Disease ◽  
Plant Secondary Metabolites ◽  
Single Copy ◽  
Transgenic Lines ◽  
Isoflavone Synthase

The isoflavones are a group of plant secondary metabolites primarily synthesized in legumes and are known for their role in improving human health and plant disease resistance. The isoflavones, especially genistein, act as precursors for the production of phytoalexins, which may induce broad-spectrum disease resistance in plants. In the present study, we screened transgenic rice lines expressing the isoflavone synthase (GmIFS1) gene from soybean for rice blast (Magnaporthe oryzae) resistance. Two homozygous transgenic lines (I2 and I10), based on single copy gene integration, were identified. The expression of GmIFS1 in transgenic lines was confirmed by qRT-PCR. Genistein was detected in the transgenic lines using LC-MS/MS. Subsequently, the transgenic lines were evaluated against the rice blast pathogen, isolate YJ54 (race IB-54). The results indicated that more than 60% of the plants in both the lines (I2 and I10) showed resistance against the blast pathogen. The progenies of one of the resistant transgenic lines (I10) also showed more than 65% resistance against rice blast. The resistance of these transgenic lines against rice blast may be attributed to the synthesis of isoflavone (genistein) in rice.

scholarly journals Paxillus involutus Forms an Ectomycorrhizal Symbiosis and Enhances Survival of PtCOMT-modified Betula pendula in vitro

2008 ◽  
Vol 57 (1-6) ◽  
pp. 235-242 ◽  
Author(s):  
H. Tiimonen ◽  
T. Aronen ◽  
T. Laakso ◽  
P. Saranpää ◽  
V. Chiang ◽  
...  
Keyword(s):  
Populus Tremuloides ◽  
Betula Pendula ◽  
Root Formation ◽  
Lateral Roots ◽  
Cauliflower Mosaic Virus ◽  
35S Promoter ◽  
Paxillus Involutus ◽  
Transgenic Lines ◽  
G Ratio

Abstract The ability of the PtCOMT (caffeate/5-hydroxyferulate O-methyltransferase from Populus tremuloides L.) - modified Betula pendula Roth. lines to form symbiosis with an ectomycorrhizal (ECM) fungus Paxillus involutus Batsch Fr. was studied in vitro. Lignin precursor gene PtCOMT was introduced into two B. pendula clones under the control of the cauliflower mosaic virus 35S promoter or the promoter of the sunflower polyubiquitin gene UbB1. Of the four transgenic lines, one 35SPtCOMT line (23) had a decreased syringyl/guaiacyl (S/G) ratio of root lignin, and two UbB1-PtCOMT lines (110 and 130) retarded root growth compared to the control clone. Both control clones and all transgenic lines were able to form ECMs with P. involutus, but the transgenic lines differed from the controls in the characteristics of the ECMs. The number of lateral roots covered with fungal hyphae and/or development of a Hartig net (HN) were reduced in line 23 with a decreased S/G ratio, and in lines 110 and 130 with slower root formation and changed root morphology, respectively. However, line 23 benefited more from the inoculation in lateral root formation than the control, and in lines 110 and 130 the percentage of viable plants increased most due to inoculation. The results show that B. pendula plants genetically transformed with the lignin gene PtCOMT could form mycorrhizal symbiosis regardless of changes in either the root S/G ratio or development. The benefits of the symbiosis were variable even in the closed in vitro system, and dependent on the clone or transgenic line and the ECM fungal symbiont.

scholarly journals Molecular Breeding of Transgenic Rice Plants Expressing a Bacterial Chlorocatechol Dioxygenase Gene

2002 ◽  
Vol 68 (8) ◽  
pp. 4061-4066 ◽  
Author(s):  
Masami Shimizu ◽  
Tetsuya Kimura ◽  
Takayoshi Koyama ◽  
Katsuhisa Suzuki ◽  
Naoto Ogawa ◽  
...  
Keyword(s):  
Transgenic Rice ◽  
Molecular Breeding ◽  
Ralstonia Eutropha ◽  
Leaf Tissue ◽  
Cauliflower Mosaic Virus ◽  
35S Promoter ◽  
Gene Encoding ◽  
Rice Plants ◽  
Transgenic Rice Plants ◽  
Dioxygenase Gene

ABSTRACT The cbnA gene encoding the chlorocatechol dioxygenase gene from Ralstonia eutropha NH9 was introduced into rice plants. The cbnA gene was expressed in transgenic rice plants under the control of a modified cauliflower mosaic virus 35S promoter. Western blot analysis using anti-CbnA protein indicated that the cbnA gene was expressed in leaf tissue, roots, culms, and seeds. Transgenic rice calluses expressing the cbnA gene converted 3-chlorocatechol to 2-chloromucote efficiently. Growth and morphology of the transgenic rice plants expressing the cbnA gene were not distinguished from those of control rice plants harboring only a Ti binary vector. It is thus possible to breed transgenic plants that degrade chloroaromatic compounds in soil and surface water.

scholarly journals Senescence-Specific Expression of RAmy1A Accelerates Non-structural Carbohydrate Remobilization and Grain Filling in Rice (Oryza sativa L.)

2021 ◽  
Vol 12 ◽  
Author(s):  
Ning Ouyang ◽  
Xuewu Sun ◽  
Yanning Tan ◽  
Zhizhong Sun ◽  
Dong Yu ◽  
...  
Keyword(s):  
Oryza Sativa ◽  
Transgenic Rice ◽  
Oryza Sativa L ◽  
Sugar Content ◽  
Soluble Sugar ◽  
Grain Filling ◽  
Leaf Sheath ◽  
Specific Expression ◽  
Total Soluble Sugar ◽  
Transgenic Lines

Remobilization of pre-anthesis NSCs (non-structural carbohydrates) is significant for effective grain filling in rice (Oryza sativa L.). However, abundant starch particles as an important component of NSCs are still present in the leaf sheath and stem at the late stage of grain filling. There are no studies on how bioengineering techniques can be used to improve the efficiency of NSC remobilization. In this study, RAmy1A was expressed under the senescence-specific promoter of SAG12, which was designed to degrade starch in the leaf sheath and stem during grain filling. RAmy1A mRNA successfully accumulated in the leaf, stem, and sheath of transgenic plants after anthesis. At the same time, the starch and total soluble sugar content in the leaf, stem, and leaf sheath were obviously decreased during the grain-filling period. The photosynthetic rate of transgenic lines was higher than that of the wild types by an average of 4.0 and 9.9%, at 5 and 10 days after flowering, respectively. In addition, the grain-filling rate of transgenic lines was faster than that of the wild types by an average of 26.09%. These results indicate an enhanced transport efficiency of NSCs from source tissues in transgenic rice. Transgenic rice also displayed accelerated leaf senescence, which was hypothesized to contribute to decreased grain weight.

scholarly journals Improved Cold-resistant Performance in Transgenic Grape (Vitis vinifera L.) Overexpressing Cold-inducible Transcription Factors AtDREB1b

HortScience ◽  
2009 ◽  
Vol 44 (1) ◽  
pp. 35-39 ◽  
Author(s):  
Wanmei Jin ◽  
Jing Dong ◽  
Yuanlei Hu ◽  
Zhongping Lin ◽  
Xuefeng Xu ◽  
...  
Keyword(s):  
Transgenic Plants ◽  
Vitis Vinifera ◽  
Blot Analysis ◽  
Freezing Point ◽  
Cold Treatment ◽  
Cauliflower Mosaic Virus ◽  
Vitis Vinifera L ◽  
35S Promoter ◽  
Transgenic Lines ◽  
H 26

Dehydration response element binding (DREB)1b is a cold-inducible transcription factor in Arabidopsis thaliana. DREB1b driven by cauliflower mosaic virus 35S promoter was genetically introduced into grape Vitis vinifera L. cv. Centennial Seedless through Agrobacterium-mediated transformation for improving its cold resistance and exploring new genetic breeding approaches to obtain cold-resistant cultivars. In this study, Southern blot analysis showed the DREB1b gene was integrated into the transgenic grapevines with one to two copies. Northern blot analysis showed the presence of DREB1b transcripts in the independent transgenic lines 3, 5, 6, and 7. Further characterization of transgenic grapevines confirmed that both electrolyte leakage conductivity and the freezing point of the transgenic plants were lower than those of wild-type plants. After the cold treatment at –4 °C for 12 h, 26% of transgenic plants wilted among which 95% plants recovered once being placed under the condition of temperature 22 to 25 °C. However, subjected to the same treatment, 98% of nontransgenic plants wilted and only 2% recovered. Our results lead to the conclusion that activity of DREB1b in the transgenic grape could significantly improve its resistance to cold stress.

Transcriptional silencing of RNAi constructs against nematode genes in Arabidopsis

Nematology ◽  
2013 ◽  
Vol 15 (5) ◽  
pp. 519-528 ◽  
Author(s):  
Tina Kyndt ◽  
Hongli Ji ◽  
Bartel Vanholme ◽  
Godelieve Gheysen
Keyword(s):  
Transcriptional Silencing ◽  
Housekeeping Genes ◽  
Cauliflower Mosaic Virus ◽  
Heterodera Schachtii ◽  
Camv 35S Promoter ◽  
35S Promoter ◽  
Expression Levels ◽  
Camv 35S ◽  
General Decline ◽  
Transgenic Lines

In this research, Arabidopsis thaliana plants were transformed with hairpin constructs targeting cyst nematode (Heterodera schachtii) genes, driven by the cauliflower mosaic virus (CaMV) 35S promoter: two housekeeping genes (the splicing factor Hs-U2AF and the vacuolar Hs-H+ATPase) and one candidate effector gene (the ubiquitin extension protein Hs-ubi). Expression of the dsRNA appeared to be extremely variable between and within homozygous T3 lines and even between tissues. Infection experiments showed up to 50% reduction in nematode infection for some transgenic lines. The results varied not only between lines containing the same construct but also between independent repetitions of the experiment. Further focusing on the Hs-U2AF-RNAi lines revealed large variations and a general decline of construct expression levels over the generations. Bisulphite sequencing of a 197 bp part of the CaMV 35S promoter revealed substantial methylation in this region and a negative correlation between the methylation level and expression of the hairpin construct. Taken together, our results show that host-generated RNAi can suffer from high levels of transcriptional silencing of the construct, leading to varying expression levels within and between transgenic lines.

scholarly journals T4 Lysozyme and Attacin Genes Enhance Resistance of Transgenic `Galaxy' Apple against Erwinia amylovora

2002 ◽  
Vol 127 (4) ◽  
pp. 515-519 ◽  
Author(s):  
Kisung Ko ◽  
John L. Norelli ◽  
Jean-Paul Reynoird ◽  
Herb S. Aldwinckle ◽  
Susan K. Brown
Keyword(s):  
Mosaic Virus ◽  
Erwinia Amylovora ◽  
Alfalfa Mosaic Virus ◽  
Cauliflower Mosaic Virus ◽  
Enzyme Linked Immunosorbent Assay ◽  
35S Promoter ◽  
In Planta ◽  
Mechanical Wounding ◽  
Genes Encoding ◽  
Transgenic Lines

Genes encoding lysozyme (T4L) from T4 bacteriophage and attacin E (attE) from Hyalophora cecropia were used, either singly or in combination, to construct plant binary vectors, pLDB15, p35SAMVT4, and pPin2Att35SAMVT4, respectively, for Agrobacterium-mediated transformation of `Galaxy' apple, to enhance resistance to Erwinia amylovora. In these plasmids, the T4L gene was controlled by the cauliflower mosaic virus 35S promoter with duplicated upstream domain and the untranslated leader sequence of alfalfa mosaic virus RNA 4, and the attE gene was controlled by the potato proteinase inhibitor II (Pin2) promoter. All transgenic lines were screened by polymerase chain reaction (PCR) for T4L and attE genes, and a double-antibody sandwich enzyme-linked immunosorbent assay for neomycin phosphotransferase II. Amplification of T4L and attE genes was observed in reverse transcriptase-PCR, indicating that these genes were transcribed in all tested transgenic lines containing each gene. The attacin protein was detected in all attE transgenic lines. The expression of attE under the Pin2 promoter was constitutive but higher levels of expression were observed after mechanical wounding. Some T4L or attE transgenic lines had significant disease reduction compared to nontransgenic `Galaxy'. However, transgenic lines containing both attE and T4L genes were not significantly more resistant than nontransgenic `Galaxy', indicating that there was no in planta synergy between attE and T4L with respect to resistance to E. amylovora.

scholarly journals Engineering Resistance to Multiple Prunus Fruit Viruses Through Expression of Chimeric Hairpins

2007 ◽  
Vol 132 (3) ◽  
pp. 407-414 ◽  
Author(s):  
Zongrang Liu ◽  
Ralph Scorza ◽  
Jean-Michel Hily ◽  
Simon W. Scott ◽  
Delano James
Keyword(s):  
Gene Silencing ◽  
Mosaic Virus ◽  
Fruit Production ◽  
Cauliflower Mosaic Virus ◽  
Ringspot Virus ◽  
35S Promoter ◽  
Plum Pox Virus ◽  
In Planta ◽  
Posttranscriptional Gene Silencing ◽  
Transgenic Lines

Prunus L. fruit production is seriously affected by several predominant viruses. The development of new cultivars resistant to these viruses is challenging but highly desired by breeders and growers. We report a posttranscriptional gene silencing-based approach for engineering multivirus resistance in plants. A single chimeric transgene, PTRAP6, was created by the fusion of 400 to 500-base pair (bp) gene fragments from six major Prunus fruit viruses, including american plum line pattern virus, peach mosaic virus, plum pox virus (PPV), prune dwarf virus (PDV), prunus necrotic ringspot virus, and tomato ringspot virus (ToRSV). Both strands of PTRAP6 were found being transcribed as an ≈2.5-kilobp transcript in planta without splicing interruption. To induce gene silencing/virus resistance, we placed two copies of PTRAP6 in an inverted repeat under the control of the cauliflower mosaic virus 35S promoter and separated by an intron spacer fragment to create PTRAP6i. Inoculation of the resulting transgenic Nicotiana benthamiana Domin. plants revealed that 12 of 28 R0 PTRAP6i transgenic lines (43%) were resistant to ToRSV ranging from mild symptoms to symptom-free phenotypes. Detailed analysis of two of three highly resistant homozygous R3 generation lines demonstrated that they were resistant to all three viruses tested, including PDV, PPV, and ToRSV. The remaining three viruses targeted by PTRAP6i were either unavailable for this study or were unable to systemically infect N. benthamiana. Transgene-wide and -specific small interfering RNA species were detected along with disappearance of transgene transcript in the resistant lines, indicating that posttranscriptional gene silencing underlies the mechanism of resistance. This work presents evidence that PTRAP6i is able to confer gene silencing-based resistance to multiple Prunus fruit viruses.

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