A suite of novel promoters and terminators for plant biotechnology. II. The pPLEX series for use in monocots

2003 ◽  
Vol 30 (4) ◽  
pp. 453 ◽  
Author(s):  
Petra H.D. Schünmann ◽  
Brian Surin ◽  
Peter M. Waterhouse
Keyword(s):  
Transgene Expression ◽  
Expression Patterns ◽  
Plant Biotechnology ◽  
Leaf Tissue ◽  
Subterranean Clover ◽  
Expression Vectors ◽  
Reading Frame ◽  
Subterranean Clover Stunt Virus ◽  
Maize Ubiquitin Promoter ◽  
Low Levels

A suite of plant expression vectors (pPLEX), constructed from the gene regulation signals from subterranean clover stunt virus (SCSV) genome, has previously been used in dicot transformation for a variety of applications in plant biotechnology. To assess their use for the transformation of monocots, a number of modifications were made to the basic vector series and assessed in rice. In their unmodified forms, the SCSV promoters directed low levels of gene expression, however, insertion of an intron between the promoter and the transgene open reading frame (analogous to the rice actin and maize ubiquitin promoter systems) increased transgene expression 50-fold. The expression patterns from the intron-modified SCSV (segments 4 and 7) promoters were very similar to those directed by the actin or ubiquitin promoters. All promoter systems investigated directed expression that appeared to be constitutive within leaf tissue, and localised to the epidermal and vascular tissues of the root. The pPLEX vectors described here are an important counterpart to the dicot pPLEX series and have the potential to be useful in monocot research and biotechnology.

A suite of novel promoters and terminators for plant biotechnology

2003 ◽  
Vol 30 (4) ◽  
pp. 443 ◽  
Author(s):  
Petra H. D. Schünmann ◽  
Danny J. Llewellyn ◽  
Brian Surin ◽  
Petra Boevink ◽  
Robert C. De Feyter ◽  
...  
Keyword(s):  
Solanum Tuberosum L ◽  
Marker Gene ◽  
Plant Biotechnology ◽  
Subterranean Clover ◽  
Expression Vectors ◽  
Crop Species ◽  
Specific Expression ◽  
Circular Dna ◽  
Subterranean Clover Stunt Virus ◽  
Gene Assay

The gene regulation signals from subterranean clover stunt virus (SCSV) were investigated for their expression in dicot plants. The SCSV genome has at least eight circular DNA molecules. Each circular DNA component contains a promoter element, a single open reading frame and a terminator. The promoters from seven of the segments were examined for their strength and tissue specificity in transgenic tobacco (Nicotiana tabacum L.), potato (Solanum tuberosum L.) and cotton (Gossypium hirsutum L.) using a GUS reporter gene assay system. While the promoters of many of the segments were poorly expressed, promoters derived from segments 4 and 7 were shown to direct high levels of expression in various plant tissues and organs. The segment 1 promoter directs predominantly callus-specific expression and, when used to control a selectable marker gene, facilitated the transformation of all three species (tobacco, potato and cotton). From the results, a suite of plant expression vectors (pPLEX) derived from the SCSV genome were constructed and used here to produce herbicide- and insect-resistant cotton, demonstrating their utility in the expression of foreign genes in dicot crop species and their potential for use in agricultural biotechnology.

scholarly journals Functional examination of lncRNAs in allotetraploid Gossypium hirsutum

BMC Genomics ◽  
2021 ◽  
Vol 22 (1) ◽  
Author(s):  
Luyao Wang ◽  
Jin Han ◽  
Kening Lu ◽  
Menglin Li ◽  
Mengtao Gao ◽  
...  
Keyword(s):  
Abiotic Stresses ◽  
Expression Patterns ◽  
Leaf Tissue ◽  
Evolutionary Model ◽  
Cotton Leaf ◽  
Functional Screening ◽  
Virus Induced Gene Silencing ◽  
Individual Gene ◽  
Cotton Species ◽  
Allotetraploid Cotton

Abstract Background An evolutionary model using diploid and allotetraploid cotton species identified 80 % of non-coding transcripts in allotetraploid cotton as being uniquely activated in comparison with its diploid ancestors. The function of the lncRNAs activated in allotetraploid cotton remain largely unknown. Results We employed transcriptome analysis to examine the relationship between the lncRNAs and mRNAs of protein coding genes (PCGs) in cotton leaf tissue under abiotic stresses. LncRNA expression was preferentially associated with that of the flanking PCGs. Selected highly-expressed lncRNA candidates (n = 111) were subjected to a functional screening pilot test in which virus-induced gene silencing was integrated with abiotic stress treatment. From this low-throughput screen, we obtained candidate lncRNAs relating to plant height and tolerance to drought and other abiotic stresses. Conclusions Low-throughput screen is an effective method to find functional lncRNA for further study. LncRNAs were more active in abiotic stresses than PCG expression, especially temperature stress. LncRNA XLOC107738 may take a cis-regulatory role in response to environmental stimuli. The degree to which lncRNAs are constitutively expressed may impact expression patterns and functions on the individual gene level rather than in genome-wide aggregate.

scholarly journals Cytoplasmic Male Sterility-Associated Mitochondrial Gene orf312 Derived from Rice (Oryza sativa L.) Cultivar Tadukan

Rice ◽  
2021 ◽  
Vol 14 (1) ◽  
Author(s):  
Ayumu Takatsuka ◽  
Tomohiko Kazama ◽  
Kinya Toriyama
Keyword(s):  
Oryza Sativa ◽  
Cytoplasmic Male Sterility ◽  
Male Sterility ◽  
Oryza Sativa L ◽  
Mitochondrial Gene ◽  
Expression Patterns ◽  
Promising Candidate ◽  
Illumina Hiseq ◽  
Reading Frame ◽  
Cms Line

Abstract Background Cytoplasmic male sterility (CMS) is a trait associated with non-functional pollen or anthers, caused by the interaction between mitochondrial and nuclear genes. Findings A Tadukan-type CMS line (TAA) and a restorer line (TAR) were obtained by successive backcrossing between the Oryza sativa cultivars Tadukan (a cytoplasmic donor) and Taichung 65 (a recurrent pollen parent). Using Illumina HiSeq, we determined whole-genome sequences of the mitochondria of TAA and screened the mitochondrial genome for the presence of open reading frame (orf) genes specific to this genome. One of these orf genes, orf312, showed differential expression patterns in TAA and TAR anthers at the meiotic and mature stages, with transcript amounts in TAR being less than those in TAA. The orf312 gene is similar to the previously described orf288, a part of which is among the components comprising WA352, a chimeric CMS-associated gene of wild-abortive-type CMS. Conclusions The orf312 gene is a promising candidate for CMS-associated gene in TAA.

Phosphorus compounds measured in a rapid and simple leaf test for the assessment of the phosphorus status of subterranean clover

1984 ◽  
Vol 24 (125) ◽  
pp. 213 ◽  
Author(s):  
GCJ Irving ◽  
D Bouma
Keyword(s):  
Leaf Tissue ◽  
Rapid Test ◽  
Subterranean Clover ◽  
Exchange Chromatography ◽  
Phosphorus Compounds ◽  
Fresh Leaf ◽  
Organic Phosphates ◽  
Phosphorus Status ◽  
Colour Development ◽  
Hydrolysis Of

Experiments were done to determine what proportion of the phosphate extracted from fresh leaf tissue by five drops of 10 N H2SO4 represents inorganic tissue phosphate, and to what extent hydrolysis of organic phosphates during and after the extraction, and during the development of the blue phosphomolybdate complex, could contribute to the values obtained. The extraction is the basis of a simple and rapid test for the assessment of the phosphorus status of subterranean clover (Bouma and Dowling 1982). Extraction of leaf tissue of subterranean clover and sunflower with 0.2 M HClO4 at O�C, which was shown to extract inorganic leaf phosphorus without causing significant hydrolysis of organic phosphates, gave values not significantly different from those in H2SO4 extracts. The rate of hydrolysis of endogenous organic phosphates in tissue, extracted and left at room temperature for periods of up to 40 min. after adding H2SO4, did not differ significantly from zero. Errors due to hydrolysis during the 30 min. previously recommended for colour development are reduced to negligible proportions by reducing the time for colour development to 10 min. and by adding citric acid at this point. Anion-exchange chromatography of 10 N H2SO4 and 0.2 M HClO4 extracts confirmed the similarity of their composition and provided estimates of the various phosphate compounds present. The extraction of fresh leaf tissue with 10 N H2SO4 provides a satisfactory estimate of the endogenous inorganic phosphorus content.

IDENTIFICATION, SEQUENCE AND EXPRESSION OF A CRUSTACEAN CARDIOACTIVE PEPTIDE (CCAP) GENE IN THE MOTHMANDUCA SEXTA

2001 ◽  
Vol 204 (16) ◽  
pp. 2803-2816 ◽  
Author(s):  
P. K. LOI ◽  
S. A. EMMAL ◽  
Y. PARK ◽  
N. J. TUBLITZ
Keyword(s):  
Central Nervous System ◽  
Nervous System ◽  
Gene Structure ◽  
Expression Patterns ◽  
Amino Acid Residues ◽  
Coding Region ◽  
Genome Database ◽  
Reading Frame ◽  
Crustacean Cardioactive Peptide ◽  
Embryonic Life

SUMMARYThe crustacean cardioactive peptide (CCAP) gene was isolated from the tobacco hawkmoth Manduca sexta. The gene has an open reading frame of 125 amino acid residues containing a single, complete copy of CCAP. Analysis of the gene structure revealed three introns interrupting the coding region. A comparison of the M. sexta CCAP gene with the Drosophila melanogaster genome database reveals significant similarities in sequence and gene structure.The spatial and temporal expression patterns of the CCAP gene in the M. sexta central nervous system were determined in all major post-embryonic stages using in situ hybridization techniques. The CCAP gene is expressed in a total of 116 neurons in the post-embryonic M. sextacentral nervous system. Nine pairs of cells are observed in the brain, 4.5 pairs in the subesophageal ganglion, three pairs in each thoracic ganglion(T1-T3), three pairs in the first abdominal ganglion (A1), five pairs each in the second to sixth abdominal ganglia (A2-A6) and 7.5 pairs in the terminal ganglion. The CCAP gene is expressed in every ganglion in each post-embryonic stage, except in the thoracic ganglia of first- and second-instar larvae. The number of cells expressing the CCAP gene varies during post-embryonic life,starting at 52 cells in the first instar and reaching a maximum of 116 shortly after pupation. One set of thoracic neurons expressing CCAP mRNA shows unusual variability in expression levels immediately prior to larval ecdysis. Using previously published CCAP immunocytochemical data, it was determined that 91 of 95 CCAP-immunopositive neurons in the M. sexta central nervous system also express the M. sexta CCAP gene, indicating that there is likely to be only a single CCAP gene in M. sexta.

Cartilage associated protein (CASP) is a novel developmentally regulated chick embryo protein

1997 ◽  
Vol 110 (12) ◽  
pp. 1351-1359
Author(s):  
P. Castagnola ◽  
M. Gennari ◽  
R. Morello ◽  
L. Tonachini ◽  
O. Marin ◽  
...  
Keyword(s):  
Chick Embryo ◽  
Developmentally Regulated ◽  
Reading Frame ◽  
Amino Terminal ◽  
Nuclear Antigens ◽  
Cartilage Extracellular Matrix ◽  
Low Levels ◽  
Specific Rabbit ◽  
Chicken Protein

A subtracted cDNA library was generated to identify cDNAs specific for chondrocyte mRNAs preferentially expressed at the hypertrophic stage with respect to early differentiation stages. The characterization of a cDNA isolated from this library that hybridizes with a 1.8 kb mRNA is described here. This mRNA is expressed at extremely low levels in dedifferentiated chondrocytes cultured in adherent conditions, at very low levels in differentiating chondrocytes and at very high levels in hypertrophic chondrocytes cultured in suspension conditions. In the developing chick embryo this mRNA is detectable in RNAs extracted from several other tissues besides cartilage. The described cDNA contains a complete open reading frame coding for a polypeptide of about 33 kDa. Homology searches with known cDNA and protein sequences have revealed that the chicken protein is related to the amino-terminal half of two mammalian nuclear antigens. By immunohistochemistry with specific rabbit antisera a strong signal was detected in the cartilage extracellular matrix of selected regions of the developing skeleton. Because of this localization of the antigen we named this protein cartilage associated protein (hereafter referred to as CASP).

scholarly journals An Alternative Platform for Protein Expression Using an Innate Whole Expression Module from Metagenomic DNA

2019 ◽  
Vol 7 (1) ◽  
pp. 9 ◽  
Author(s):  
Dae-Eun Cheong ◽  
So-Youn Park ◽  
Ho-Dong Lim ◽  
Geun-Joong Kim
Keyword(s):  
Gene Expression ◽  
Regulatory Element ◽  
Expression System ◽  
Expression Patterns ◽  
Gene Clusters ◽  
Single Element ◽  
Reading Frame ◽  
Cis Acting ◽  
Dna Libraries ◽  
Expression Module

Many integrated gene clusters beyond a single genetic element are commonly trapped as the result of promoter traps in (meta)genomic DNA libraries. Generally, a single element, which is mainly the promoter, is deduced from the resulting gene clusters and employed to construct a new expression vector. However, expression patterns of target proteins under the incorporated promoter are often inconsistent with those shown in clones harboring plasmids with gene clusters. These results suggest that the integrated set of gene clusters with diverse cis- and trans-acting elements is evolutionarily tuned as a complete set for gene expression, and is an expression module with all the components for the expression of a nested open reading frame (ORF). This possibility is further supported by truncation and/or serial deletion analysis of this module in which the expression of the nested ORF is highly fluctuated or reduced frequently, despite being supported by plentiful cis-acting elements in the spanning regions around the ORF such as the promoter, ribosome binding site (RBS), terminator, and 3′-/5′-UTRs for gene expression. Here, we examined whether an innate module with a naturally overexpressed gene could be considered as a scaffold for an expression system. For a proof-of-principle study, we mined a complete expression module with an innately overexpressed ORF in E. coli from a metagenomics DNA library, and incorporated it into a vector that had no regulatory element for expressing the insert. We obtained successful expression of several inserts such as MBP, GFPuv, β-glucosidase, and esterase using this simple construct without tuning and codon optimization of the target insert.

Heritable transgene expression pattern imposed onto maize ubiquitin promoter by maize adh-1 matrix attachment regions: tissue and developmental specificity in maize transgenic plants

2004 ◽  
Vol 22 (12) ◽  
Author(s):  
Fran�ois Torney ◽  
Anne Partier ◽  
V�ronique Says-Lesage ◽  
Isabelle Nadaud ◽  
Pierre Barret ◽  
...  
Keyword(s):  
Transgenic Plants ◽  
Expression Pattern ◽  
Transgene Expression ◽  
Matrix Attachment Regions ◽  
Ubiquitin Promoter ◽  
Maize Ubiquitin Promoter

Characterization of a gene locus from Erwinia amylovora with regulatory functions in exopolysaccharide synthesis of Erwinia spp.

1998 ◽  
Vol 44 (7) ◽  
pp. 657-666 ◽  
Author(s):  
Phillip Aldridge ◽  
Frank Bernhard ◽  
Peter Bugert ◽  
David L Coplin ◽  
Klaus Geider
Keyword(s):  
Structural Gene ◽  
Erwinia Amylovora ◽  
Genomic Library ◽  
Expression Vectors ◽  
External Signal ◽  
Wild Type ◽  
Reading Frame ◽  
Gus Reporter ◽  
Acid Protein ◽  
Erwinia Stewartii

In a genomic library of Erwinia amylovora, a locus has been identified that can suppress an Erwinia stewartii rcsA mutant. In addition, the locus induced a mucoid sticky phenotype of colonies in a wild-type strain of Erwinia stewartii and increased exopolysaccharide synthesis in several species of bacteria belonging to the genus Erwinia. An open reading frame was identified at this locus encoding a 225 amino acid protein that contained a helix-turn-helix motif typical of transcriptional regulators. The corresponding gene was subsequently named rcsV (regulator of capsular synthesis affecting viscosity). A mutant of rcsV in wild-type Erwinia amylovora had no detectable phenotype and produced typical levels of amylovoran under laboratory conditions. The rcsV gene on a high copy number plasmid under the control of its own promoter did not alter amylovoran production, in contrast to in-frame fusions of the structural gene in expression vectors. Since even the lac promoter was inert in the expression of rcsV, a DNA-binding protein could inhibit transcription of the gene in Erwinia amylovora. On the other hand, an Erwinia amylovora rcsA mutant was suppressed by rcsV when its promoter was replaced and the structural gene fused in-frame with lacZ' or malE. Northern blots, with total RNA from Erwinia amylovora, or promoter analysis using the GUS reporter gene did not show expression of rcsV in Erwinia amylovora, although primer extension analysis did. RcsV could be a component involved in the regulation of amylovoran synthesis, and gene expression may require an unknown external signal during the life cycle or pathogenesis of Erwinia amylovora. Key words: amylovoran, fire blight, rcsA-like activator, fusion protein.

Increased Duration of Transgene Expression in the Lung with Plasmid DNA Vectors Harboring Adenovirus E4 Open Reading Frame 3

1999 ◽  
Vol 10 (11) ◽  
pp. 1833-1843 ◽  
Author(s):  
Nelson S. Yew ◽  
John Marshall ◽  
Malgorzata Przybylska ◽  
Donna M. Wysokenski ◽  
Robin J. Ziegler ◽  
...  
Keyword(s):  
Plasmid Dna ◽  
Transgene Expression ◽  
Open Reading Frame ◽  
Reading Frame ◽  
Dna Vectors
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