scholarly journals Identification of Chlamydomonas reinhardtii endogenous genic flanking sequences for improved transgene expression

2017 ◽  
Vol 92 (6) ◽  
pp. 1232-1244 ◽  
Author(s):  
Cristina López‐Paz ◽  
Dianyi Liu ◽  
Sa Geng ◽  
James G. Umen
Keyword(s):  
Chlamydomonas Reinhardtii ◽  
Transgene Expression ◽  
Flanking Sequences

A novel T-DNA vector design for selection of transgenic lines with simple transgene integration and stable transgene expression

2005 ◽  
Vol 32 (8) ◽  
pp. 671 ◽  
Author(s):  
Song Chen ◽  
Christopher A. Helliwell ◽  
Li-Min Wu ◽  
Elizabeth S. Dennis ◽  
Narayana M. Upadhyaya ◽  
...  
Keyword(s):  
Transgene Expression ◽  
Marker Gene ◽  
Direct Repeat ◽  
Transgene Silencing ◽  
Selective Marker ◽  
Hairpin Rna ◽  
Transgenic Lines ◽  
Flanking Sequences ◽  
Dna Vector ◽  
Selection Of

Plants transformed with Agrobacterium frequently contain T-DNA concatamers with direct-repeat (d / r) or inverted-repeat (i / r) transgene integrations, and these repetitive T-DNA insertions are often associated with transgene silencing. To facilitate the selection of transgenic lines with simple T-DNA insertions, we constructed a binary vector (pSIV) based on the principle of hairpin RNA (hpRNA)-induced gene silencing. The vector is designed so that any transformed cells that contain more than one insertion per locus should generate hpRNA against the selective marker gene, leading to its silencing. These cells should, therefore, be sensitive to the selective agent and less likely to regenerate. Results from Arabidopsis and tobacco transformation showed that pSIV gave considerably fewer transgenic lines with repetitive insertions than did a conventional T-DNA vector (pCON). Furthermore, the transgene was more stably expressed in the pSIV plants than in the pCON plants. Rescue of plant DNA flanking sequences from pSIV plants was significantly more frequent than from pCON plants, suggesting that pSIV is potentially useful for T-DNA tagging. Our results revealed a perfect correlation between the presence of tail-to-tail inverted repeats and transgene silencing, supporting the view that read-through hpRNA transcript derived from i / r T-DNA insertions is a primary inducer of transgene silencing in plants.

Chemical regulation of Fea1 driven transgene expression in Chlamydomonas reinhardtii

2017 ◽  
Vol 26 ◽  
pp. 323-329 ◽  
Author(s):  
Paula Barjona do Nascimento Coutinho ◽  
Christine Friedl ◽  
Rainer Buchholz ◽  
Stephanie Christine Stute
Keyword(s):  
Chlamydomonas Reinhardtii ◽  
Transgene Expression ◽  
Chemical Regulation

scholarly journals Human and pigSRY 5′ flanking sequences can direct reporter transgene expression to the genital ridge and to migrating neural crest cells

2006 ◽  
Vol 235 (3) ◽  
pp. 623-632 ◽  
Author(s):  
Alexandre Boyer ◽  
Nicolas Pilon ◽  
Diana L. Raiwet ◽  
Jacques G. Lussier ◽  
David W. Silversides
Keyword(s):  
Neural Crest ◽  
Transgene Expression ◽  
Neural Crest Cells ◽  
Genital Ridge ◽  
Flanking Sequences ◽  
Reporter Transgene

Hypoxia stimulates human preproendothelin-1 promoter activity in transgenic mice

1997 ◽  
Vol 273 (4) ◽  
pp. L848-L855 ◽  
Author(s):  
Catherine R. Aversa ◽  
Suzanne Oparil ◽  
Jaime Caro ◽  
Huaibin Li ◽  
Shuang-Dan Sun ◽  
...  
Keyword(s):  
Transgenic Mice ◽  
Transgene Expression ◽  
Regulatory Mechanisms ◽  
Pulmonary Vasculature ◽  
Pulmonary Tissue ◽  
Hypoxic Induction ◽  
Flanking Sequences ◽  
Bronchiolar Epithelium

Significant elevations in endothelin (ET)-1 levels accompany many diseases, but the underlying regulatory mechanisms are unclear. To investigate the in vivo regulation of human preproendothelin-1 (PPET-1), we examined the activity of the PPET-1 promoter in transgenic mice exposed to hypoxia. Mice expressing one of three PPET-1 promoter-luciferase (PPET-1/LUC) reporter transgenes (≈2.5 kb, 138 bp, or none of the 5′-flanking sequences of the PPET-1 gene) were generated. LUC expression was reduced in mice with a truncated 138-bp PPET-1 promoter. Exposure of mice bearing the 2.5-kb PPET-1/LUC transgene to hypoxia (10% O2for 24 h) increased LUC expression sixfold in pulmonary tissue but only twofold in other tissues. In situ hybridization revealed the strongest transgene expression in the pulmonary vasculature and bronchiolar epithelium. These data are consistent with the hypothesis that hypoxic induction of the PPET-1 gene leads to increased pulmonary production of ET-1 in diseases associated with low O2tension.

Strategies to facilitate transgene expression in Chlamydomonas reinhardtii

Planta ◽  
2009 ◽  
Vol 229 (4) ◽  
pp. 873-883 ◽  
Author(s):  
Alke Eichler-Stahlberg ◽  
Wolfram Weisheit ◽  
Ovidiu Ruecker ◽  
Markus Heitzer
Keyword(s):  
Chlamydomonas Reinhardtii ◽  
Transgene Expression

Validated Nuclear-Based Transgene Expression Regulated by the Fea1 Iron-Responsive Promoter in the Green Alga Chlamydomonas reinhardtii

2019 ◽  
Vol 61 (5) ◽  
pp. 305-316 ◽  
Author(s):  
Paula Barjona do Nascimento Coutinho ◽  
Christine Friedl ◽  
Marcus Heilmann ◽  
Rainer Buchholz ◽  
Stephanie Christine Stute
Keyword(s):  
Chlamydomonas Reinhardtii ◽  
Green Alga ◽  
Transgene Expression

1,25-dihydroxyvitamin D3 increases osteoblast-specific transgene expression regulated by human osteocalcin gene flanking sequences

Bone ◽  
1996 ◽  
Vol 19 (3) ◽  
pp. 162
Author(s):  
N.A. Sims ◽  
E.M. Gardiner ◽  
C.P. White ◽  
M.L. Drummond ◽  
N.A. Morrison ◽  
...  
Keyword(s):  
Transgene Expression ◽  
Dihydroxyvitamin D3 ◽  
1,25 Dihydroxyvitamin D3 ◽  
Osteocalcin Gene ◽  
Flanking Sequences ◽  
Human Osteocalcin

Evaluating nuclear transgene expression systems in Chlamydomonas reinhardtii

2013 ◽  
Vol 2 (4) ◽  
pp. 321-332 ◽  
Author(s):  
Anil Kumar ◽  
Vanessa R. Falcao ◽  
Richard T. Sayre
Keyword(s):  
Chlamydomonas Reinhardtii ◽  
Transgene Expression ◽  
Expression Systems

scholarly journals Exploring the Impact of Terminators on Transgene Expression in Chlamydomonas reinhardtii with a Synthetic Biology Approach

Life ◽  
2021 ◽  
Vol 11 (9) ◽  
pp. 964
Author(s):  
Katrin Geisler ◽  
Mark A. Scaife ◽  
Paweł M. Mordaka ◽  
Andre Holzer ◽  
Eleanor V. Tomsett ◽  
...  
Keyword(s):  
Synthetic Biology ◽  
Chlamydomonas Reinhardtii ◽  
Transgene Expression ◽  
Model Organism ◽  
Optimal Size ◽  
Gfp Reporter Gene ◽  
The Many ◽  
High Level ◽  
The Impact ◽  
Synthetic Biology Approach

Chlamydomonas reinhardtii has many attractive features for use as a model organism for both fundamental studies and as a biotechnological platform. Nonetheless, despite the many molecular tools and resources that have been developed, there are challenges for its successful engineering, in particular to obtain reproducible and high levels of transgene expression. Here we describe a synthetic biology approach to screen several hundred independent transformants using standardised parts to explore different parameters that might affect transgene expression. We focused on terminators and, using a standardised workflow and quantitative outputs, tested 9 different elements representing three different size classes of native terminators to determine their ability to support high level expression of a GFP reporter gene. We found that the optimal size reflected the median size of element found in the C. reinhardtii genome. The behaviour of the terminator parts was similar with different promoters, in different host strains and with different transgenes. This approach is applicable to the systematic testing of other genetic elements, facilitating comparison to determine optimal transgene design.

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