Development of a nuclear transformation system with a codon-optimized selection marker and reporter genes in Pyropia yezoensis (Rhodophyta)

2014 ◽  
Vol 26 (4) ◽  
pp. 1863-1868 ◽  
Author(s):  
Ryo Hirata ◽  
Toshiki Uji ◽  
Satoru Fukuda ◽  
Hiroyuki Mizuta ◽  
Asao Fujiyama ◽  
...  
Keyword(s):  
Reporter Genes ◽  
Pyropia Yezoensis ◽  
Selection Marker ◽  
Nuclear Transformation ◽  
Transformation System

scholarly journals A Codon-Optimized Bacterial Antibiotic Gene Used as Selection Marker for Stable Nuclear Transformation in the Marine Red Alga Pyropia yezoensis

2013 ◽  
Vol 16 (3) ◽  
pp. 251-255 ◽  
Author(s):  
Toshiki Uji ◽  
Ryo Hirata ◽  
Satoru Fukuda ◽  
Hiroyuki Mizuta ◽  
Naotsune Saga
Keyword(s):  
Red Alga ◽  
Pyropia Yezoensis ◽  
Selection Marker ◽  
Nuclear Transformation

scholarly journals Transformation of coffee (Coffea Arabica L. cv. Catimor) with the cry1ac gene by biolistic, without the use of markers

2010 ◽  
Vol 70 (2) ◽  
pp. 387-393 ◽  
Author(s):  
Z De Guglielmo-Cróquer ◽  
I. Altosaar ◽  
M. Zaidi ◽  
A. Menéndez-Yuffá
Keyword(s):  
Reporter Genes ◽  
Leaf Miner ◽  
Selection Marker ◽  
Marker Genes ◽  
Cry1ac Gene ◽  
Reverse Transcription Pcr ◽  
Gus Reporter ◽  
Leucoptera Coffeella ◽  
Coffee Plants ◽  
Coffea Arabica L

The transformation of coffee plantlets with the cry1ac gene of Bacillus thuringiensis was achieved by biolistic using either the whole pUBC plasmid or only the ubi-cry1ac-nos genetic cassette. The cry1ac gene was inserted into coffee plants in order to confer resistance to the leaf miner Leucoptera coffeella, an insect responsible for considerable losses in coffee crops. Bearing in mind that the genetic cassettes used for this study lack reporter genes and/or selection marker genes, the parameters for the transformation procedure by biolistic were previously standardised with a plasmid carrying the gus reporter gene. The presence of the cry1ac gene in young plantlet tissues was determined by PCR, Southern blot and reverse transcription-PCR. Our results show that the obtainment of viable coffee plantlets, transformed by bombardment with the cry1ac gene and without selection markers nor reporter genes, is feasible.

scholarly journals Enhancement of Ganoderic Acid Accumulation by Overexpression of an N-Terminally Truncated 3-Hydroxy-3-Methylglutaryl Coenzyme A Reductase Gene in the Basidiomycete Ganoderma lucidum

2012 ◽  
Vol 78 (22) ◽  
pp. 7968-7976 ◽  
Author(s):  
Jun-Wei Xu ◽  
Yi-Ning Xu ◽  
Jian-Jiang Zhong
Keyword(s):  
Ganoderma Lucidum ◽  
Coenzyme A ◽  
Catalytic Domain ◽  
Selection Marker ◽  
Transformation System ◽  
Protein Subunit ◽  
Ganoderic Acid ◽  
Content Type ◽  
Ganoderic Acids ◽  
Coenzyme A Reductase

ABSTRACTGanoderic acids produced byGanoderma lucidum, a well-known traditional Chinese medicinal mushroom, exhibit antitumor and antimetastasis activities. Genetic modification ofG. lucidumis difficult but critical for the enhancement of cellular accumulation of ganoderic acids. In this study, a homologous genetic transformation system forG. lucidumwas developed for the first time using mutatedsdhB, encoding the iron-sulfur protein subunit of succinate dehydrogenase, as a selection marker. The truncatedG. lucidumgene encoding the catalytic domain of 3-hydroxy-3-methylglutaryl coenzyme A reductase (HMGR) was overexpressed by using theAgrobacterium tumefaciens-mediated transformation system. The results showed that the mutatedsdhBsuccessfully conferred carboxin resistance upon transformation. Most of the integrated transfer DNA (T-DNA) appeared as a single copy in the genome. Moreover, deregulated constitutive overexpression of the HMGR gene led to a 2-fold increase in ganoderic acid content. It also increased the accumulation of intermediates (squalene and lanosterol) and the upregulation of downstream genes such as those of farnesyl pyrophosphate synthase, squalene synthase, and lanosterol synthase. This study demonstrates that transgenic basidiomyceteG. lucidumis a promising system to achieve metabolic engineering of the ganoderic acid pathway.

Improvement of Chloroplast Transformation Using CRISPR/Cas9

2020 ◽  
Vol 14 (3) ◽  
pp. 401-407
Author(s):  
Ning Tang ◽  
Yumei Xia ◽  
Yijie Zhan ◽  
Junhao Dan ◽  
Mulan Yu ◽  
...  
Keyword(s):  
Dna Damage ◽  
Homologous Recombination ◽  
Gene Editing ◽  
Transformation Efficiency ◽  
Chloroplast Transformation ◽  
Nuclear Transformation ◽  
Important Research ◽  
Transformation System ◽  
Repair Mechanism ◽  
Damage Repair

Chloroplasts are organelles that contain genetic materials (DNA) in higher plant cells. The special genetic characteristics of chloroplasts mean that plasmid transformation has important research value, so it has become an important research direction second to nuclear transformation. Although the techniques of chloroplast genome modification have been successfully applied in tobacco and extended to other high plants, there are still many limitations. Exogenous genes are integrated into the chloroplast genome through homologous recombination. Therefore, the low efficiency of homologous recombination directly limits transformation efficiency. Gene editing with fixed-point cutting function and DNA damage repair mechanism may effectively improve the efficiency. In the present study, we aimed to use CRISPR/Cas9 to cut the site between two homologous recombinant fragments in chloroplast transformation to improve the efficiency by activating the DNA damage repair mechanism. The Cas9 gene and gRNA were added to the chloroplast transformation system of tobacco by co-transformation or integration into a transformation vector. The acquired resistant plants were screened by multiple selection of spectinomycin and chloroplast DNA was isolated for molecular detection by PCR. The results showed that the efficiency of chloroplast transformation increased by 6–10 times with the addition of gene editing technology. Although the transformation efficiency was still far below the level of nuclear transformation, this study may help to increase the efficiency of the plant chloroplast transformation system, and expand the types of plant receptors.

scholarly journals Stable Nuclear Transformation System for the Coccolithophorid Alga Pleurochrysis carterae

2016 ◽  
Vol 6 (1) ◽  
Author(s):  
Hirotoshi Endo ◽  
Megumi Yoshida ◽  
Toshiki Uji ◽  
Naotsune Saga ◽  
Koji Inoue ◽  
...  
Keyword(s):  
Nuclear Transformation ◽  
Transformation System ◽  
Pleurochrysis Carterae

Development of an integrative transformation system for the opportunistic pathogenic yeastCandida lusitaniae usingURA3 as a selection marker

Yeast ◽  
2004 ◽  
Vol 21 (2) ◽  
pp. 95-106 ◽  
Author(s):  
Fabienne François ◽  
Florence Chapeland-Leclerc ◽  
Jean Villard ◽  
Thierry Noël
Keyword(s):  
Selection Marker ◽  
Transformation System ◽  
Integrative Transformation ◽  
Opportunistic Pathogenic

scholarly journals Integrative Transformation System for the Metabolic Engineering of the Sphingoid Base-Producing Yeast Pichia ciferrii

2003 ◽  
Vol 69 (2) ◽  
pp. 812-819 ◽  
Author(s):  
Jung-Hoon Bae ◽  
Jung-Hoon Sohn ◽  
Chang-Seo Park ◽  
Joon-Shick Rhee ◽  
Eui-Sung Choi
Keyword(s):  
Metabolic Engineering ◽  
De Novo ◽  
Integration Site ◽  
Fold Increase ◽  
Selection Marker ◽  
Serine Palmitoyltransferase ◽  
Transformation System ◽  
Wild Type ◽  
Gene Encoding ◽  
Integrative Transformation

ABSTRACT We have developed an integrative transformation system for metabolic engineering of the tetraacetyl phytosphingosine (TAPS)-secreting yeast Pichia ciferrii. The system uses (i) a mutagenized ribosomal protein L41 gene of P. ciferrii as a dominant selection marker that confer resistance to the antibiotic cycloheximide and (ii) a ribosomal DNA (rDNA) fragment of P. ciferrii as a target for multicopy gene integration into the chromosome. A locus within the nontranscribed region located between 5S and 26S rDNAs was selected as the integration site. A maximum frequency of integrative transformation of approximately 1,350 transformants/μg of DNA was observed. To improve the de novo synthesis of sphingolipid, the LCB2 gene, encoding a subunit of serine palmitoyltransferase, which catalyzes the first committed step of sphingolipid synthesis, was cloned from P. ciferrii and overexpressed under the control of the P. ciferrii glyceraldehyde-3-phosphate dehydrogenase promoter. After transformation of an LCB2 gene expression cassette, several transformants that contained approximately five to seven copies of transforming DNA in the chromosome and exhibited about 50-fold increase in LCB2 mRNA relative to the wild type were identified. These transformants were observed to produce approximately two times more TAPS than the wild type.

scholarly journals Generation of Transgenic Rice without Antibiotic Selection Marker through Agrobacterium-mediated Co-transformation System

2012 ◽  
Vol 22 (9) ◽  
pp. 1152-1158
Author(s):  
Soo-Kwon Park ◽  
Tack-Min Kwon ◽  
Jong-Hee Lee ◽  
Dong-Jin Shin ◽  
Woon-Ha Hwang ◽  
...  
Keyword(s):  
Transgenic Rice ◽  
Selection Marker ◽  
Transformation System ◽  
Antibiotic Selection
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