A set of dual promoter vectors for high throughput cloning, screening, and protein expression in eukaryotic and prokaryotic systems from a single plasmid.


BACKGROUND:The ability to produce the same recombinant protein in both prokaryotic and eukaryotic cells offers many experimental opportunities. However, the cloning of the same gene into multiple plasmids is required, which is time consuming, laborious and still may not produce soluble, stable protein in sufficient quantities. We have developed a set of expression vectors that allows for ligation-independent cloning and rapid functional screening for protein expression in both E. coli and S. cerevisiae. RESULTS:A set of expression vectors was made that can express the same open reading frame in E. coli (via the T7 phage promoter) and in S. cerevisiae (via the CUP1 or MET25 promoter). These plasmids also contain the essential elements for replication and selection in both cell types and have several advantages: they allow for cloning of genes by homologous recombination in yeast, protein expression can be determined before plasmid isolation and sequencing, and a GST-fusion tag is added to aid in soluble expression and purification. We have also included a TEV recognition site that allows for the specific cleavage of the fusion proteins to yield native proteins. CONCLUSIONS:The dual promoter vectors can be used for rapid cloning, expression, and purification of target proteins from both prokaryotic and eukaryotic systems with the ability to study post-translation modifications.


BMC Biotechnol


BMC biotechnology


Sinah N,Williams CA,Piper RC,Shields SB




Has Abstract


2012-08-23 00:00:00










  • Identification and characterization of an Endo-glucanase secreted from cellulolytic Escherichia coli ZH-4.

    abstract:BACKGROUND:In the previous study, the cellulolytic Escherichia coli ZH-4 isolated from bovine rumen was found to show extracellular cellulase activity and could degrade cellulose in the culture. The goal of this work was to identify and characterize the secreted cellulase of E. coli ZH-4. It will be helpful to re-under...

    journal_title:BMC biotechnology

    pub_type: 杂志文章


    authors: Pang J,Wang J,Liu Z,Zhang Q,Qi Q

    更新日期:2019-08-27 00:00:00

  • Mimotopes selected with a neutralizing antibody against urease B from Helicobacter pylori induce enzyme inhibitory antibodies in mice upon vaccination.

    abstract:BACKGROUND:Urease B is an important virulence factor that is required for Helicobacter pylori to colonise the gastric mucosa. Mouse monoclonal antibodies (mAbs) that inhibit urease B enzymatic activity will be useful as vaccines for the prevention and treatment of H. pylori infection. Here, we produced murine mAbs agai...

    journal_title:BMC biotechnology

    pub_type: 杂志文章


    authors: Li Y,Ning Y,Wang Y,Peng D,Jiang Y,Zhang L,Long M,Luo J,Li M

    更新日期:2010-11-30 00:00:00

  • Enhanced in vitro osteogenic differentiation of human fetal MSCs attached to 3D microcarriers versus harvested from 2D monolayers.

    abstract:BACKGROUND:Mesenchymal stem cells (MSCs) are of great interest in bone regenerative medicine due to their osteogenic potential and trophic effects. However, challenges to large-scale production of MSCs can hinder the translation of MSC therapies. 3D Microcarrier (MC)-based MSC culture presents a scalable and cost-effec...

    journal_title:BMC biotechnology

    pub_type: 杂志文章


    authors: Shekaran A,Sim E,Tan KY,Chan JK,Choolani M,Reuveny S,Oh S

    更新日期:2015-10-31 00:00:00

  • Glycosyltransferase efficiently controls phenylpropanoid pathway.

    abstract:BACKGROUND:In a previous study, anthocyanin levels in potato plants were increased by manipulating genes connected with the flavonoid biosynthesis pathway. However, starch content and tuber yield were dramatically reduced in the transgenic plants, which over-expressed dihydroflavonol reductase (DFR). RESULTS:Transgeni...

    journal_title:BMC biotechnology

    pub_type: 杂志文章


    authors: Aksamit-Stachurska A,Korobczak-Sosna A,Kulma A,Szopa J

    更新日期:2008-03-05 00:00:00

  • Identification of residues important for the activity of aldehyde-deformylating oxygenase through investigation into the structure-activity relationship.

    abstract:BACKGROUND:Aldehyde-deformylating oxygenase (ADO) is a key enzyme involved in the biosynthetic pathway of fatty alk(a/e)nes in cyanobacteria. However, cADO (cyanobacterial ADO) showed extreme low activity with the k cat value below 1 min-1, which would limit its application in biofuel production. To identify the activi...

    journal_title:BMC biotechnology

    pub_type: 杂志文章


    authors: Wang Q,Bao L,Jia C,Li M,Li JJ,Lu X

    更新日期:2017-03-16 00:00:00

  • Developing a xylanase XYNZG from Plectosphaerella cucumerina for baking by heterologously expressed in Kluyveromyces lactis.

    abstract:BACKGROUND:Xylanase can replace chemical additives to improve the volume and sensory properties of bread in the baking. Suitable baking xylanase with improved yield will promote the application of xylanase in baking industry. The xylanase XYNZG from the Plectosphaerella cucumerina has been previously characterized by h...

    journal_title:BMC biotechnology

    pub_type: 杂志文章


    authors: Zhan FX,Wang QH,Jiang SJ,Zhou YL,Zhang GM,Ma YH

    更新日期:2014-12-16 00:00:00

  • Purification of functional baculovirus particles from silkworm larval hemolymph and their use as nanoparticles for the detection of human prorenin receptor (PRR) binding.

    abstract:BACKGROUND:Baculovirus, which has a width of 40 nm and a length of 250-300 nm, can display functional peptides, receptors and antigens on its surface by their fusion with a baculovirus envelop protein, GP64. In addition, some transmembrane proteins can be displayed without GP64 fusion, using the native transmembrane do...

    journal_title:BMC biotechnology

    pub_type: 杂志文章


    authors: Kato T,Suzuki F,Park EY

    更新日期:2011-06-02 00:00:00

  • Identification of an antibody fragment specific for androgen-dependent prostate cancer cells.

    abstract:BACKGROUND:Prostate cancer is the most-diagnosed non-skin cancer among males in the US, and the second leading cause of cancer-related death. Current methods of treatment and diagnosis are not specific for the disease. This work identified an antibody fragment that binds selectively to a molecule on the surface of andr...

    journal_title:BMC biotechnology

    pub_type: 杂志文章


    authors: Williams RM,Hajiran CJ,Nayeem S,Sooter LJ

    更新日期:2014-09-03 00:00:00

  • Effect of Fe nanoparticle on growth and glycolipid biosurfactant production under solid state culture by marine Nocardiopsis sp. MSA13A.

    abstract:BACKGROUND:Iron is an essential element in several pathways of microbial metabolism, and therefore low iron toxicity is expected on the usage of Fe nanoparticles (NPs). This study aims to determine the effect of Fe NPs on biosurfactant production by marine actinobacterium Nocardiopsis sp. MSA13A under solid state cultu...

    journal_title:BMC biotechnology

    pub_type: 杂志文章


    authors: Kiran GS,Nishanth LA,Priyadharshini S,Anitha K,Selvin J

    更新日期:2014-05-21 00:00:00

  • Metabolic evolution of Corynebacterium glutamicum for increased production of L-ornithine.

    abstract:BACKGROUND:L-ornithine is effective in the treatment of liver diseases and helps strengthen the heart. The commercial applications mean that efficient biotechnological production of L-ornithine has become increasingly necessary. Adaptive evolution strategies have been proven a feasible and efficient technique to achiev...

    journal_title:BMC biotechnology

    pub_type: 杂志文章


    authors: Jiang LY,Chen SG,Zhang YY,Liu JZ

    更新日期:2013-06-01 00:00:00

  • Efficient CPP-mediated Cre protein delivery to developing and adult CNS tissues.

    abstract:BACKGROUND:Understanding and manipulating gene function in physiological conditions is a major objective for both fundamental and applied research. In contrast to other experimental settings, which use either purely genetic or gene delivery (viral or non-viral) strategies, we report here a strategy based on direct prot...

    journal_title:BMC biotechnology

    pub_type: 杂志文章


    authors: Gitton Y,Tibaldi L,Dupont E,Levi G,Joliot A

    更新日期:2009-04-24 00:00:00

  • Efficient generation of Rosa26 knock-in mice using CRISPR/Cas9 in C57BL/6 zygotes.

    abstract:BACKGROUND:The CRISPR/Cas9 system is increasingly used for gene inactivation in mouse zygotes, but homology-directed mutagenesis and use of inbred embryos are less established. In particular, Rosa26 knock-in alleles for the insertion of transgenes in a genomic 'safe harbor' site, have not been produced. Here we applied...

    journal_title:BMC biotechnology

    pub_type: 杂志文章


    authors: Chu VT,Weber T,Graf R,Sommermann T,Petsch K,Sack U,Volchkov P,Rajewsky K,Kühn R

    更新日期:2016-01-16 00:00:00

  • A novel MVA-mediated pathway for isoprene production in engineered E. coli.

    abstract:BACKGROUND:To deal with the increasingly severe energy crisis and environmental consequences, biofuels and biochemicals generated from renewable resources could serve as a promising alternative for replacing petroleum as a source of fuel and chemicals, among which isoprene (2-methyl-1,3-butadiene) in particular is of g...

    journal_title:BMC biotechnology

    pub_type: 杂志文章


    authors: Yang J,Nie Q,Liu H,Xian M,Liu H

    更新日期:2016-01-20 00:00:00

  • Resveratrol alleviates lipopolysaccharide-induced inflammation in PC-12 cells and in rat model.

    abstract:BACKGROUND:Spinal cord injury (SCI) remains a huge medical problem nowadays as there is no hospital providing the versatile strategies for repairing central nervous system and restoring function. Herein, we focused on PC-12 cells as an important research tool and studied the potential role of resveratrol (RSV) in infla...

    journal_title:BMC biotechnology

    pub_type: 杂志文章


    authors: Zhang G,Liu Y,Xu L,Sha C,Zhang H,Xu W

    更新日期:2019-01-28 00:00:00

  • Multimodal in vivo imaging reveals limited allograft survival, intrapulmonary cell trapping and minimal evidence for ischemia-directed BMSC homing.

    abstract:BACKGROUND:Despite positive reports on the efficacy of stem cell therapy for the treatment of cardiovascular disease, the nature of stem cell homing to ischemic tissues remains elusive. RESULTS:We used a mouse model of peripheral tissue ischemia to study the survival and homing capacity of dual reporter gene (eGFP/Luc...

    journal_title:BMC biotechnology

    pub_type: 杂志文章


    authors: Everaert BR,Bergwerf I,De Vocht N,Ponsaerts P,Van Der Linden A,Timmermans JP,Vrints CJ

    更新日期:2012-12-03 00:00:00

  • Nicotiana benthamiana is a suitable transient system for high-level expression of an active inhibitor of cotton boll weevil α-amylase.

    abstract:BACKGROUND:Insect resistance in crops represents a main challenge for agriculture. Transgenic approaches based on proteins displaying insect resistance properties are widely used as efficient breeding strategies. To extend the spectrum of targeted pathogens and overtake the development of resistance, molecular evolutio...

    journal_title:BMC biotechnology

    pub_type: 杂志文章


    authors: Prado GS,Bamogo PKA,de Abreu JAC,Gillet FX,Dos Santos VO,Silva MCM,Brizard JP,Bemquerer MP,Bangratz M,Brugidou C,Sérémé D,Grossi-de-Sa MF,Lacombe S

    更新日期:2019-03-08 00:00:00

  • Novel anti-CD38 humanized mAb SG003 possessed enhanced cytotoxicity in lymphoma than Daratumumab via antibody-dependent cell-mediated cytotoxicity.

    abstract:BACKGROUND:In vivo use of monoclonal antibodies has become routine clinical practice in the treatment of human cancer. CD38 is an attractive target, because it has double roles, as a receptor and an ectoenzyme. Daratumumab, an anti-CD38 antibody, is currently in the clinical trials for multiple myeloma. RESULTS:Here w...

    journal_title:BMC biotechnology

    pub_type: 杂志文章


    authors: Yu T,Qiao C,Lv M,Tang L

    更新日期:2019-05-22 00:00:00

  • Genetic and spectrally distinct in vivo imaging: embryonic stem cells and mice with widespread expression of a monomeric red fluorescent protein.

    abstract:BACKGROUND:DsRed the red fluorescent protein (RFP) isolated from Discosoma sp. coral holds much promise as a genetically and spectrally distinct alternative to green fluorescent protein (GFP) for application in mice. Widespread use of DsRed has been hampered by several issues resulting in the inability to establish and...

    journal_title:BMC biotechnology

    pub_type: 杂志文章


    authors: Long JZ,Lackan CS,Hadjantonakis AK

    更新日期:2005-07-04 00:00:00

  • Production of soluble mammalian proteins in Escherichia coli: identification of protein features that correlate with successful expression.

    abstract:BACKGROUND:In the search for generic expression strategies for mammalian protein families several bacterial expression vectors were examined for their ability to promote high yields of soluble protein. Proteins studied included cell surface receptors (Ephrins and Eph receptors, CD44), kinases (EGFR-cytoplasmic domain, ...

    journal_title:BMC biotechnology

    pub_type: 杂志文章


    authors: Dyson MR,Shadbolt SP,Vincent KJ,Perera RL,McCafferty J

    更新日期:2004-12-14 00:00:00

  • Characterization of purified and xerogel immobilized novel lignin peroxidase produced from Trametes versicolor IBL-04 using solid state medium of corncobs.

    abstract:BACKGROUND:Cost-effective production of industrially important enzymes is a key for their successful exploitation on industrial scale. Keeping in view the extensive industrial applications of lignin peroxidase (LiP), this study was performed to purify and characterize the LiP from an indigenous strain of Trametes versi...

    journal_title:BMC biotechnology

    pub_type: 杂志文章


    authors: Asgher M,Iqbal HM,Irshad M

    更新日期:2012-08-03 00:00:00

  • Integration, abundance, and transmission of mutations and transgenes in a series of CRISPR/Cas9 soybean lines.

    abstract:BACKGROUND:As with many plant species, current genome editing strategies in soybean are initiated by stably transforming a gene that encodes an engineered nuclease into the genome. Expression of the transgene results in a double-stranded break and repair at the targeted locus, oftentimes resulting in mutation(s) at the...

    journal_title:BMC biotechnology

    pub_type: 杂志文章


    authors: Michno JM,Virdi K,Stec AO,Liu J,Wang X,Xiong Y,Stupar RM

    更新日期:2020-02-24 00:00:00

  • Different genome-wide transcriptome responses of Nocardioides simplex VKM Ac-2033D to phytosterol and cortisone 21-acetate.

    abstract:BACKGROUND:Bacterial degradation/transformation of steroids is widely investigated to create biotechnologically relevant strains for industrial application. The strain of Nocardioides simplex VKM Ac-2033D is well known mainly for its superior 3-ketosteroid Δ1-dehydrogenase activity towards various 3-oxosteroids and oth...

    journal_title:BMC biotechnology

    pub_type: 杂志文章


    authors: Shtratnikova VY,Sсhelkunov MI,Fokina VV,Bragin EY,Shutov AA,Donova MV

    更新日期:2021-01-13 00:00:00

  • Selection of oleaginous yeasts for fatty acid production.

    abstract:BACKGROUND:Oleaginous yeast species are an alternative for the production of lipids or triacylglycerides (TAGs). These yeasts are usually non-pathogenic and able to store TAGs ranging from 20 % to 70 % of their cell mass depending on culture conditions. TAGs originating from oleaginous yeasts can be used as the so-call...

    journal_title:BMC biotechnology

    pub_type: 杂志文章


    authors: Lamers D,van Biezen N,Martens D,Peters L,van de Zilver E,Jacobs-van Dreumel N,Wijffels RH,Lokman C

    更新日期:2016-05-27 00:00:00

  • Stable high-level expression of factor VIII in Chinese hamster ovary cells in improved elongation factor-1 alpha-based system.

    abstract:BACKGROUND:Recombinant factor VIII (FVIII), used for haemophilia A therapy, is one of the most challenging among the therapeutic proteins produced in heterologous expression systems. Deletion variant of FVIII, in which the entire domain B is replaced by a short linker peptide, was approved for medical use. Efficacy and...

    journal_title:BMC biotechnology

    pub_type: 杂志文章


    authors: Orlova NA,Kovnir SV,Gabibov AG,Vorobiev II

    更新日期:2017-03-24 00:00:00

  • Decreasing translation error rate in Escherichia coli increases protein function.

    abstract:BACKGROUND:Over-expressed native or recombinant proteins are commonly used for industrial and pharmaceutical purposes, as well as for research. Proteins of interest need to be purified in sufficient quantity, quality and specific activity to justify their commercial price and eventual medical use. Proteome quality was ...

    journal_title:BMC biotechnology

    pub_type: 杂志文章


    authors: Musa M,Radman M,Krisko A

    更新日期:2016-03-11 00:00:00

  • Evaluating the use of a CpG free promoter for long-term recombinant protein expression stability in Chinese hamster ovary cells.

    abstract:BACKGROUND:Methylated CpG dinucleotides in promoters are associated with the loss of gene expression in recombinant Chinese hamster ovary (CHO) cells during large-scale commercial manufacturing. We evaluated a promoter devoid of CpG dinucleotides, CpGfree, in parallel with a similar CpG containing promoter, CpGrich, fo...

    journal_title:BMC biotechnology

    pub_type: 杂志文章


    authors: Ho SC,Koh EY,Soo BP,Mariati,Chao SH,Yang Y

    更新日期:2016-10-18 00:00:00

  • Expression of Vitreoscilla hemoglobin enhances production of arachidonic acid and lipids in Mortierella alpina.

    abstract:BACKGROUND:Arachidonic acid (ARA, C20:4, n-6), which belongs to the omega-6 series of polyunsaturated fatty acids and has a variety of biological activities, is commercially produced in Mortierella alpina. Dissolved oxygen or oxygen utilization efficiency is a critical factor for Mortierella alpina growth and arachidon...

    journal_title:BMC biotechnology

    pub_type: 杂志文章


    authors: Zhang H,Feng Y,Cui Q,Song X

    更新日期:2017-08-30 00:00:00

  • An adenovirus prime/plasmid boost strategy for induction of equipotent immune responses to two dengue virus serotypes.

    abstract:BACKGROUND:Dengue is a public health problem of global significance for which there is neither an effective antiviral therapy nor a preventive vaccine. It is a mosquito-borne viral disease, caused by dengue (DEN) viruses, which are members of the Flaviviridae family. There are four closely related serotypes, DEN-1, DEN...

    journal_title:BMC biotechnology

    pub_type: 杂志文章


    authors: Khanam S,Rajendra P,Khanna N,Swaminathan S

    更新日期:2007-02-15 00:00:00

  • Structural insights from random mutagenesis of Campylobacter jejuni oligosaccharyltransferase PglB.

    abstract:BACKGROUND:Protein glycosylation is of fundamental importance in many biological systems. The discovery of N-glycosylation in bacteria and the functional expression of the N-oligosaccharyltransferase PglB of Campylobacter jejuni in Escherichia coli enabled the production of engineered glycoproteins and the study of the...

    journal_title:BMC biotechnology

    pub_type: 杂志文章


    authors: Ihssen J,Kowarik M,Wiesli L,Reiss R,Wacker M,Thöny-Meyer L

    更新日期:2012-09-24 00:00:00

  • Comparison and transfer testing of multiplex ligation detection methods for GM plants.

    abstract:BACKGROUND:With the increasing number of GMOs on the global market the maintenance of European GMO regulations is becoming more complex. For the analysis of a single food or feed sample it is necessary to assess the sample for the presence of many GMO-targets simultaneously at a sensitive level. Several methods have be...

    journal_title:BMC biotechnology

    pub_type: 杂志文章


    authors: Ujhelyi G,Dijk JP,Prins TW,Voorhuijzen MM,Hoef AM,Beenen HG,Morisset D,Gruden K,Kok EJ

    更新日期:2012-01-19 00:00:00