scholarly journals Baculovirus-Transfer Vector for Eukaryotic Expression and Immunoaffinity Purification of Gal4-Fusion Proteins

BioTechniques ◽  
1999 ◽  
Vol 26 (5) ◽  
pp. 808-814 ◽  
Author(s):  
Guoqing Chen ◽  
Albert J. Courey
Keyword(s):  
Fusion Proteins ◽  
Eukaryotic Expression ◽  
Transfer Vector ◽  
Immunoaffinity Purification ◽  
Baculovirus Transfer Vector

scholarly journals Targeted regulation of plasmid DNA expression in eukaryotic cells with a methylated-DNA-binding activator

2021 ◽  
Author(s):  
Isioma Enwerem-Lackland ◽  
Eric Warga ◽  
Margaret Dugoni ◽  
Jacob Elmer ◽  
Karmella A. Haynes
Keyword(s):  
Dna Binding ◽  
Plasmid Dna ◽  
Fusion Proteins ◽  
Genomic Dna ◽  
Transcriptional Start Site ◽  
Eukaryotic Expression ◽  
Expression Vectors ◽  
Binding Region ◽  
Methylated Dna ◽  
Dependent Activity

Purpose: Targeted regulation of transfected extra-chromosomal plasmid DNA typically requires the integration of 9 - 20 bp docking sites into the plasmid. Here, we report an elegant approach, The Dpn Adaptor Linked Effector (DAL-E) system, to target fusion proteins to 6-methyladenosine in GATC, which appears frequently in popular eukaryotic expression vectors and is absent from endogenous genomic DNA. Methods: The DNA-binding region from the DpnI endonuclease binds 6-methyladenosine within the GATC motif. We used a Dpn-transcriptional activator (DPN7-TA) fusion to induce gene expression from transiently transfected pDNAs. Results: We validated methylation-dependent activity of DPN7-TA with a panel of target pDNAs. We observed stronger transactivation when GATC targets were located upstream of the transcriptional start site in the target pDNA. Conclusion: DAL-E, consisting of a 108 aa, 12 kD DNA-binding adaptor and a 4 bp recognition site, offers a genetically-tractable, tunable system that can potentially be redesigned to recruit a variety of regulators (e.g. activators, silencers, epigenome editors) to transfected plasmid DNA.

scholarly journals A Method for Rapid Screening, Expression, and Purification of Antimicrobial Peptides

2021 ◽  
Vol 9 (9) ◽  
pp. 1858
Author(s):  
Yingli Zhang ◽  
Zhongchen Li ◽  
Li Li ◽  
Ben Rao ◽  
Lixin Ma ◽  
...  
Keyword(s):  
Antimicrobial Peptides ◽  
Fusion Proteins ◽  
Expression System ◽  
Inhibition Zone ◽  
Eukaryotic Expression ◽  
Biologically Active ◽  
Rapid Screening ◽  
Expression Vectors ◽  
Expression And Purification ◽  
E Coli

In this study, a method for the rapid screening, expression and purification of antimicrobial peptides (AMPs) was developed. AMP genes were fused to a heat-resistant CL7 tag using the SLOPE method, and cloned into Escherichia coli and Pichia pastoris expression vectors. Twenty E. coli and ten P. pastoris expression vectors were constructed. Expression supernatants were heated, heteroproteins were removed, and fusion proteins were purified by nickel affinity (Ni-NTA) chromatography. Fusion proteins were digested on the column using human rhinovirus (HRV) 3C protease, and AMPs were released and further purified. Five AMPs (1, 2, 6, 13, 16) were purified using the E. coli expression system, and one AMP (13) was purified using the P. pastoris expression system. Inhibition zone and minimum inhibitory concentration (MIC) tests confirmed that one P. pastoris¬-derived and two E. coli-derived AMPs have the inhibition activity. The MIC of AMP 13 and 16 from E. coli was 24.2 μM, and the MIC of AMP 13 from P. pastoris was 8.1 μM. The combination of prokaryotic and eukaryotic expression systems expands the universality of the developed method, facilitating screening of a large number of biologically active AMPs, establishing an AMP library, and producing AMPs by industrialised biological methods.

Efficient eukaryotic expression of fluorescent scFv fusion proteins directed against CD antigens for FACS applications

2004 ◽  
Vol 285 (2) ◽  
pp. 265-280 ◽  
Author(s):  
Matthias Peipp ◽  
Domenica Saul ◽  
Karin Barbin ◽  
Joerg Bruenke ◽  
Susan J. Zunino ◽  
...  
Keyword(s):  
Fusion Proteins ◽  
Eukaryotic Expression ◽  
Cd Antigens

Identification of common and distinct epigenetic re-programming properties of Core-Binding Factor (CBF) Fusion Proteins

2016 ◽  
Vol 228 (03) ◽  
Author(s):  
J Loke ◽  
A Ptasinska ◽  
MR Imperato ◽  
SA Assi ◽  
P Cauchy ◽  
...  
Keyword(s):  
Fusion Proteins ◽  
Core Binding Factor ◽  
Binding Factor
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