Efficient lipofection with cisplatin-resistant human tumor cells.

L Huang; K Son; X Gao; D Hages; Y Y Yang; S A Holden; B Teicher; J Kirkwood; J S Lazo

PubMed · 8729909

Efficient lipofection with cisplatin-resistant human tumor cells.

L Huang·K Son·X Gao·D Hages·Y Y Yang·S A Holden·B Teicher·J Kirkwood·J S Lazo

Abstract

Seven of seven different cisplatin-resistant human tumor cell lines showed elevated lipofection activity as compared with their sensitive parent cells, although the degree of enhancement was not quantitatively correlated with the degree of cisplatin resistance. Enhanced transfection was seen by using the same reporter gene driven by three different promoter/enhancer sequencer or by using different reporter genes driven by the same promoter/enhancer. Cells resistant to actinomycin D, bleomycin, and nitrogen mustards were not more transfectable than the sensitive parent cells. Although the mechanism of enhanced transfection in cisplatin-resistant cells is not known, data indicated that enhanced transcription, multidrug-resistant phenotype, and methallothionein overexpression do not play a role.

Explore related subjects

Chloramphenicol O-Acetyltransferase Cisplatin Drug Resistance, Neoplasm Drug Screening Assays, Antitumor Enhancer Elements, Genetic Gene Expression Regulation, Neoplastic Genes, Reporter Genetic Therapy Humans Liposomes Luciferases Metallothionein Neoplasms Transfection Tumor Cells, Cultured Journal Article Research Support, U.S. Gov't, P.H.S.

Keep this discovery

Explore connections, maps & timelines

BibTeX RIS

L Huang, K Son, X Gao, D Hages, Y Y Yang, S A Holden, B Teicher, J Kirkwood, J S Lazo. Efficient lipofection with cisplatin-resistant human tumor cells.. https://pubmed.ncbi.nlm.nih.gov/8729909/

Cite the original work for its findings. Save a collection to share your selection of sources.

KEEP EXPLORING

Related citations

PubMed2007

Versatile DNA fragmentation and directed evolution with nucleotide exchange and excision technology.

Mimicking natural evolution by DNA shuffling is a commonly used method for the optimization of DNA and protein properties. Here, we present an advancement of this approach whereby a gene library is amplified using a standard polymerase chain reaction (PCR), but incorporates dUTP as a fragmentation-defining exchange nucleotide, together with the four standard dNTPs. Incorporated uracil bases are excised using uracil-DNA-glycosylase, and the DNA backbone subsequently is cleaved with piperidine. This oligonucleotide pool is then reassembled with an internal primer extension procedure using a proofreading polymerase to increase yield, and, finally, is amplified by PCR. Denaturing polyacrylamide urea gels demonstrate this method to produce adjustable fragmentation size ranges dependent on the dUTP:dTTP ratios. Using the model protein, chloramphenicol acetyltransferase I, the sequencing of shuffled gene libraries based on a PCR containing 33% dUTP revealed a low mutation rate, of approx 0.1%, with an average parental fragments size of 86 bases, even without the use of a fragment-size separation. Nucleotide exchange and excision technology (NExT) DNA shuffling is, thus, reproducible and easily executed, making it superior to competing techniques. Additionally, NExT fragmentation outcome can be predicted using the computer software, NExTProg.

Chloramphenicol O-Acetyltransferase↗

PubMed2006

Influence of global fluorination on chloramphenicol acetyltransferase activity and stability.

Varied levels of fluorinated amino acid have been introduced biosynthetically to test the functional limits of global substitution on enzymatic activity and stability. Replacement of all the leucine (LEU) residues in the enzyme chloramphenicol acetyltransferase (CAT) with the analog, 5',5',5'-trifluoroleucine (TFL), results in the maintenance of enzymatic activity under ambient temperatures as well as an enhancement in secondary structure but loss in stability against heat and denaturants or organic co-solvents. Although catalytic activity of the fully substituted CAT is preserved under standard reaction conditions compared to the wild-type enzyme both in vitro and in vivo, as the incorporation levels increase, a concomitant reduction in thermostability and chemostability is observed. Circular dichroism (CD) studies reveal that although fluorination greatly improves the secondary structure of CAT, a large structural destabilization upon increased levels of TFL incorporation occurs at elevated temperatures. These data suggest that enhanced secondary structure afforded by TFL incorporation does not necessarily lead to an improvement in stability.

Chloramphenicol O-Acetyltransferase↗

PubMed2006

Production of milligram quantities of recombinant proteins from PCR-amplified DNAs in a continuous-exchange cell-free protein synthesis system.

Explore the source record for details and available documents.

Chloramphenicol O-Acetyltransferase↗