PMID: 8604299Feb 15, 1996Paper

Transfecting mammalian cells: optimization of critical parameters affecting calcium-phosphate precipitate formation

Nucleic Acids Research
M JordanF M Wurm

Abstract

DNA-calcium phosphate co-precipitates arise spontaneously in supersaturated solutions. Highly effective precipitates for transfection purposes, however, can be generated only in a very narrow range of physico-chemical conditions that control the initiation and growth of precipitate complexes. The concentrations of calcium and phosphate are the main factors influencing characteristics of the precipitate complex, but other parameters, such as temperature, DNA concentration and reaction time are important as well. An example for this is the finding that almost all of the soluble DNA in the reaction mix can be bound into an insoluble complex with calcium phosphate in <1 min. Extending the reaction time to 20 min results in aggregation and/or growth of particles and reduces the level of expression. With improved protocols we gained better reproducibility and higher efficiencies both for transient and for stable transfections. Up to 60% of cells stained positive for beta-gal and transient production of secreted proteins was improved 5- to 10-fold over results seen with transfections using standard procedures. Similar improvements in efficiency (number of recombinant cell colonies) were observed with stable transfections, using co-tra...Continue Reading

References

May 1, 1990·Protein Engineering·L R PaborskyC M Gorman
Jul 1, 1990·Biologicals : Journal of the International Association of Biological Standardization·F M Wurm
Mar 25, 1989·Nucleic Acids Research·G R MacGregor, C T Caskey
Aug 1, 1987·Molecular and Cellular Biology·C Chen, H Okayama
Apr 1, 1973·Analytical Biochemistry·J A Zivin, J F Snarr
Apr 1, 1973·Virology·F L Graham, A J van der Eb
Jun 1, 1981·Journal of Applied Toxicology : JAT·J S LeeA A Seawright
Jul 1, 1980·Proceedings of the National Academy of Sciences of the United States of America·G Urlaub, L A Chasin
Jan 1, 1982·Proceedings of the National Academy of Sciences of the United States of America·A LoyterF H Ruddle
Sep 15, 1993·Proceedings of the National Academy of Sciences of the United States of America·W S PearD Baltimore
Apr 26, 1994·Proceedings of the National Academy of Sciences of the United States of America·B A KeytJ Ogez

❮ Previous
Next ❯

Citations

Feb 24, 2001·The Journal of Gene Medicine·J FominayaA Bernad
Sep 5, 2002·Biotechnology and Bioengineering·M BoorsmaM F Bachmann
Apr 15, 2006·Biotechnology Letters·Madiha DerouaziFlorian M Wurm
Sep 30, 2006·Biotechnology Letters·Frederic GrosjeanFlorian M Wurm
Jan 30, 2004·Biochimica Et Biophysica Acta·Jeanette LindellFlorian Wurm
May 18, 2005·The International Journal of Biochemistry & Cell Biology·Shuang ShiDalong Ma
May 5, 2004·Fish & Shellfish Immunology·Suhee HongChris J Secombes
Jan 27, 2004·Journal of Biotechnology·D S Anson, M Limberis
Jul 20, 2002·Archives of Biochemistry and Biophysics·Donghui MaKandiah Jeyaseelan
Aug 6, 2003·Journal of Molecular Biology·Martin BarilLéa Brakier-Gingras
Aug 23, 2001·Biomaterials·M OkazakiJ C Elliott
Mar 6, 1998·Journal of Biotechnology·U von Stockar, L A van der Wielen
Apr 21, 1999·Current Opinion in Biotechnology·F Wurm, A Bernard
Mar 31, 2011·Journal of Medicinal Chemistry·Jürgen EinsiedelPeter Gmeiner
Nov 13, 2008·Cytotechnology·E J Schlaeger, K Christensen
Nov 13, 2008·Cytotechnology·Philippe GirardFlorian M Wurm
Jun 6, 2006·Nature Immunology·Anouk CarauxClaude Roth

❮ Previous
Next ❯

Related Concepts

Trending Feeds

COVID-19

Coronaviruses encompass a large family of viruses that cause the common cold as well as more serious diseases, such as the ongoing outbreak of coronavirus disease 2019 (COVID-19; formally known as 2019-nCoV). Coronaviruses can spread from animals to humans; symptoms include fever, cough, shortness of breath, and breathing difficulties; in more severe cases, infection can lead to death. This feed covers recent research on COVID-19.

Blastomycosis

Blastomycosis fungal infections spread through inhaling Blastomyces dermatitidis spores. Discover the latest research on blastomycosis fungal infections here.

Nuclear Pore Complex in ALS/FTD

Alterations in nucleocytoplasmic transport, controlled by the nuclear pore complex, may be involved in the pathomechanism underlying multiple neurodegenerative diseases including Amyotrophic Lateral Sclerosis and Frontotemporal Dementia. Here is the latest research on the nuclear pore complex in ALS and FTD.

Applications of Molecular Barcoding

The concept of molecular barcoding is that each original DNA or RNA molecule is attached to a unique sequence barcode. Sequence reads having different barcodes represent different original molecules, while sequence reads having the same barcode are results of PCR duplication from one original molecule. Discover the latest research on molecular barcoding here.

Chronic Fatigue Syndrome

Chronic fatigue syndrome is a disease characterized by unexplained disabling fatigue; the pathology of which is incompletely understood. Discover the latest research on chronic fatigue syndrome here.

Evolution of Pluripotency

Pluripotency refers to the ability of a cell to develop into three primary germ cell layers of the embryo. This feed focuses on the mechanisms that underlie the evolution of pluripotency. Here is the latest research.

Position Effect Variegation

Position Effect Variagation occurs when a gene is inactivated due to its positioning near heterochromatic regions within a chromosome. Discover the latest research on Position Effect Variagation here.

STING Receptor Agonists

Stimulator of IFN genes (STING) are a group of transmembrane proteins that are involved in the induction of type I interferon that is important in the innate immune response. The stimulation of STING has been an active area of research in the treatment of cancer and infectious diseases. Here is the latest research on STING receptor agonists.

Microbicide

Microbicides are products that can be applied to vaginal or rectal mucosal surfaces with the goal of preventing, or at least significantly reducing, the transmission of sexually transmitted infections. Here is the latest research on microbicides.