Disclaimer Introduction This document was originally designed to focus on fire situations in chemistry laboratories. While it continues to mention laboratories, the information contained herein is broadly applicable to almost all workplaces. Fire is the most common serious hazard that one faces in a typical chemistry laboratory.
Transfection products are manufactured by Altogen Biosystems. Transfection Transfection is the use of chemical liposomes, chemicals or mechanical electroporation means to transfer nucleic acids into cells in order to alter mRNA expression or the genetic makeup of a genome i. Transfection is used to develop genetic therapies for diseases such as obesity, AIDS, cancer and cardiac issues.
Gene silencing called RNA interference or RNAi is a technique through which the The lab protocols levels of a gene are turned off or knocked down. Although a gene may be silenced at either the transcriptional or post-transcriptional stages, RNAi is specifically a post-transcriptional gene silencing mechanism achieved by transfecting a dsRNA into a cell.
Non-viral delivery of DNA or RNA requires a transfection reagent or transfection method with multi-faceted capabilities. Attributes of a good transfection reagent include protecting the payload from harsh environments that can lead to degradation from RNases and DNases by forming a tight bilayer hydrophobic exterior, hydrophilic interior.
A successful transfection reagent also readily loads with a high ratio of active pharmaceutical ingredient API to delivery agent ratio.
This ultimately leads to lower manufacturing costs of the transfection reagent since lower concentrations can be used per test sample. Perhaps the most crucial aspect of a transfection reagent is size. With an understanding of cellular pore size and an extensive innate response to foreign material in an organism, transfection reagents must have final structure sizes that can bypass mechanisms that trigger immune responses.
Optimal transfection reagent compound size should fall in a range of nm. Chemical transfection is the uptake of a test reagent either through the liposome binding to the cell membrane, endocytosis or interaction with a cell surface marker. Mechanical transfection is the physical movement of proteins or nucleic acids into the cell, via electrical current.
Other methods exist, such as viral delivery of DNA into cells termed transduction. A similar process to transduction of delivering plasmid DNA into bacteria or non-animal eukaryotic cells is called transformation.
Whether it is by chemical or mechanical transfection, the idea of transfection is to create an opening in the cell membrane to allow exogenous genetic material to pass into the cytoplasm. The delivered payload either remains in the cytoplasm for transcriptional regulation or continues to the nucleus for integration into the genomic DNA and stable expression of the payload plasmid DNA.
Transfection is used to deliver nucleic acids to cells in the creation of transgenic mouse models, genetically modified plants and even genetically modified pets. In Vitro Transfection Manipulation of gene expression by transfection of cultured cells enables a fully controlled, clean environment for the study of apoptosis, cell cycle, target gene expression and mechanism of action.
Although cell culture studies do not require a transfection reagent to protect the payload from the harsh environment of circulation in animals, it must provide protection from degrading factors in cell culture medium and FBS i. Liposomes are small capsules made of polymers similar to those that make up the cellular membrane.
When formulated with nucleic acids, lipid transfection reagents engage with the cell membrane and release their cargo into the cells. Lipid based transfection reagents utilize the semi-permeable feature of cell membranes to transport nucleic acid molecules to the interior of a cell.
The transfection complexes overcome the cell membrane by endocytosis and the nucleic acids are released into cytoplasm.Database of research protocols in bioscience including molecular biology, cell biology, immunology, plant biology, developmental biology, neuroscience, bioinformatics, genetics, etc.
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These are protocols that are used in the Bowdish lab. We hope that these are in sufficient detail as to be helpful for you but if not please don’t be shy. This is a compilation of laboratory protocols, local sources of research resources, and safety information for lab researchers. For assistance with these resources, please contact the author of this guide or your librarian. Yeast Methods: The Forsburg lab website is an excellent source of S. pombe information: Yast-Two-Hybrid Interaction Trap: See the protocols developed by the Brent lab C. elegans Methods General.
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