%0 Journal Article %1 incorporation %A Haque, F %A Geng, J %A Montemagno, C %A Guo, P %D 2013 %J Nat Protoc %K phage %N 2 %P 373-392 %R 10.1038/nprot.2013.001 %T Incorporation of a viral DNA-packaging motor channel in lipid bilayers for real-time, single-molecule sensing of chemicals and double-stranded DNA %U http://www.ncbi.nlm.nih.gov/pubmed/23348364 %V 8 %X Over the past decade, nanopores have rapidly emerged as stochastic biosensors. This protocol describes the cloning, expression and purification of the channel of the bacteriophage phi29 DNA-packaging nanomotor and its subsequent incorporation into lipid membranes for single-pore sensing of double-stranded DNA (dsDNA) and chemicals. The membrane-embedded phi29 nanochannel remains functional and structurally intact under a range of conditions. When ions and macromolecules translocate through this nanochannel, reliable fingerprint changes in conductance are observed. Compared with other well-studied biological pores, the phi29 nanochannel has a larger cross-sectional area, which enables the translocation of dsDNA. Furthermore, specific amino acids can be introduced by site-directed mutagenesis within the large cavity of the channel to conjugate receptors that are able to bind specific ligands or analytes for desired applications. The lipid membrane-embedded nanochannel system has immense potential nanotechnological and biomedical applications in bioreactors, environmental sensing, drug monitoring, controlled drug delivery, early disease diagnosis and high-throughput DNA sequencing. The total time required for completing one round of this protocol is around 1 month.

@article{incorporation, abstract = {Over the past decade, nanopores have rapidly emerged as stochastic biosensors. This protocol describes the cloning, expression and purification of the channel of the bacteriophage phi29 DNA-packaging nanomotor and its subsequent incorporation into lipid membranes for single-pore sensing of double-stranded DNA (dsDNA) and chemicals. The membrane-embedded phi29 nanochannel remains functional and structurally intact under a range of conditions. When ions and macromolecules translocate through this nanochannel, reliable fingerprint changes in conductance are observed. Compared with other well-studied biological pores, the phi29 nanochannel has a larger cross-sectional area, which enables the translocation of dsDNA. Furthermore, specific amino acids can be introduced by site-directed mutagenesis within the large cavity of the channel to conjugate receptors that are able to bind specific ligands or analytes for desired applications. The lipid membrane-embedded nanochannel system has immense potential nanotechnological and biomedical applications in bioreactors, environmental sensing, drug monitoring, controlled drug delivery, early disease diagnosis and high-throughput DNA sequencing. The total time required for completing one round of this protocol is around 1 month.}, added-at = {2014-02-24T11:54:25.000+0100}, author = {Haque, F and Geng, J and Montemagno, C and Guo, P}, biburl = {https://www.bibsonomy.org/bibtex/2e89a5ad951f1a372a35fccbb497d2aae/ross_mck}, description = {Incorporation of a viral DNA-packaging motor chan... [Nat Protoc. 2013] - PubMed - NCBI}, doi = {10.1038/nprot.2013.001}, interhash = {2862d016cc3c7403d936263283a6a0ec}, intrahash = {e89a5ad951f1a372a35fccbb497d2aae}, journal = {Nat Protoc}, keywords = {phage}, month = feb, number = 2, pages = {373-392}, pmid = {23348364}, timestamp = {2014-02-24T11:54:25.000+0100}, title = {Incorporation of a viral DNA-packaging motor channel in lipid bilayers for real-time, single-molecule sensing of chemicals and double-stranded DNA}, url = {http://www.ncbi.nlm.nih.gov/pubmed/23348364}, volume = 8, year = 2013 }