Linked Life Data

21356979

Source:http://linkedlifedata.com/resource/pubmed/id/21356979

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:dateCreated
2011-3-1
pubmed:abstractText
INTRODUCTIONThis protocol describes a method for establishing a green fluorescent protein (GFP) calibration curve using dilutions of recombinant GFP and blue fluorescent beads. The total fluorescence intensity (TFI) per mRNA molecule is first calculated by imaging serial dilutions of purified enhanced GFP (eGFP) to determine the TFI within a specific volume. A calibration curve of fluorescence intensity in a given voxel per molecule of GFP is then used to determine the number of GFP molecules in the sample of formaldehyde fixed cells to be imaged. This is followed by a method for detection of single molecules in formaldehyde-fixed and live cells. These cells have been cotransfected with mRNA reporter and MCP-xFP plasmids, where MCP-xFP refers to a fluorescent protein fused to the MS2 capsid protein. It is important to collect micrographs and establish the calibration curve on the same day that the cells are imaged, using the same equipment configuration, camera settings, and image acquisition parameters.
pubmed:language
eng
pubmed:journal
pubmed:status
PubMed-not-MEDLINE
pubmed:author
pubmed:volume
2007
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
pdb.prot4871
pubmed:year
2007
pubmed:articleTitle
Imaging real-time gene expression in living systems with single-transcript resolution: image analysis of single mRNA transcripts.
pubmed:affiliation
Department of Anatomy and Structural Biology, Albert Einstein College of Medicine, Bronx, NY 10461, USA.
pubmed:publicationType
Journal Article