Gfp-scap Plasmid
Gfp-scap Plasmid - Related Videos
11:27
Analysis of SCAP N-glycosylation and Trafficking in Human Cells
0 Views •
Cited by 17 •
2016
We describe a modified method for membrane fraction isolation from human cells and sample preparation for the detection of SCAP N-glycosylation and total protein by using western blot. We further introduce a GFP-labeling method to monitor SCAP trafficking using confocal microscopy. This protocol can be used in regular biology laboratories.
08:16
Plasmid Purification
0 Views •
2023
Plasmid purification is a technique used to isolate and purify plasmid DNA from genomic DNA, proteins, ribosomes, and the bacterial cell wall. A plasmid is a small, circular, double-stranded DNA that is used as a carrier of specific DNA molecules. When introduced into a host organism via transformation, a plasmid will be replicated, creating numerous copies of the DNA fragment under study. In this video, a step-by-step generalized procedure is described for how to perform plasmid purification.
01:28
Plasmids
0 Views •
2025
Plasmids are extrachromosomal DNA molecules found in bacteria, archaea, and some eukaryotic microbes like yeast. These small, circular DNA structures typically contain fewer than 30 genes, although some may exist linearly. Plasmids vary in their number within a cell, known as copy number. Single-copy plasmids are present in one copy per cell and multi-copy plasmids are present in multiple copies, reaching over 100 copies per cell.Plasmids usually replicate independently of the chromosomal DNA...
07:11
Homemade Site Directed Mutagenesis of Whole Plasmids
0 Views •
Cited by 91 •
2009
Site directed mutagenesis of whole plasmids is a simple way to create slightly different variations of an original plasmid. Here we demonstrate an easy and cost effective way to introduce base substitutions into a plasmid using standard reagents.
03:44
Generation of GFP-Tagged Recombinant Respiratory Syncytial Virus Using a T7 Expression System
0 Views •
2026
Source: Bouillier, C. et. al., Amplification, and Titration of Recombinant Respiratory Syncytial Viruses. J. Vis. Exp. (2019)This video demonstrates the generation of a green fluorescent protein (GFP)-expressing recombinant respiratory syncytial virus using a T7-based reverse genetics system. The co-transfection of plasmids encoding viral proteins and a GFP-tagged genome leads to protein expression and initiates virus rescue.
FAQs
SCAP links cellular sterol status with SREBP trafficking. When sterol levels fall, SCAP helps move SREBP from the endoplasmic reticulum to the Golgi. There, SREBP processing activates genes that support cholesterol and fatty-acid synthesis. This makes SCAP a regulatory connection between lipid abundance and the transcriptional response that restores lipid balance.
The GFP tag provides a fluorescence signal that marks the intracellular location of SCAP-SCAP? Actually, GFP-SCAP allows researchers to follow where SCAP is positioned in cultured cells as sterol conditions change. Changes in fluorescence distribution can indicate sterol-dependent trafficking, helping connect SCAP movement with regulation of SREBP signaling and cellular lipid metabolism.
Localization changes provide a visual readout of the regulatory state controlled by sterols. Under low-sterol conditions, movement of the GFP-SCAP signal is relevant because SCAP supports SREBP transport toward the Golgi, where processing activates lipid-synthesis genes. Comparing these patterns helps researchers relate cellular fluorescence observations to cholesterol homeostasis rather than treating localization as an isolated image.
A typical analysis uses cultured cells containing the GFP-SCAP plasmid, followed by fluorescence-based observation of SCAP distribution under differing sterol conditions. Researchers compare where the signal appears and how its pattern changes when sterol availability changes. This approach provides a cellular view of trafficking behavior that can be linked to the SREBP pathway and lipid regulation.
GFP-SCAP imaging can show whether SCAP localization responds to sterol status in a manner consistent with regulated SREBP transport. These observations help investigators study how cells coordinate cholesterol and fatty-acid synthesis with lipid availability. The method therefore connects intracellular trafficking patterns with broader mechanisms that maintain cellular lipid homeostasis.
Because SCAP participates in sterol-sensitive control of SREBP signaling, the plasmid can help investigate cellular mechanisms associated with dysregulated cholesterol metabolism. Fluorescence-based tracking may reveal altered localization or trafficking behavior in experimental cell systems. Such observations provide mechanistic context for understanding how disrupted lipid regulation could contribute to disease-related metabolic changes.