Accelerated Sensor Of Action Potentials

 By restriction enzymes then amplified by polymerase chain reaction to make many to millions of copies of a single fragment.

that goes into creating a clone. First, scientist remove a somatic cell from an animal that they

134). They are loops of DNA that are separate from the chromosomal DNA and can self-replicate in a cell, found mostly in bacteria (Brown, 2011; Addgene, 2015). Lederberg and William Hayes discovered that plasmids were being transferred from one cell to another, not the chromosomal DNA (Brown, 2011, p. 135). This discovery lead to plasmids being an essential tool for scientists. Scientists can engineer plasmids to have specific genes to introduce into new cells (Brown, 2011, p. 134). On a plasmid loop there will be an origin of replication (ORI) and a multiple cloning site (MCS) where the gene of interest is inserted (Bio-Rad, 2015). This region has specific restriction enzyme recognition sites, which are cut by the enzymes to open up the DNA where the new gene will be inserted (Jove Science Education Database, 2015). Most plasmids will also contain an antibiotic resistance gene allowing cell survival in environments containing antibiotics (Jove Science Education Database, 2015).

This lab is about moving genes from one thing to another using plasmids. Plasmid has the ability to replicate, so it replicates independently, and separately from the chromosomal DNA. Plasmid are one or more small piece of DNA and they enter cells as a double strand DNA. When they enter the cell as a doubke strand they do not invade he chromosomal DNA. We will also transform bacteria into GFP which is mainly from the jelly fish Aequorea Victoria. The GFP causes the the jelly fish to fluorescent and glow in the dark. After the transformation, bacteria starts to make the GFP which causes them to glow a green color under a ultraviolet light.

Green Fluorescent Protein, produced by the bioluminescent jellyfish Aequorea victoria, is a protein that fluoresces green under ultraviolet light. Since its discovery, properties of the protein have been improved by mutations in the gene resulting in the expansion of its spectrum, which now contains brighter variants and multiple different colors. GFP is used in a wide variety of applications and technologies. Its many different applications have contributed greatly, and continue to do so, in numerous fields of study including, but not limited to, cellular and molecular biology, microbiology, biotechnology, and medicine.

This process uses enzymes to cut and insert piece of DNA into a plasmid vector. This vector is then transfected into cells. The different pieces of DNA that were being inserted into this plasmid were a control and three different mutants (an alpha-5-tailless mutant, full-length alpha-5, and full-length alpha-2). These pieces of DNA code for integrins. Once these pieces of DNA were inserted into the plasmid vector, the plasmid needed to be inserted into a cell where it would be able to replicate. In these tests, rat intestinal epithelial wild-type cells (cells that would be found in nature, not cells grown in the laboratory) were used. To insert the plasmid into these cells lipofectamine plus was used. Lipofectamine plus acts like a detergent and opens the membrane of the cell so the plasmid can enter. Also, the plasmids that were transfected into cells were all treated with G-418, an antibiotic. When the cells were plated and allowed to multiply, only those containing the plasmid with the antibiotic would live. This made it possible to know which cells actually received the plasmid and properly underwent mitosis, and to eliminate those cells that never took up the plasmid in the first place.

Green Fluorescent Protein (GFP) is a fluorescent protein found in jellyfish that causes organisms to fluoresce. This protein was the first fluorescent protein to be discovered and has been highly useful in a broad range of cell biology disciplines; because of it’s highly useful reporter genes and the ability to use multicolor protein tracking in living cells it has been useful in many scientific experiments such as E. Coli transformations. There have been many other fluorescent proteins that have been cloned from a wide variety of marine invertebrates, therefore making GFP not the only standing fluorescent protein present. (Shaner 2014). The scientific uses of GFP have included

-Plasmid is a small, circular, double-stranded DNA molecule that naturally exist in bacterial cells. The range of length are from hundreds base pairs to thousands base pairs. This is a great tool for clone, transfer, and manipulate genes.

It started out as one idea, and then developed into something more valuable as many different layers began to uncover. More and more was discovered, and so it began to spread worldwide, engrossing scientists with their unique abilities. Although, it’s important to take note of the fact that a large portion of this interest began with the discovery of a plasmid to act as a vector. A vector is, simply put, a vehicle for foreign DNA. Vectors insert this genetic material into its system, and relocate elsewhere in order for the DNA to replicate and spread throughout the new location. This information proved to be crucial, for this ‘vector-like’ ability allowed for scientists to mix and match different DNA strands in order to genetically modify certain organisms. In order for scientists to accomplish this task, they must first be able to insert DNA into the plasmid vectors themselves. To do so, a restriction enzyme is used, which, depending on what restriction enzyme is selected, will cut the DNA at certain points in the base pairing sequence. This cutting process is done to both the foreign material, as well as the plasmid vector being used for transport. Once completed, both parts will have split into two “sticky ends,” which are separated pieces of the base pairings that are ready to be connected to a matching pair of DNA. It’s important scientists use the same restriction enzyme for both, in order for the same base pairings to be cut from each, leaving sticky ends that can correspond to the opposite strand of the other organisms DNA. The base pairings of each are then lined up with one another in order to create a singular strands with the separate pieces taking on the role as one singular genetic strand. This is accomplished through the help of DNA ligase, which performs as a sort of glue that bonds corresponding nitrogen bases. Once the insertion into the plasmid is completed, it is then time to

The effect of CaMKII on synapse specificity is controlled by two factors: the rate of CaMKII’s inactivation and its mobility. Starting with the rate of inactivation of CaMKII, the fluorescence lifetime of the GFP, in other words, the activation of CaMKII, lasted only two minutes. This went along with the observation calcium levels fall back to initial concentrations within milliseconds. This does not allow calcium to diffuse into neighboring spines and activate CaMKII due to the short time limit. Because of the fast rate of inactivation of CaMKII, CaMKII is forced to stick to one dendritic spine. Hence why, the activation of CaMKII follows synaptic specificity. The second factor is mobility of the activation of CaMKII. Using fluorescence decay, it was concluded that CaMKII had really slow mobility of diffusion out of a dendritic spine. Because CaMKII can’t diffuse out of the spine fast enough, this also confines the activation of CaMKII to one

In the case of the “off” center response, the Rod is not exposed to light and therefore, Rhodopsin does not absorb the light photon and the GPCR does not activated Transducin. For this case, since Transducin is not activated, there becomes an increase in intracellular [cGMP]. The cGMP molecules are able to bind to the cGMP-gated sodium channels subsequently causing a net flux of sodium inside the photoreceptor. This net influx causes a graded action potential to fire in the photoreceptor. This action potential fires down the cell causing an increase in NT release into the synaptic cleft. The NT transmitter involved is Glutamate, once again. This photoreceptor cells makes a synapse with the “on” center bipolar cell and extends over to the “off”

The authors in this article are trying to better understand how information is processed in the brain, especially the diseased brain. In order to research this information there first needs to be a precise method of spatio-temporal recording of electrical activity in larger neuronal circuits and individual neurons. The authors hypothesized that fluorescent indicators would be useful tools in recording brain activity because they would allow monitoring of a genetically defined neuronal circuit and they would not require chemical access to visualize and provide optical recording of brain activity. Since the current techniques could not meet the authors’ requirements to study neuronal activity in vivo, they sought out to create a voltage

Rods are extremely sensitive to light and can react to a single photon. The light is absorbed by the pigment rhodopsin and the energy acquired results in decrease in cGMP which in turn causes the closing of Na+ channels. This hyperpolarizes the cell causing a release of synaptic vesicles.

The polymerase chain reaction or PCR for short can be used to create many copies of DNA. This allows the DNA to then be visualized using a dye like ethidium bromide after gel electrophoresis. The process has been refined over the years, however the basic steps are similar.

The phototransduction cascade converts light into electrical signals in the rod and cone cells, and in the photosensitive ganglion cells of the retina of the eye. This biological cycle converts photons into electrical signals in the retina. Rod photoreceptors in vertebrates are responsible for initiating vision by allowing light to isomerize 11-cis retinal chromophore of rhodopsin. This results in the rhodopsin to be excited and activates transducin to stimulate cCMP hydrolysis by the PDE, leading to less cGMP in the outer segment of photoreceptors. The decrease in cGMP closes channels, which haults the entry of sodium and calcium. This will lead to a hyperpolarized cell, as well as reducing the synaptic release of neurotransmitter glutamate.