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What is the purpose of ethidium bromide?
Ethidium bromide is an intercalating agent that attaches itself between the helix of a DNA. Because the ethidium molecule lights up when illuminated by an ultraviolet light, it is used often in biochemistry laboratories so that fragment of DNA that has been separated by gel can be visualized.
What is the role of Cscl and ethidium bromide in plasmid purification?
The CsCl forms a gradient and the molecules migrate according to their density until they float at their individual isopycnic points (the point in the gradient that equals the buoyant density of the molecule). However, plasmid DNA and contaminating chromosome have about the same density and cannot be separated easily. This is rectified, however, by the addition of ethidium bromide. Density is a function of AT/GC ratio, but it is also a function of conformation. For supercoiled DNA, there is more DNA per unit volume than for relaxed DNA. Intercalation of ethidium bromide into DNA causes the helix to unwind (negative supercoiling) and become more relaxed. However, negative supercoiling only relaxes the DNA to a point. Further unwinding induces supercoiling in the opposite direction. When the DNA is circular and the ends are connected, the plasmid "kinks up" into a very tight knot. Thus, ethidium bromide causes the plasmid density to be increased.
If ethidium bromide have a higher melting point than 120 C 260 - 262 C could it be autoclaved with the medium you are using without being degarded or its?
Although theoretically the menting point of entidium bromide is high, it is generally added to media/gel after boiling and media or gel and the media or gel is still in the molten liquid state
What are the two products when liquid bromine is added to calcium iodide?
When liquid bromine is added to calcium iodide, the reaction results in the formation of calcium bromide and iodine. Calcium bromide is a salt composed of calcium cations (Ca2+) and bromide anions (Br-), while iodine is a diatomic molecule composed of two iodine atoms (I2). This reaction is a double displacement reaction where the bromine displaces the iodine in calcium iodide to form the two new products.
What happens to bromine when bromine atom gains an electron and forms the bromide ion Br -?
When a bromine atom gains an electron to form the bromide ion (Br-), it becomes negatively charged. The added electron gives the ion a full outer shell of electrons, making it stable. The bromide ion will be attracted to positively charged ions or molecules.
What is the purpose of ethidium bromide?
Ethidium bromide is an intercalating agent that attaches itself between the helix of a DNA. Because the ethidium molecule lights up when illuminated by an ultraviolet light, it is used often in biochemistry laboratories so that fragment of DNA that has been separated by gel can be visualized.
How does ethidium bromide interact with double stranded DNA Does it increase or remove supercoiling?
when ethidium ion intercalates between two dna base pairs in a circular dna it causes the dna to unwind by 26 degrees, thereby decreasing twist and increasing writhe. in a circular dna which is negatively supercoiled, if ethidium is added it will become relaxed and if more ethidium is added dna becomes positively supercoiled
Why glycerol used in agarose el electrophoresis?
Glycerol is added to the loading buffer in agarose gel electrophoresis to make the sample denser than the surrounding buffer. This helps the sample sink into the well and prevents it from mixing with the buffer during loading. Additionally, glycerol increases the density of the sample and helps it sink into the gel.
What is the buffer solution and why is it added to the agarose?
You use a buffer when making agarose gels so that when the gel is used for electrophoresis, the gel is able to conduct electricity. The buffer contains ions from the buffer salts that will facilitate conduction. that was good
What is used to make the DNA visible on the gel?
YES!! You can use a simple Agarose gel to separate to view the DNA on electrophoresis. Use 0.8 - 1% gel for 5-10kbp , 2% for 0.2 - 1kbp. If the fragments are really tiny, use an Acrylamide gel (vertical gel) to electrophorese and they will show right out. This is to offset the instability of high concentration gels.
What is the role of Cscl and ethidium bromide in plasmid purification?
The CsCl forms a gradient and the molecules migrate according to their density until they float at their individual isopycnic points (the point in the gradient that equals the buoyant density of the molecule). However, plasmid DNA and contaminating chromosome have about the same density and cannot be separated easily. This is rectified, however, by the addition of ethidium bromide. Density is a function of AT/GC ratio, but it is also a function of conformation. For supercoiled DNA, there is more DNA per unit volume than for relaxed DNA. Intercalation of ethidium bromide into DNA causes the helix to unwind (negative supercoiling) and become more relaxed. However, negative supercoiling only relaxes the DNA to a point. Further unwinding induces supercoiling in the opposite direction. When the DNA is circular and the ends are connected, the plasmid "kinks up" into a very tight knot. Thus, ethidium bromide causes the plasmid density to be increased.
If ethidium bromide have a higher melting point than 120 C 260 - 262 C could it be autoclaved with the medium you are using without being degarded or its?
Although theoretically the menting point of entidium bromide is high, it is generally added to media/gel after boiling and media or gel and the media or gel is still in the molten liquid state
What happens when silver nitrate is added to bromide?
When silver nitrate is added to a solution containing bromide ions, a white precipitate of silver bromide will form. This is due to a chemical reaction between the silver nitrate and bromide ions, resulting in the insoluble silver bromide precipitate.
What do you see when chlorine is added to a solution of bromide ions?
When chlorine is added to a solution of bromide ions, a red-brown color may appear due to the formation of bromine. This is because chlorine can oxidize bromide ions to bromine.
What is the main difference between protein electrophoresis and nucleic acid electrophoresis?
There are many similarities and differences between protein and DNA electrophoresis.Similarities:PAGE protein and DNA electrophoresis both cause separation by size, creating bands that are viewed by the scientist or a machine. The smallest segments more the fastest due to less friction with the surface of their medium or equipment.The movement of charges through the medium is what causes the DNA or proteins to move. Electrons move from the negative to positive end of the gel or capillary tube.Differences:In PAGE protein electrophoresis, a polyacrylamide gel is used to prevent convection from altering the movement of the proteins. If the proteins are charged, and there is a worry that the charge will affect the mobility of the protein segments, 1% SDS can be added to get rid of the mass/charge issue. This way, only the mass of the segment determines how far it moves. In DNA capillary electrophoresis, the size of the capillary is so small that it does not have room for convection to occur (it is only 20-50 microns wide). Thus, there is no medium in the capillary but DNA itself.In protein electrophoresis, the proteins are often dyed so their movement can be viewed with the naked eye, or a machine. With DNA capillary electrophoresis, DNA strands are made through DNA replication with dNTPs that are fluorescently labeled for the different nucleotides. Each base is labeled a different color. A fine laser lights up the DNA strand in the capillary tube and reads what color fluoresces. This is how the nucleotide is identified.Protein PAGE electrophoresis is used to determine the purity of a protein sample. It can also be used to see how large the chains are that make up a multi-chain protein if a denaturing agent is added. DNA electrophoresis is used to get the order of nucleotides in a DNA sequence. It is done by chopping the DNA sequence into many smaller bits and sequencing them, then putting them back together by lining them up according to sequence overlaps. This is called the "shotgun" method. Protein electrophoresis can figure out the order of about 15-20 amino acids by a similar method, but DNA electrophoresis can get up to 1000 nucleotides (~300 amino acids). DNA electrophoresis is limited by the low probability that the DNA sequence would be cut into a segment greater than 1000 nucleotides.
What happens to Bromide ions when added to silver nitrate?
When bromide ions are added to silver nitrate, a white precipitate of silver bromide is formed. This is because silver ions react with bromide ions to form the insoluble silver bromide precipitate. This reaction is commonly used in qualitative analysis to detect the presence of bromide ions.
What is th blue dye added to the sample before the electrophoresis is performed?
In biochemistry labs, the traditional answer for a protein gel (polacrylamide gel electrophoresis) is bromphenol blue. For a DNA gel (agarose gel electrophoresis), traditionally the same dark blue dye bromphenol blue was combined with the lighter, slower migrating blue dye xylene cyanol. Oftentimes nowwe only use the bromphenol blue, or even substitute for it with Orange G, which is a UV-transparent dye that more easily enables the visualization of smaller molecular weight nucleic acids that migrate in the same region.
