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Why does DNA ligase require ATP or NAD plus to connect two Okazaki fragments?
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What is the result when DNA ligase has completed its job?
Two new strands of DNA. <--- Gradpoint/NovaNet
What is the methodology for producing recombinant DNA to be used in gene cloning?
1. A vector such as plasmid is needed along with a host cell. Restriction enzymes and DNA ligase are enzymes that are used to introduce foreign DNA into a vector.
What is the process of making a genetically modified organism in a lab?
Contrary to the common understanding of the word, the process which we use to create transgenic organisms is called "cloning". This doesn't refer to copying an organism as often believed. Cloning as scientists mean it refers to creating DNA which does not naturally occur in any organisms. This is also called transgenic DNA.To make a genetically modified organism, we need:transgenic DNA - made by cloningan organismThe transgenic DNA of course needs to be made for a specific purpose. Depending on how big it needs to be, it can be made in different ways. Very small stretches of DNA can be synthesised chemically. Larger bits of DNA are usually made by extracting a cell's DNA and then using a technique called polymerase chain reaction (PCR) to take only one particular bit of the DNA out. We then use restriction enzymes and ligase to cut and paste different bits of DNA together. Usually, the end product will be a plasmid.The finished plasmid (transgenic DNA) then needs to be introduced into the target organism - and there are different ways of doing it for every type of organisms. Some bacteria for example will simply take up plasmids and integrate them into their own DNA without a complicated process around it being necessary. For bigger organisms like animals, it's very difficult - usually impossible - to modify an already living organism. For these, the only option is usually to take an oocyte (egg cell, e.g. a flower seed) and introduce the plasmid into it. If the egg survives this procedure and develops (which is very difficult to achieve), the resulting organism will be transgenic, or genetically modified.
What are the steps of using recombinant DNA technology?
1 Isolate DNA 2 Cut DNA with a restriction enzyme 3 Mix the DNA's and join then together by using DNA ligase 4 Insert the recombinant plasmid into a host bacterium 5 Allow the bacterium to reproduce
What is the role of a plasmid in genetic engineering?
A plasmid vector available today is made with a specific host in mind. For example, if you decide to express a gene in a bacteria, there will be plasmids available with features that suit the particular organism that you wish to transform and they will be different from plasmids used to transfect for example, yeast. However, generally, a plasmid will have at the very least an origin of replication recognizable by the desired organism, a promoter upstream of the multiple cloning site that is recognizable by the organisms, and a selection marker such as an antibiotic resistance gene.The process of expressing a gene from one organism in another host via plasmid vectors begin with the isolation of the gene from the original organism. For the sake of this example, suppose the insulin gene in humans is the gene of interest. First, beta cells from the Islets of Langerhans will have to be lysed and total RNA will be isolated from the cell. Because DNA is filled with many introns that are hard to get rid of, gene isolation from higher eukaryotes almost always start from the mRNA stage because the introns were already sliced out in mRNA processing. The RNA will be then subjected to reverse-transcriptase polymerase chain reaction with primers specific for the insulin gene. The insulin gene will subsequently be selectively amplified and the reaction mixture can then be purified to contain only cDNA of the insulin gene.With the purified cDNA, a process called molecular cloning is used to get the gene into the plasmid. The plasmid and the gene are both cut with compatible restriction enzymes. The cuts on the plasmid has to be in the multiple cloning site the the cuts on the gene has to be outside of the open reading frame for the cloning to produce an effective vector. (Review molecular biology for the necessity of promoters and an intact open reading frame) The cut plasmid and gene fragments are then placed together and ligated. The ligated product should theoretically now contain the gene inside the multiple cloning site directly following the promoter. The promoter may express the gene constitutively or it may be inducible, requiring certain conditions to be met before it is turned on.The plasmid with the cloned insulin gene can now be transformed into competent bacteria hosts (or yeast if desired, however it will not be as efficient). Competence describes the ability of bacteria to take up DNA from its surroundings. The most commonly used host, E. coli, are artificially made to be competent by treatment with a high concentration calcium solution in a cold environment, while others, such as B. subtilis, are naturally competent. All bacteria can be made competent with electroporation but E. Coli is most often used because of its easily satisfied nutrient requirements and very short generation time. The plasmid and competent E. coli is placed together in a cold environment to initiate the uptake of the plasmid into E. coli cells. The mixture is then heat shocked and bacterial growth medium with the necessary selection agent is added to start the incubation process. If the selection marker on the plasmid is an antibiotic resistance gene, for example ampicillin resistance, a medium with ampicillin will be used to incubate the bacterial culture because only the cells that contain the plasmid will be resistant to the antibiotic while cells that have failed to take up the plasmid will die. The cells can then be incubated for as long as needed and split into different cultures if needed because they now contain the plasmid and will express the gene carried on the plasmid.A plasmid can be considered as a suitable vehicle for cloning, because 1. It can be isolated from the cells2. It possesses a single restriction site for one or more restriction enzymes.3.Insertion of a linear molecule at one of these sites does not alter its replication properties.4.Reinsertion of these vectors to the host cell can identified and selectable.5.They do not occur free in nature and found in bacterial cells.Ex: for plasmid cloning vectors are pBr322,pACYC18,pUC,pUN121.
What DNA replication enzyme attaches okazaki fragments as a continuous strand of DNA?
Ligase joins okazaki fragments to each other to form a continuous strand of DNA
Name the enzyme joining Okazaki fragments?
DNA ligase.
What is DNA ligase functions?
Ligase joins together the Okazaki fragments into a continuous strand of DNA.
What is the function of DNA ligase?
Ligase joins together the Okazaki fragments into a continuous strand of DNA.
What is role of the DNA ligase in DNA replication?
It joins Okazaki fragments
What stand of DNA replicated in okasaki fragments?
The lagging strand of DNA contains okazaki fragments, which are later joined by DNA ligase
The enzyme that connects the fragments ( Okazaki fragments) are called?
DNA ligase is the enzyme responsible for connecting the Okazaki fragments on the lagging strand during DNA replication. It catalyzes the formation of phosphodiester bonds to join the individual nucleotides together.
What are okazaki fragments used to elongate?
Okazaki fragments are used to elongate the lagging strand. These fragments are used as primers for RNA polymerase to fill up the gaps in the newly formed complimentary DNA on the lagging strand. DNA ligase then seals up the gaps.
What is the function of DNA ligase in DNA replication?
it ties the chunks of dna together
During DNA replication the short sections of new DNA known as okazaki fragments which are eventually linked together by ligase?
lagging
What is the difference between ligase and lyase?
ligase mean to join..eg DNA ligase is used to join okazaki fragments ..&lyase mens to break down..lysozyme in our saliva & tears is lyase which disssolves bacterial cell wall
What enzyme glues the DNA together after replication?
Question is unclear . If you mean pieces of DNA , thae enzyme is DNA ligase which joins pieces of DNA called Okazaki fragments together .
