A student is a lab isolated DNA from a transgenic mouse, a model used to study...

A student is a lab isolated DNA from a transgenic mouse, a model used to study heart failure and wanted to perform the following:

1- Amplify DNA
2- Clone the gene of interest
3- Sequence the sequence of the gene that causes heart failure
What does he need to do for each step

Expert Solution

1. Polymerase chain reaction or PCR is commonly employed to amplify the DNA. PCR helps in generating several copies of DNA sequence. For amplification using PCR, the following components are required - (1) DNA template (2) Taq polymerase (enzyme) (3) DNA primers (4) dNTPs (5) buffer solution (6) bi valent cations (magnesium or manganeese). The procedures include - Initialization, denaturation, annealing, extension, final elongation and final hold.

2. Molecular clonning of DNA includes production of identical copies of the gene or DNA of interest. A standard clonning protocol includes the following steps - (1) selection of the clonning vector and host organism (2) Vector DNA preparation (3) creating the typical recombinant DNA (4) Introducing recombinant DNA into host organism (5) Selecting the organism that contains recombinant DNA (6) Clone screening which conatins the DNA insert.

3. Sequencing includes the understanding of the exact nucleotide sequence of the DNA or gene of insert. Sequencing of a DNA may be done using sanger method or maxam -gilbert sequencing.

A typical sanger sequencing is based on the incorporation (selective) of chain terminating dNTPs with the help of DNA polymerase during the process of DNA replication.

The maxam and gilbert method employs the use of radioactive labelling at 5' end of the DNA and chemical treatments produces breaks at certain regions of nucleotide bases. The DNA fragments are seen with the help of electrophoresis and location of each nucleotide is estimated.

gladiator answered 1 year ago

Creating a transgenic organism A. is the result of deleting DNA from specific genes of an...

Creating a transgenic organism A. is the result of deleting DNA from specific genes of an organism. B. is a form of natural selection. C. is the result of adding DNA to an organism from another species. D. happens spontaneously. E. benefits all organisms. How are artificial selection and natural selection similar? A. Both require human intervention. B. Both work to create a new species. C. Both select for a specific trait(s) within a population. D. Both processes only affect...

An experimental mouse model of Leishmania infection was used by Nathan and Miller to investigate the...

An experimental mouse model of Leishmania infection was used by Nathan and Miller to investigate the relative contribution of reactive oxygen intermediates (ROIs) and reactive nitrogen intermediates (RNIs) in the control of leishmaniasis. i) How did the authors establish that RNIs but not ROIs are essential and sufficient to limit an infection? ii) How are these toxic intermediates generated by redox reactions and what mechanisms protect the parasite against their toxicity?

List 3 similarities in how DNA is isolated from bacteria and from yeast.

Isolated naked bacterial DNA from which proteins are removed is supercoiled. DNA in bacterial chromosomes is...

Isolated naked bacterial DNA from which proteins are removed is supercoiled. DNA in bacterial chromosomes is also supercoiled. When naked DNA is nicked, supercoiling is abolished. In contrast nickling chromosomal DNA does not abolish supercoiling. Why?

As a reminder, PCR is the technique used to replicate DNA in the lab. PCR is...

As a reminder, PCR is the technique used to replicate DNA in the lab. PCR is based on the way DNA is replicated in cells. During PCR template DNA plus other ingredients are placed in a test tube and the solution goes through repeated cycles of heating and cooling. The 'ingredients' of a basic PCR reaction are: DNA template, primer, nucleotides, DNA polymerase, and any salts required for polymerase function (buffer). No ligase is required in PCR but it is...

Working with neurons in the lab, you noticed that a batch of isolated neurons from Science...

Working with neurons in the lab, you noticed that a batch of isolated neurons from Science Express is not working appropriately. Your research advisor asks you to sort it out. Doing several experiments you noticed that most neurons either failed to start or failed to get back to the resting phase (state). You suspect that the special batch of neurons might carry a mutation and was mistakenly sent by this company. What kind of mutation do you suspect these neurons...

Lab Exercise 10 - ISOLATION OF DNA FROM PLANTS Introduction Deoxyribonucleic acid (DNA) is located in...

Lab Exercise 10 - ISOLATION OF DNA FROM PLANTS Introduction Deoxyribonucleic acid (DNA) is located in the nucleus of eukaryotic cells (animals, plants, fungi, and protists). DNA contains information to direct the cell in the manufacture of proteins. Proteins control development, organ function, metabolism, enzymatic reactions, photosynthesis, muscle action, brain activity, and many other cellular processes. DNA is often referred to as the “blueprint for life”. DNA is a polymer composed of the nucleotide bases guanine (G), adenine (A), thymine...

. You have extracted extrachromosomal DNA and genomic DNA from E. coli in the lab. What...

. You have extracted extrachromosomal DNA and genomic DNA from E. coli in the lab. What is the differentiating steps during the DNA isolation so that your target DNA is not contaminated with the other unwanted DNA (i.e. you are targeting extrachromosomal DNA, so you don’t want the genomic DNA, and vice-versa)? Why and how that step can differentiate between these groups of DNA

You have previously used radiation therapy to induce cell apoptosis in a mouse tumor model in...

You have previously used radiation therapy to induce cell apoptosis in a mouse tumor model in which FGF is over-expressed. Recently, you notice that this therapy is no longer effective and the tumor continued to grow rapidly. Genomic sequencing revealed that those tumor cells have acquired a new loss-of-function mutation in the tumor-suppressor gene Pten, whose protein product normally acts as a phosphatase to dephosphorylate PIP3 into PIP2. Based on this information, you have an idea of why the tumor...

You have constructed four different libraries: a genomic library made from DNA isolated from human brain...

You have constructed four different libraries: a genomic library made from DNA isolated from human brain tissue, a genomic library made from DNA isolated from human muscle tissue, a human brain cDNA library, and a human muscle cDNA library. You wish to clone the human gene encoding myostatin, which is expressed only in muscle cells. A. Assuming the human genome is 3×10^9 bp and that the average insert size in the genomic libraries is 100 kb, how frequently will a...

Question