We have mRNA prepared from human cells but PCR needs DNA. What should we do?
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- The temperature at which the primers and target DNA hybridize may be changed to influence the stringency of PCR amplification. What effect will changing the hybridization temperature have on the amplification? Let's say you have a certain yeast gene A and want to check whether it has a human equivalent. How might managing the hybridization's rigor benefit you?You want to amplify the following sequence of DNA from a gene of interest that you are studying. To do this, you will use PCR. Design a forward and reverse primer. Show where the primers anneal to the template sequence. Template DNA: 5’ ATGACGGAATATAAGCTGGTGGTGGTGG---GGCTGCATGAGCTGCAAGTGTGTGCTCTCCTAA 3’ 3’ TACTGCCTTATATTCGACCACCACCACC---CCGACGTACTCGACGTTCACACACGAGAGGATT 5’ (PLEASE EXPLAIN STEP - BY STEP) Answer : Foward primer 5": Reverse primer 3"EcoRI recognizes GA-A-T-T-C sequence and cleave/ cut between G and A. How will the DNA fragments look like if EcoRI is used for the DNA below? How many fragments are produced? -'3 5'-AAAGATTTGAATTTCGAATTCAATTTAAGAATTCCCTTAGAATTTCC
- What advantages do cDNA libraries provide over genomic DNA libraries? Describe cloning applications where the use of a genomic library is necessary to provide information that a cDNA library cannot.You obtained the sequence of the frog gene X you amplified in Question #16 through a process called automated sequencing. In automated sequencing, you are given a printout of the sense strand of your DNA. The printout is shown below. The first thing you need to do is use the correct reading frame. Having done this, the next thing to do is to write out the mRNA sequence using this sense strand reading frame. The last thing to do is to translate the sequence. Do these steps in the space below. The reading frame DNA sequence is: 2.The mRNA sequence is: The polypeptide sequence is:Q4) What’s the evolutionary purpose of restriction enzymes? Why is the bacterial DNA not harmed in this process? Q5) Restriction enzymes look for a very particular KIND OF sequence. What is that called. Give me an example of one.
- automated sequencing is used are given a printout of the sense strand of your DNA. The printout is linked. The first thing you need to do is use the correct reading frame. Having done this, the next thing to do is to write out the mRNA sequence using this sense strand reading frame. The last thing to do is to translate the sequence The reading frame DNA sequence is: The mRNA sequence is: The polypeptide sequence is:CRISPR/Cas9 can be used in genome editing. Among the following statements, which one is correct? The CRISPR and the Cas9 parts both recognise the DNA target sequence and then recruit an endonuclease for cutting it The Cas9 part recognises the DNA target sequence and the CRISPR part cuts it The CRISPR and the Cas9 parts both recognise and cut the DNA target sequence. The CRISPR part recognises the DNA target sequence and Cas9 cuts it.You’re working in a research lab, and your current task is to clone the gene that codes for tyrosinase from potatoes. You grind up some potato, extracts the DNA from it and digests the DNA with two different restriction enzymes (separately, not together): EcoRI and BamHI. You then obtain the cloning vector, pUC19, and digest it with the same two enzymes. You then run a gel which is shown to the right. You mix the potato DNA (digested with the enzyme you specified in part B) with cloning vector DNA (digested with the same enzyme). You then add the mixture to E. coli cells and carry out a transformation procedure so that the cells can each update a plasmid. You then plate the cells on a plate containing antibiotic. What antibiotic would be in the plate? Why would there be antibiotic in the plate? Be specific! Unfortunately, you don’t get a single bacterial colony to grow on the plate. Not even one! You review your procedure and realize that when mixing the digested potato DNA…
- To amplify a section of DNA using the polymerase chain reaction (PCR), all you need to load into the tube is 1) a buffer solution, 2) the DNA you want to amplify, 3) some DNA nucleotides, 4) a polymerase (like Taq polymerase), and O an RNA polymerase a set of forward and reverse primers some phospholipids for a cell membrane some ribosomesCloning Genes Is a Multistep Process The following DNA sequence contains a six-base sequence that is a recognition and cutting site for a restriction enzyme. What is this sequence? Which enzyme will cut this sequence? (See Figure 13.5 for help.) 5 CCGAGGAAGCTTAC 3 3 GGCTCCTTCGAATG 5Which of the following best describes the process of DNA sequencing? a. DNA is separated on a gel, and the different bands are labeled with fluorescent nucleotides and scanned with a laser. b. A laser is used to fluorescently label the nucleotides present within the DNA, the DNA is run on a gel, and then the DNA is broken into fragments. c. Nucleotides are scanned with a laser and incorporated into the DNA that has been separated on a gel, and then the DNA is amplified with PCR. d. Fragments of DNA are produced in a reaction that labels them with any of four different fluorescent dyes, and the fragments then are run on a gel and scanned with a laser. e. DNA is broken down into its constituent nucleotides, and the nucleotides are then run on a gel and purified with a laser.
