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1) Briefly outline the steps of DNA extraction as carried out in a lab for the purposes of sequencing, with reference to a published protocol from a company that supplies regents and kits for DNA extraction (e.g Qiagen, ThermoFischer, Invitrogen, or similar). Please Include the weblink for the published protocol you used as a reference. (Word limits: 300)
2) Why was there a greater A260 absorbance reading for your DNA sample that was incubated at higher temperatures. (Word Limits: 100)
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- In a protocol for DNA sample preparation for agarose gel electrophoresis, what volume of 4X loading buffer must be added to 21 micro L of DNA to obtain a 1X buffer solution?Whether done manually or automated, DNA sequencing gels are always made of polyacrylamide rather than agarose. Why can't agarose be used for a sequencing gel, as it is for other DNA gel electrophoresis?Briefly outline the steps of DNA extraction as carried out in a lab for the purposes of sequencing, with reference to a published protocol from a company that supplies regents and kits for DNA extraction (e.g Qiagen, ThermoFischer, Invitrogen, or similar). You should note the refinements compared to the protocol used in your practical session.
- You have a very concentrated sample of DNA which you need to dilute for sequencing 1:6000 with a final volume of 10 μl. How would you do this in four steps, using a serial dilution? Do not pipet less than 1 ul and do not make more than 10 ul per dilution. Describe how to do each dilution using the format suggested below.To carry out sequencing, you need to include 600 ng of DNA and a total of 6.4 pmol of sequencing primer. If your concentration of DNA (determined by A260 reading on a Nanodrop Spectrophotometer) is 89.0 ng/μL, how many μL of this sample do you need for a total of 600 ng?A 22-kb piece of DNA has the following restriction sites:A batch of this DNA is first fully digested by HpaI alone, then another batch is fully digested by HindIII alone, and finally, a third batch is fully digested by both HpaI and HindIII together. The fragments resulting from each of the three digestions are placed in separate wells of an agarose gel, separated by gel electrophoresis, and stained by ethidium bromide. Draw the bands as they would appear on the gel.
- What is the useful of the Development of highly sensitive and low-cost DNA agarose gel electrophoresis detection systems, and evaluation of non-mutagenic and loading dye-type DNA-staining reagents in modern medicine today?Consider the DNA segment with a sequence: 3'-TACGGTACGGGATTG-5'. If the given DNA sample was subjected to pyrosequencing, sketch the expected profile of the sequencing output. Assume that the sequential flooding of nucleotides follows the order G-C-A-T.You have isolated genomic DNA and plasmid DNA from a bacterial culture. Please answer the following questions: 1. What solution did you use to elute DNA from the silica resin column? 2. What is the function of the chaotropic salt in DNA isolation? 3. Why do we add the chaotropic salt for genomic DNA before lysis and for plasmid isolation we add the salts after the lysis step?
- what type of gel(in terms of material) can be more suitable for the electrophoresis of 500-1000bp DNA fragment and why?As a technique for detecting genetic variations, RFLP has substantial drawbacks. Name one such drawback, explain why it is unique to RFLP analysis with specific reference to the technique, and discuss why DNA sequencing overcomes this drawback. Please leave the link for any sources used. Thanks!What would spread (fragmented) bands in the electrophoresis gel tell you about the quality of your DNA? Explain briefly, yet clearly