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- at Arrange Window Help 4) 57% D Tue 4 May Tools Slide Show 2 ARMS PCR nations Slide Show Review View A A =、=<|EE|三v| v v A Sha v 20 Quick Styles D Sha Text Arrange Convert to SmartArt Picture Shapes ab x x. AV v Aa v D. Av A、申。 Box Briefly outline how MLPA probe-sets can simultaneously detect multiple targets using a single set of PCR primers. E Notes Comments MAY 4 Pr w MacBook AirKha Vu Danels Include: 8DX : Safehy Jor bromie tnto lab repart! Name Section date sheet MAPPING PRACTICE #1 Below is a restriction map for the plasmid PGEN 101 (total length = 20 Kb). Using this map as a guide, give the number of restriction fraqments along with their associated lengths that would result from digesting PGEN 101 with the restriction enzymes EcoRI, BamHI and a combination of ECORI and BamHI. BamHI 3.2 Kb 1.7 Kb EcoRI BamHI PGEN 101 8.7 Kb 5.5 Kb .9 Kb EcoRI ECORI DIGESTION PERFORMED SIZES OF FRAGMENTS OBTAINED 10.4 kb , 0.9kb, 8.7 Kb EcoRI 3.2 Kb, 16. 8kb BamHI EcoRI + BamHI5' 3' O 60°C 5' O 95°C Primer 1 O 75°C ORF For your PCR reaction above, the primer set has a much higher AT (low GC) content than what would be considered normal. Due to this difference in AT content, the optimal annealing temperature for the primers is going to have to be altered from the "normal" temperature cycle: 95°C 60°C ⇒ 75°C Which of the temperatures will have to be altered to accommodate this difference the primer set? Primer 25¹ 3' 5' annealing with
- PCR Analysis la: The 2x PCR Master mix has four main components. Two of these are PCR Buffer and MgCl2. Based on your knowledge of DNA replication and PCR, what are the other two components and what are their functions? 1b: Explain what would happen if the PCR reaction did not contain Taq polymerase? Assuming all other required PCR components are present, would any part of the PCR reaction occur? Explain your reasoning. 1c: Why does a PCR reaction require a primer? Would you expect this primer to be composed of DNA or RNA? Explain your reasoning. 1d: Should PCR primers be complementary to each other? Explain your reasoning le: As you know, experiments include controls. In this lab, a negative control was included along with your experimental samples. Which components do you think the negative control would contain? Hint: Think about which component would be left out of the negative control relative to the experimental samples. Explain why. 1f: What would the PCR product be for the…Analyzing Cloned Sequences A base change (A to T) is the mutational event that created the mutant sickle cell anemia allele of beta globin. This mutation destroys an MstII restriction site normally present in the beta globin gene. This difference between the normal allele and the mutant allele can be detected with Southern blotting. Using a labeled beta globin gene as a probe, what differences would you expect to see for a Southern blot of the normal beta globin gene and the mutant sickle cell gene?1. The final amount of each primer required in a PCR reaction is 0.025 nanomol. If the total volume of the PCR reaction is equal to 100 µl and the stock concentration of each primer is equal to 10 µM. Calculate the volume of stock primer that needs to be added in order to ensure a primer amount of 0.025 nanomol. 2. A 250 ml phosphate buffer solution contains 204.12 µg KH2PO4 and 87.09 µg K2HPO4, respectively. Determine the molarities in micromolar (µM) of the abovementioned compounds in the phosphate buffer solution. [Mw KH2PO4 = 136.08 g/mol and Mw K2HPO4 = 174.18 g/mol] 3. You are required to use the pET 28b expression vector in a cloning experiment. If the vector is 5.368 kilobasepairs in length and the mass of the DNA that you have added represent 8.5 x 109 molecules. Determine the mass of vector DNA that you have added to your reaction? [1basepair ~ 660 Da or g/mol, Avogadro’s constant = 6.023 x 1023 molecules/mol]
- A cloning vector map is shown below. EcoRI Bam Ban Hind P-galactosidase Amp Bam Bam EcoRI Ori C Which restriction site is best for inserting a DNA fragment for selection of chimeric plasmid containing colonies? 1) They're all equally good. 2) Hindll 3) EcoRI 4) BamHIPCHEM4321. An agarose gel electrophoresis pattern of the plasmid PSPM4321 digestion (restriction) is shown below. Draw a restriction map of a plasmid with the appropriate restriction sites based on the data given below. Hindlll Hindll BamHI +BamHI Figure 1: 1% agarose gel electrophoresis of pCHEM4321 40 24 16 12 12 8 4 4 + |1. Briefly explain why the total size of the pMBBS plasmid in the Restriction Enzymes practical is 3000bp (base pairs) 2. Briefly explain why the cut sites on the pMBBS plasmid in each Restriction Enzymes (EcoRI, BamHI, and XhoI) just 1 each 3. Briefly explain what led to the 5 fragments formed which linked to the 500bp, 1000bp, 1500bp, 2000bp, 2500bp sizes
- Select all that are true about DNA gel electrophoresis O This technique can be used to separate DNA based on the size of the fragment O Since DNA is positively charged it will travel to the negative electrode O After a gel is run, the larger fragments end up being further away from the well The rate at which DNA fragments travel is logarithmically and is inversely proportional to its size7 of 11 Identify the incorrect statements O Each primer should have 40-60% GC content The annealing temperatures of the two primers should be within 5°C of each other The recommended final concentration of each DNTP is 2 M The pH of PCR buffer is usually between 8.0 and 9.5 V A pair of primers should have complementarity at their 3' endsOO HUAWEI Nova 2 Plus DUAL CAMERA estion To study the function of any gene of interest you would perform the loss and gain of function approaches by either deleting or re-expressing the gene of interest, which of the following can be used to determine and quantify the activity of the gene? red Oa. Microscopy d out of O b. Western blotting O C. PCR/OLA on O d. Gene knockdown O e. DNA hybridization stion Which one of the following is NOT correct regarding Bacterial Biosensors? O a. produced by all Pseudomonas species ed O b. Encoded by LacZ gene out of O c. Encoded by Lux genes O d. Chemical compounds used for the quantitative assessment of water pollution O e. In the presence of pollutants the bioluminescent decreases. stion CD4-Pseudomonas Exotoxin Fusion Protein is another biotechnology strategy for HIV therapeutic setting. This construct will target and kill: Select one: d O a. Any lymphocyte put of O b. HIV (virus only) O C. Non infected TH O d. Infected TH cells O e. Infected TH…