they were labeled, and where they will be stored. if someone else needed to pick
ID: 3164346 • Letter: T
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they were labeled, and where they will be stored. if someone else needed to pick up where you left off for the day, woukd they know what you did? Would they know where 13. Plates will be incubated at 30°C for approxi mately 18-24 hrs. Day 2: Write up and Data Analysis 14. Describe the general appearance of each plate after 24 hours of incubation at 30 C. Also, indicate whether any of the colonies are blue and whether any of them glow in the dark. Be specific about which plate(s) have which type of colonies. 15. Write a conclusion to your experiment. Were the results as expected Why? What does this mean? 16. Answer the post lab questions Post-Laboratory Questions: 1. If the e bacteria were bioluminescent after 24 hours of incubation at 30'C what did that result tell us? sing the puc19 vector illustrated in Figure 7.2, if you ligated in rDNA and then transformed your your plate and find about 100 colonies, 50 are blue and 50 are white. Could you explain this result? You have created a rDNA plasmid using the pGEM*T Easy vector system by Promega (a map of us vector can be accessed at the Promega website (Promega.com). Search by vector name. Wha w int o competent cells as described in this lab and after 24 hours of incubation, you return to 3. results would you expect on the following plates: . X-Gal; IPTG; 40 ug/ml ampicillin · X-Gal; I PTG, 40 ?/rnl hygromycin .. ,..: : 40pg/ml ampicillin 4. After combining the cell extracts with a strain of non-bioluminescent E. coli, a single bacterium ing colonies of millions of bacteria. How can they all be glowing if we only transformed a single bacterium? is transformed; however, when we analyze the plates for bioluminescence we are looking at glow-Explanation / Answer
Answer:
1) The bioluminiscence gene is a virulent gene which produces proteins which luminates under stress conditions. The bacteria which show bioluminiscence property will show it only after incubation for 24 hours at 30 degree celcius. This means that the bioluminiscence property is shown before even reaching the stationary stage., i.e it is showing it in the growing stage.
2) 50 showing white and 50 showing blue means the effeciency of transformation was 50%. The colonies showing white were transformed successfully and the colonies showing blue were not transformed successfully.
3) The p-GEM vector contains an amp resistance gene and the MCS is at LacZ site. If the rDNA is ligated at lacz site, then:
X-Gal, IPTG; 40ug/ml ampicillin, it will show white colonies which will have successfull transformation.
4) The cell extracts, i.e the cell lysates should contain the bioluminiscent gene which was transformed into the non-bioluminiscent E.coli. Now we are seeing that many more bacteria of the same glowing, this is because of conjugation of the gene between the individuals of same species. The gene is transported at a very fast rate to almost a lot of individual bacteria so that millions of bacteria are glowing at the same time.
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