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Which of the following is a reason that cDNA clones of eukaryotic genes are capable of being expressed in prokaryotic cells?
I. cDNA clones do not contain any of the introns present in the genomic DNA
II. Prokaryotes have the basic translational machinery needed to express these genes
III. Prokaryotes are capable of making similar post-transcriptional modifications as eukaryotes
cDNA clones are incredibly powerful tools that can be used to express eukaryotic proteins in prokaryotic cells. One of the primary reasons that this is possible is because both eukaryotes and prokaryotes contain very similar systems of translating mRNA transcripts. Another reason is because the cDNA clone does not contain any of the information that would normally be spliced (introns). Prokaryotes do not make the same post-transcriptional modifications that eukaryotic cells do, and must have introns removed by external means in order to create viable protein products.
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Which of the following DNA sequences could most likely be cleaved by an endonuclease?
Endonucleases cut DNA at restriction sites. These restriction sites are palindromic, meaning that they read the same forward as they do backwards.
The sequence 5'-CCTAGG-3' will have the complementary strand 3'-GGATCC-5'. Since this sequence is read the same forward as it is backwards, it is a palindromic sequence that can be potentially cleaved by an endonuclease.
5'-CCT|AGG-3'
3'-GGA|TCC-5'
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When performing recombinant DNA techniques, it is often necessary to cut desired DNA sequences with restriction enzymes. Where are these enzymes typically isolated from?
The correct answer is bacteria. Both bacteria and archae utilize restriction enzymes as a defense mechanism to cut foreign intracellular DNA (specifically DNA of bacteriophage). Restriction enzymes are named based on the organism in which they were discovered - for example EcoRI is named after E.coli, and was the first restriction enzyme, found in the R strain.
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After a double digest with EcoRI and HindIII, a gel electrophoresis shows that you have several restriction fragment bands. You see bands of lengths 3kb, 4kb and 5kb. From previous tests, you know that there are two EcoRI restriction sites and two HindIII restriction sites in the DNA fragment you are studying. How many restriction fragments are really in your reaction?
The correct answer is five. Although you can only see three bands in your gel, a DNA fragment with four different restriction sites will be cut into five distinct pieces (if you're having trouble picturing this, drawing it out helps). In this case, some of the bands must be matching sizes, so they do not appear to be separate on your gel. For instance, there may be two 3kb bands, two 4kb bands, and one 5kb band.
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