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Quiz #4 key
BE 109 SP 03 MON/WED
4/14/03
Name_____________________
Write your name on this quiz and then answer the following questions in three sentences
or LESS. You have fifteen minutes to answer ten questions.
1. With microarrays we can measure ___________ levels, although in some cases we
might rather measure ____________ levels in our cells, because this would give us more
direct information about a cell’s functional state. Hint: think about the role of each type
of molecule in the central dogma of biology.
Gene expression (mRNA) …protein
2. What types of genes will be on your microarrays this week? Hint: what was the paper
handed out in lecture last time about?
Human DNA repair genes.
3. In microarray experiments, what are the probes and what are the targets?
The probes are oligonucleotides that we have spotted onto the array – each spot
contains olignucleotides for a specific gene. The targets are labeled cDNA from
our cell samples which can hybridize to matching probe spots.
4. If we are not careful, our RNA samples may be degraded. Why? What can we do to
prevent this?
RNAses, which degrade RNA, are everywhere. We prevent this by spraying
equipment, benchtops and gloves with RNAse away, and by using certified
RNAse free pipette tips and eppendorf tubes.
5. Why do you need to shield your reactions from light?
Both Cy3 and Cy5 are light sensitive and easily degraded. Prolonged light
exposure causes photobleaching.
6. What is reverse transcription?
RNA -> DNA.
7. Two control chips will be run between the two classes. What do they control for?
The untreated vs. oligofectamine-treated chip controls for oligofectamine effects.
The oligofectamine-treated vs. oligofectamine-treated chip controls for cDNA
synthesis, labeling, and microarray hybridization effects.
8. What is a major difference between Cy3 and Cy5 that will affect how we treat the
results of our experiments?
Cy5 is a bigger molecule and thus does not get integrated as well into the cDNA.
9. How do you control for that (last question)?
We spike equal amounts of an internal control (alien mRNA) into each of our two
samples before labeling. This allows to normalize our data properly.
10. What is the purpose of having more than one spot on our array for each gene?
This is another control – controls for printing/spotting and hybridization. If the
four duplicates show reproducibility we can be more confident in our results.