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Lecture 5

BIOB11H3 Lecture Notes - Lecture 5: Northern Blot, Nonsense Mutation, Alternative Splicing


Department
Biological Sciences
Course Code
BIOB11H3
Professor
Dan Riggs
Lecture
5

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OP5: RNA splicing (RNA splicing and related phenomena)
Smart Pages: Figs 11-28, 29, 30, 32, 33, 34, 35, 36; also Figure 12.63
Vocabulary Words: consensus sequence/frameshift/missense mutation/snRNP/spliceosome
ribozyme/group 1 & group 2 introns/lariat/branch point/magic bullet/siRNA/microRNA
Central Ideas:
1. Conserved (consensus) sequences exist at the junctions of introns/exons
2. These sequences are recognized by spliceosome components and interactions occur
by RNA::RNA base pairing.
3. Self-splicing RNAs exist in nature and can be categorized as group I or group II
introns. This involves elaborate intrastrand basepairing, forming complex 3D
structures. Self-splicing implies that RNA has catalytic activity, like an enzyme-thus
the term ‘ribozyme’ was coined.
4. Northern blotting (RNA gel blotting) can be used to demonstrate the processivity of
RNA splicing as well as alternative splicing (we will get to this as a means of gene
regulation in chapter 12.
5. The consequences of incorrect splicing: frameshifts and the generation of missense
and/or nonsense mutations, abolishing protein function.
6. Small interfering RNAs (siRNAs) and microRNAs (miRNAs) play roles in development
and defense by mediating destruction of dsRNA or inhibiting translation.
Questions a scientist might ask:
1. What is the functional difference between a missense and a nonsense mutation?
2. Is there any evidence that some introns might be transposable elements?
3. How were the consensus sequences at the intron/exon junctions discovered?
Sample multiple choice question:
The following events occur in spliceosome mediated RNA maturation. Order them and
indicate which is the fourth from the beginning.
a. U2 snRNA interacts with the branch point A.
b. U1 snRNA interacts with the splice donor site.
c. Cleavage occurs at the splice donor site.
d. A lariat is formed by the intron.
e. The adjacent exons are ligated together.
Analytical Questions:
1. How could you use microscopy techniques to show that a gene contains introns?
2. See question 1 on page 428; question 1, page 432; question 19, page 454.
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