Human Anatomy & Physiology (11th Edition)
Human Anatomy & Physiology (11th Edition)
11th Edition
ISBN: 9780134580999
Author: Elaine N. Marieb, Katja N. Hoehn
Publisher: PEARSON
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3. In future lectures we will describe a technique known as Northern blotting that can be
used to detect RNA transcribed from a particular gene. Briefly, in this method a specific
RNA is detected using a short segment of cloned DNA as a probe. The DNA probe,
which is radioactive, is complementary to the RNA that the researcher wishes to detect.
After the radioactive probe DNA basepairs to the RNA, the RNA is visualized as a dark
(radioactive) band on an X-ray film. As shown here, the method of Northern blotting can
be used to determine the amount of a particular RNA transcribed in a given cell type. If
one type of cell produces twice as much of a particular mRNA as occurs in another cell,
the band will appear twice as intense. Also, the method can distinguish if alternative
RNA splicing has occurred to produce an RNA that has a different molecular mass.
875 nucleotides
675 nucleotides
1
Northern blot
2
-
3
Lane 1 is a sample of RNA isolated
from nerve cells.
Lane 2 is a sample of RNA isolated
from kidney cells. Nerve cells
produce twice as much of this RNA
as do kidney cells.
Lane 3 is a sample of RNA isolated
from spleen cells. Spleen cells
produce an alternatively spliced
version of this RNA that is about
200 nucleotides longer than theRNA
produced in nerve and kidney cells.
The question:
Let's suppose a researcher was interested in the effects of mutations on the expression
of a particular structural gene in eukaryotes. The gene has one intron that is 450
nucleotides long. After this intron is removed from the pre-mRNA, the mRNA transcript
is 1100 nucleotides in length. Diploid somatic cells have two copies of this gene.
Describe (or draw) the expected results of a Northern blot using mRNA from the cytosol
of somatic cells, which were obtained from the following individuals:
Lane 1: A normal individual
Lane 2: An individual homozygous for a deletion that removes the -50 to -100 region of
the gene that encodes this mRNA
Lane 3: An individual heterozygous in which one gene is normal and the other gene had
a deletion that removes the -50 to -100 region
Lane 4: An individual homozygous for a mutation that introduces an early stop codon
into the middle of the coding sequence of the gene
Lane 5: An individual homozygous for a three-nucleotide deletion that removes the AG
sequence at the 3' splice site
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Transcribed Image Text:3. In future lectures we will describe a technique known as Northern blotting that can be used to detect RNA transcribed from a particular gene. Briefly, in this method a specific RNA is detected using a short segment of cloned DNA as a probe. The DNA probe, which is radioactive, is complementary to the RNA that the researcher wishes to detect. After the radioactive probe DNA basepairs to the RNA, the RNA is visualized as a dark (radioactive) band on an X-ray film. As shown here, the method of Northern blotting can be used to determine the amount of a particular RNA transcribed in a given cell type. If one type of cell produces twice as much of a particular mRNA as occurs in another cell, the band will appear twice as intense. Also, the method can distinguish if alternative RNA splicing has occurred to produce an RNA that has a different molecular mass. 875 nucleotides 675 nucleotides 1 Northern blot 2 - 3 Lane 1 is a sample of RNA isolated from nerve cells. Lane 2 is a sample of RNA isolated from kidney cells. Nerve cells produce twice as much of this RNA as do kidney cells. Lane 3 is a sample of RNA isolated from spleen cells. Spleen cells produce an alternatively spliced version of this RNA that is about 200 nucleotides longer than theRNA produced in nerve and kidney cells. The question: Let's suppose a researcher was interested in the effects of mutations on the expression of a particular structural gene in eukaryotes. The gene has one intron that is 450 nucleotides long. After this intron is removed from the pre-mRNA, the mRNA transcript is 1100 nucleotides in length. Diploid somatic cells have two copies of this gene. Describe (or draw) the expected results of a Northern blot using mRNA from the cytosol of somatic cells, which were obtained from the following individuals: Lane 1: A normal individual Lane 2: An individual homozygous for a deletion that removes the -50 to -100 region of the gene that encodes this mRNA Lane 3: An individual heterozygous in which one gene is normal and the other gene had a deletion that removes the -50 to -100 region Lane 4: An individual homozygous for a mutation that introduces an early stop codon into the middle of the coding sequence of the gene Lane 5: An individual homozygous for a three-nucleotide deletion that removes the AG sequence at the 3' splice site
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