Differential gene expression Flashcards
How are so many different cell types produced from the same genome?
Differential gene expression enables different cell types to express unique subsets of genes that give each cell type
their unique characteristics
Within a given organism, all cells have the same genome but not all genes are expressed in every cell
Differential gene expression
The term used to describe the difference in mRNA species found in different cell types/or in response to different stimuli
what triggers differential gene expression
developmental and environmental cues
promoter elements/sequences
Under the control of specific transcription factors, determine when and where specific genes will be expressed
Specific transcription factors
Dictate whether a gene will e expressed or not. There are hundreds of specific TFs, which are activated by different stimuli
Analysis of a reporter gene expression
GUS reporter gene
This technique enables us to explore whether specific gene promoter elements are activated in different cells or tissues
what does the introduction of a reporter gene construct into a cell or tissue do
it can determine whether the transcription factors required to switch on a particular promoter are present in that cell type.
reverse transcription-polymerase chain reaction
(RT-PCR)
Enables us to explore whether specific mRNAs, encoded by their respective genes are present in a sample by amplifying a DNA copy of the mRNA (called cDNA) using the RT-PCR technique
PCR reaction using a primer specific for cDNA
cDNA
Melt and separate strands with heat
Add PCR primers
Add heat-stable DNA polymerase - Taq polymerase
Results in strong amplification of the cDNA sequence
how do we reverse transcribe mRNA into cDNA
Isolate RNA from cells
Add oligo dT primer which binds to polyA tail on mRNA
Add reverse transcriptase, a viral enzyme that can make a DNA copy of an RNA molecule
Now we have RNA;DNA hybrids
Adding PCR primers and Taq polymerase amplifies and makes
retroviruses
have RNA genomes and use reverse transcriptase to make DNA copies of their genomes for integration into the host genome
cDNA library
represents a DNA copy generated using reverse transcriptase of all mRNAs expressed by a particular cell type or tissue
agarose gel electrophoresis
DNA is negatively charged and migrates from - to +
Agarose forms large pores through which DNA can migrate
