Transcription Flashcards
What are promoters?
promoters are DNA Sequence that contain a region called operator.
Mechanism by which prokaryotes control transcription.?
- promoters
- sigma factors
What are operators?
DNA sequence in the promoter that contains the binding site for repressor proteins.
Positive regulation of operon?
in prokaryotes
Inducible operon. Helps turn transcription on
- gene is normally “off” because of a repressor bound to the operator
- inducer binds to repressor, repressor fall off the DNA.
- transcription is turn “on”
( example the lac operon)
Negative regulation of operon?
in prokaryotes
Repressible operon. Will turn transcription off.
- genes is normally “ on”
- a corepresor binds ( activate) to a repressor
- corepresor + repressor ( activated repressor) binds to the operator
- transcription is block
( example the trip operon)
Transcription regulation in eukaryotes ?
- Regulatory transcription factors
- DNA methylation.
How Transcription factors regulate transcription in eukaryotes?
TF clear the way for BTF( increase affinity) enhancement
TF block BTF -> –> reduce transcription
How DNA methylation regulate( block) transcription in eukaryotes?
A methyl group is added via methyl transferase to a cytosine.
The CG sequence is used as recognition.
- methylated bases serve as a binding site for MGDs
MGDs block transcription.
What is MGDs and what they do.
MGDs are Methyl C-pG binding domain protein.
- they block transcription factor assembly
- block chromatin remodeling
- recruit histone deacetylases.
Properties of methylation?
Cell, tissue and person specific
Programmed during early development
Influence by the environment
Depends on what the mother of my mom did when she was pregnant with my mom.
What is epigenetics?
The total collection DNA structural alterations
- even though all the cell in a human body have the same genome( DNA information) the cells differ in the methylation pattern.
Example: my brain cells and my liver cells have the same DNA sequence but they differ in their methylation pattern.
What is imprinting?
Specific gene is methylated different in the 2 stran becaes depending in what parent came from
- only one copy is methylated
- having only one functional copy
Epigenetics relationship in animal cloning?
Problems: a few genes fail to be demythylated( reset failed)
And the new animal ( clone) start in off position.
Methylation of the original donor of DNA can be different from that of the clone and therefore appearance and behavior may differ .
How transcription ( expression) can be measured.
- Report assay
- northern blot
- microarray analysis
How report assay is used to determine levels of transcription.?
-Report assay measure enhancer/promoter activity.
- -Procedure:Insert promoter/enhancer of the gene of interest in front of a reporter gene.
- the more transcription, the more mRNA, the protein is made( usually a fluoxetine protein GFP)
How does northern blog is used to analyze level of transcription?
If a gene is transcribed a lot , there is a lot of mRNA at anytime inside the cell. Provide a relative quantization of the amount of mRNA and therefore the relative amount of transcription.
Procedure:
Extract the total mRNA from different cell types.
Separate the deferent mRNA by gel electrophoresis.
Transfer from gel to nitrocellulose
Add a gene specific probe label with a radioactive
A single radioactive band should be seen.why only 1 band?
( one band because the radioactive probe will bind to just one containing a complementary RNA molecules)
Th
How is microarray analysis used to determine transcription levels?
Provides the relative amount of each mRNA molecule in the cell.
Procedures:
-Isolate cytoplasmatic mRNA from cells and converted to cDNA.( used reverese transcriptase)
-add label the single strand of cDNA with a dye.
- add the label cDNA to the DNA microarray
(each gene spot has a single stranded DNA fragment)
-labeled cDNA will go to the spot with the corresponding gene (corresponding single DNA fragment)
The more mRNA. The moe cDNA will bind to the gene spot
Diseases associated with transcription factors?
- cancer
- fragile syndrome
- prader-willi & Angelman syndrome( caused by abnormal imprinting)
Evidence that a transcription couple-DNA repair system exist?
Transcribed DNA sequence are repaired much more efficiently than non- transcribed strands, if damaged is introduced to both.
-procedure
When DNA POL II gets to a damaged nucleotide, it stalls.
Certain proteins recognize the stall RNA POL II and will bind to it.
Those proteins recruit RNA REPAIR enzymes
Damaged is fixed, RNA POL continues.
( clockayne Syndrome: defects associated with repair system
What is the clockayne syndrome?
Syndrome caused by defects with the Elaine syndrome.
Suffer from : premature aging, short stature, and have muscle/ bone defecs.
