Lecture 16 Flashcards
miRNAs in RNAi are encoded by…
endogenous genes
miRNAs regulate… similar to…
gene expression similar to a dimmer switch (turning some on / some off)
miRNAs require blank before activation
processing
miRNAs are initially expressed as a… which may have up to…
long pri-miRNA which may have up to 5 miRNA precursors
the pri-miRNA is then processed to a…
70 bp long pre-miRNA
what 2 enzymes process pri-miRNA into pre-miRNA?
Drosha and Pasha
after processing, the pre-miRNA enters the…
cytoplasm
pre-miRNA is cut to 22 bps long by…
Dicer
after being cut by Dicer, the pre-miRNA is now…
a mature DS miRNA
mature miRNA is integrated into the RISC via…
argonaute, which can use one strand as a guide
dsRNA can be synthesized in the lab; you need blank and it can be made via blank
sequence of the target mRNA
PCR
3 methods for editing genome
1) TALEN
2) Zinc-finger nucleases (ZFN)
3) CRISPR
ZFNs are artificial…
restriction enzymes
ZFNs are made by the fusion of…
a zinc-finger TF domain and a DNA-cleavage domain
ZFN specificity is attained by engineering the…
zinc-finger TF domain
ZFNs can only be used in…
pairs - nuclease domain needs to dimerize
TALENs stands for…
transcription activator-like effector nuclease
TALENs are restriction enzymes that can be engineered to…
cut specific sequences
TALENs are made by the fusion of the…
DNA binding domain of a bacterial TF (TAL) to a DNA cleavage domain
the DNA cleavage domain is not sequence-specific but the researcher can…
engineer the TAL domain to be sequence-specific
TALs recognize DNA through…
34 AA repeat sequence
TALs recognize DNA through 34 AA repeat sequence, but the blank and blank position are hypervariable
12th and 13th
you can use one TALEN to induce blank or two to induce blank
a single break, a double break
CRISPR
clustered regularly interspaced short palindromic sequences (DNA seq)
CRISPR are derived from…
bacteriophages that had previously infected the bacteria
CRISPR is used to…
detect and destroy DNA from similar viruses during subsequent infections
upon viral infection, bacteria use CRISPR-associated (Cas) proteins to…
cut off a piece of viral DNA
this DNA sequence is integrated into the…
bacterial genome in between specific spacer regions
by storing this DNA sequence, the bacteria will…
1) remember this virus
2) will be able to counterattack it if it invades again
repeat sequences separate…
the spacer/viral sequences
