Topic.3.Cell.Determination&Stem.cells.powerpoint.3.1.IPSC.Gene.editing Flashcards
Induced pluripotent cells(iPSC)
adult somatic cells genetically reprogrammed to an ESC state due to forced gene expression (don’t need to destroy embryos anymore).
Insert transcription factors: Oct 4,Sox2,c-Myc,klf4)
What was this forced gene expression in iPSC?
retrovirus to get master regulatory genes into PSCs
What is a retrovirus?
Retrovirus is an RNA virus that uses reverse transcriptase for replication-generate DNA from an RNA template. The DNA is then integrated into genome through integrase enzyme
T/F HIV is a retrovirus
true
Lentivirus are a family of viruses that are responsible for diseases like AIDS, they are a subgroup of retrovirus. How are they treated?
Lentiviral vectors in gene therapy is a method by which genes can be inserted, modified, or deleted in organisms using lentiviruses. They delete genes that are bad and replace them with good genes, then repackage so it can enter another cell.
It might be concerning to use iPSC because..
retrovirus can randomly insert genes causing mutations and cancer
CRISPR
Clustered Regularly Interspersed Short Palindromic Repeats
What is CRISPR?
In prokaryotic DNA, found spaces in between DNA bases, of viral origin.
its immune defense system that degrades invading DNA
CRISPR Cas9 complex made out of:
Cas9:DNA cutting protein
SgRNA:single guide RNA
crRNA binds to targeted DNA sequence
tracrRNA bps with crRNA so Cas9 can locate target
PAM is DNA on target DNA so Cas9 can bind and cleave DNA
The double strands made by CrispR-cas9 can be repaired by?
NHEJ: Non-homologous end joining: DNA ends are ligated
back together, but usually with the introduction of a small
insertion or deletion
HR: Homology-directed repair: donor DNA with homologous
sequences at either end can be integrated into the site
dCas9
Variation of Cas9, can’t cut DNA but can bind to DNA preventing endogenous gene expression
