Difference between revisions of "Team:NTU-Singapore/Demonstrate"
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<p>We tried to activate an endogenous gene, ADAR3 and ADAR2 in HEK293 cells. ADAR3 is an RNA-specific adenosine deaminase involved in A-I conversion. This gene is specifically expressed in human neuronal cells and predisposes stem cells towards a neuronal fate. Our advisor, Dr. Yuan Ming, is also using dCas9-VPR for the induction of ADAR3 and ADAR2 expression in H1 human embryonic stem cells. We used the gRNA designed by Dr. Yuan Ming to test out if our truncated dCas9 could activate endogenous ADAR2 and ADAR3 in HEK293 and H1 cells.</p> | <p>We tried to activate an endogenous gene, ADAR3 and ADAR2 in HEK293 cells. ADAR3 is an RNA-specific adenosine deaminase involved in A-I conversion. This gene is specifically expressed in human neuronal cells and predisposes stem cells towards a neuronal fate. Our advisor, Dr. Yuan Ming, is also using dCas9-VPR for the induction of ADAR3 and ADAR2 expression in H1 human embryonic stem cells. We used the gRNA designed by Dr. Yuan Ming to test out if our truncated dCas9 could activate endogenous ADAR2 and ADAR3 in HEK293 and H1 cells.</p> | ||
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Revision as of 21:16, 19 October 2016
CRISPRy! I choose you!
To infinity! And beyond!!
Project Truncation
Knowing that several of our truncated mutants is able to activate our reporter gene, we now turned our attention to using dCas9 for the activation of an endogenous gene. The difference between reporter gene and endogenous gene activation is the copy number of our template within the cell, with the former in larger amounts than the latter.
We tried to activate an endogenous gene, ADAR3 and ADAR2 in HEK293 cells. ADAR3 is an RNA-specific adenosine deaminase involved in A-I conversion. This gene is specifically expressed in human neuronal cells and predisposes stem cells towards a neuronal fate. Our advisor, Dr. Yuan Ming, is also using dCas9-VPR for the induction of ADAR3 and ADAR2 expression in H1 human embryonic stem cells. We used the gRNA designed by Dr. Yuan Ming to test out if our truncated dCas9 could activate endogenous ADAR2 and ADAR3 in HEK293 and H1 cells.
ADAR2 activation in HEK293 cells with only one gRNA used.
