1. CRISPR is cheap, easy to do, and powerful… is it also dangerous?
Anne B. Allison, Ph.D.
Teaching Demonstration
University of Richmond
3/20/17

2. Previously: natural genetic change
mechanisms of genetic variation
mobile genetic elements
how viruses interact with genomes
There is no natural mechanism for generating entirely new, long stretches of genetic sequence

3. Today: CRISPR how this new gene-editing tool works
promises and limitations of this technology
CRISPR provides a relatively easy and precise way to alter genomes
Big questions:
What specific measures may help policy keep pace with rapidly evolving technologies?
To what extent should dual-use research be private, restricted, or prohibited?

4. What are some ways people alter genetic information?
Let's take a few minutes to think-pair-share 

5. Biologists learn about gene function by altering genetic information
The past four decades have relied on techniques that are resource-intensive and/or lack precision:
-  targeted gene editing requires extensive training, cost, and time
- dominant negative mutants often target more than one molecular pathway
- RNA interference may have thousands of off-target effects
By comparison, CRISPR is relatively easy and precise

6. Cutting DNA can lead to:
How CRISPR works
Clustered Regularly Interspaced Short Palindromic Repeats
discovered as adaptive immune system in bacteria and engineered as a genome-editing tool
Cutting DNA can lead to:
     - disrupting gene
     - repairing a gene

7. CRISPR-Cas9 relies on extensive complementary basepairing to locate target DNA in a sea of chromatin
5’ ccntggtgcatctgactcctgtg 3’
3’ ggnaccacguagacugaggacac 5’
CRISPR guide RNA
Cas9 protein
3’ ggnaccacgtagactgaggacac 5’

8. Identifying on- and off-target sites

9. CRISPR may bind 20 or more off-target sites throughout the genome
This low off-target frequency (~6.5x10-9) is modest and impressive, nevertheless, one off-target cleavage could have disastrous consequences
Shengdar Q Tsai, et al. Nature Biotechnology (2015)

10. Components of the human genome
T. Ryan Gregory Nature Reviews Genetics (2005)

11. Recommended sources of information
Primary:
Jinek M, Chylinski K, Fonfara I, Hauer M, Doudna JA, Charpentier E. (2012). A programmable dual-RNA-guided DNA endonuclease in adaptive bacterial immunity. Science, 337(6096):
doi: /science
Shengdar Q Tsai, et al. (2015). GUIDE-seq enables genome-wide profiling of off-target cleavage by CRISPR-Cas nucleases. Nature Biotechnology, 33: 187–197. doi: /nbt
News:
‘Rewriting the Code of Life’ by Michael Specter in New Yorker January 2, 2017 issue
‘Do CRISPR Enthusiasts have their head in the sand about the safety of gene editing?’ by Sharon Begley in STAT News July 18, 2016
‘Genetically Engineer Almost Anything’ by Tim De Chant and Eleanor Nelsen in NOVA Next July 17, 2014
Video:
Dr. Jennifer Doudna’s iBiology video series:

12. You may access these slides and related materials at

13. Gene drives engineer evolution

14. Genetic code

15. One letter alters hemoglobin protein function
G A G G T G
C T C C A C
G A G G U G
glutamic acid valine
mutation
DNA
RNA
protein
normal protein is soluble
mutant protein forms fibers which distort shape of red blood cells causing these cells to clog blood vessels leading to a range of serious symptoms

16. Malaria has exerted selective pressure for the sickle cell trait in human populations
MAPS SHOWING MALARIA DISTRIBUTION AND MATCHING DISTRIBUTION OF HUMANS CARRYING THE HbS (SICKLE CELL) GENE