Lecture 10: Genetic Engineering

Question 1

Heritability for many traits is

Answer 1

h of around 0.5

Question 2

Zinc finger nucleases

Answer 2

little bits that anneal to palindromic sequences on either side of desired cut site, the ZNF binds and then cleaves

• we WANT homologous recombination that includes the donor plasmid
• but sometimes we get off target nonhomologous end joining

Question 3

TALENs

Answer 3

transcription activator-like effector genome editing

- also induces DSB and homology directed repair

Question 4

diff between crispr and zfns/talens

Answer 4

you have to build a protein complex for ZFNs and TALENs while crispr you just make a guide RNA

Question 5

CRISPR

Answer 5

Clustered Regularly interspaced Short Palindromic Repeats

• bacterial immunity to bacteriophages
• the viral DNA is incorporated into the bacterial CRISPR sequence so that it can be recognized and destroyed later on

Question 6

Parts of the CRISPR cas 9 system

Answer 6

• Cas 9 protein which cleaves
• the CRISPR RNA (CrRNA for the sequence to be targeted c= complementary)
• the transactivating RNA (tracrRNA) (acts as a scaffold)

TracrRNA and crRNA are combined to make one guide RNA in this system

Question 7

Briefly, how do you use CRISPR

Answer 7

1. design a crRNA for a sequence to be removed (can include multiple genes in the crRNA so multiple genes can be targeted)
2. make a guide rna of the tracer plus comp rna
3. create a genetic sequence you want to incorporate
4. inject the above elements. Example in monkeys, culture egg in petri dish, inject sperm, then after arouns 9 hours inject the cas9 mRNA and guide rna. Transfer the embryo into a surrogate mother.

Question 8

CRISPR has been used in…

Answer 8

mice, monkey, mushrooms, and…humans…

Question 9

CRISPR babies

Answer 9

twin girls in China were first humans with germline genetic modification

• targeted CCR5 which produces a protein important for WBCs and immune function. Deletion of this gene is linked to HIV resistance
• Performed by Dr. He Jiankiu, no papers on it tho (got put in prison for this)

Question 10

Logistic hurdles of Genetic Engineering

Answer 10

• gene identity
• selectivity
• efficiency of DNA repair
• Unintended consequences

Question 11

Logistic hurdle: Gene identity

Answer 11

we know traits have a genetic basis but we often don’t know all the genes involved

• much of our info on the genetics comes from GWAS (probz with reproducibility, small effect of the genes found, and variance of gene effect in different populations
• polygenic scores using common SNPs explain only 20% of the variance seen in traits but usually lower
• also pleiotropy (could have unintended consequences)

Question 12

Examples of single gene disorders

Answer 12

PKU, HD

Question 13

Logistic hurdle: Gene selectivity

Answer 13

CRISPR is imperfect and sometimes off target effects occur. Other non-specific effects on other genes.

• if we use crispr to edit a quantitative trait (which is influenced by many, many genes) that comes with a high risk of side effects
• therefore, genetic modification is most suitable for editing monogenic traits

Question 14

Logistic hurdle: Efficiency of DNA repair

Answer 14

• we don’t want to delete a gene allele we want to REPLACE it with another allele which is much trickier than just deleting
• repair is inefficient (doesn’t happen in every cell, can accidentally not incorporate the donor plasmid) and also unintended errors can occur

Question 15

mosaicism vs chimera

Answer 15

mosaicism - one zygote, induce a genetic change at the four cell stage and the person becomes a mosaic

chimera - two zygotes, one normal and one genetically edited and then fusion or exchange of cells from genetically edited zygote into normal person.

Question 16

Efficiency of gene editing in Embryos

Answer 16

early experiments suggested that CRISPR/Cas9 editing in non-viable human embryos is inefficient (only 50% of 54 embryos had a modification) and that there is also evidence of off-target effects
- current ongoing experiments in China, Sweden, and the US

Question 17

Synteny

Answer 17

In comparative genomics, synteny is the preserved order of genes on chromosomes of related species which results from descent from a common ancestor. Basically the concept that genes will be found in the same order on the same chromosome between species descended from a common ancestor.

Question 18

Consent

Answer 18

Adults can consent but young children and babies cannot consent to genetic engineering.

Question 19

Public attitudes toward genetic modification

Answer 19

There’s support for genetic modification to prevent disease but little support of modification to “improve” function (yes to cure/prevent disease, no to enhance)
- generally support for ‘’ is higher in men, highly educated ppl, and non-religious

Question 20

Family planning and economic forces

Answer 20

• ppl already consider abortion if they know the child will face significant health issues or early mortality
• rising population globally and reduced fertility rate (especially in countries with one child policy) attitudes might change to support the production of genetically modified embryos.

Question 21

Risk for mutations increases with…

Answer 21

Age
- Ppl are now having families later and later. Risk for mutations increases with age as does potential for certain disorders like ASD and SZ.

Question 22

Who owns your DNA? Can you patent a gene? And why would you bother?

Answer 22

You’d bother because money. Patenting a risk variant of BRCA1 would allow you exclusive access to a for-profit testing system.
- for now, we own our own natural genes but synthetic gene patents could still occur

Question 23

Myriad genetics

Answer 23

lost the patent suit to own the risk variant of BRCA1 which invalidated a lot of other gene patents.

Question 24

Good side of gene patents

Answer 24

• reserved for synthetic genes only
• could provide financial incentive for people to get into research
• could lead to unprecedented development in the field
• regulation and protection would be more manageable

Question 25

23andMe GWAS

Answer 25

GWAS using data from 23andMe on MDD identified a couple diff variants of significance (n= ~70 000)

Question 26

Protecting your DNA

Answer 26

do you want:

• the government to have your DNA?
• the healthcare system?
• insurance companies?

all could pose issues - privacy, discriminatory practices

Question 27

DNA testing and the law

Answer 27

• the golden state killer was identified using familial DNA evidence
• fertility doctors used their own sperm and fathered 15-200 children thinking no one would know (norman barwin, cecil jacobson, jan karbaat)

Question 28

Genetic discrimination

Answer 28

Possibility that you would be treated differently if you have a gene mutation that increases the risk of a certain disorder.

Question 29

GINA and Bill S-201

Answer 29

GINA - (USA) Genetic information non-discrimination act

Bill s-201 - same thing but Canadian

Question 30

Problems with defining a certain gene as a ‘good’ gene

Answer 30

• if you suggest that only one gene is a good gene, you are not considering the fact increased genetic diversity is super important and also suggesting that you know all the functions of a gene.
• traits good in one context may be bad in another