Lecture 10: Genetic Engineering Flashcards
Heritability for many traits is
h of around 0.5
Zinc finger nucleases
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
TALENs
transcription activator-like effector genome editing
- also induces DSB and homology directed repair
diff between crispr and zfns/talens
you have to build a protein complex for ZFNs and TALENs while crispr you just make a guide RNA
CRISPR
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
Parts of the CRISPR cas 9 system
- 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
Briefly, how do you use CRISPR
- design a crRNA for a sequence to be removed (can include multiple genes in the crRNA so multiple genes can be targeted)
- make a guide rna of the tracer plus comp rna
- create a genetic sequence you want to incorporate
- 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.
CRISPR has been used in…
mice, monkey, mushrooms, and…humans…
CRISPR babies
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)
Logistic hurdles of Genetic Engineering
- gene identity
- selectivity
- efficiency of DNA repair
- Unintended consequences
Logistic hurdle: Gene identity
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)
Examples of single gene disorders
PKU, HD
Logistic hurdle: Gene selectivity
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
Logistic hurdle: Efficiency of DNA repair
- 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
mosaicism vs chimera
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.
Efficiency of gene editing in Embryos
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
Synteny
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.
Consent
Adults can consent but young children and babies cannot consent to genetic engineering.
Public attitudes toward genetic modification
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
Family planning and economic forces
- 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.
Risk for mutations increases with…
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.
Who owns your DNA? Can you patent a gene? And why would you bother?
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
Myriad genetics
lost the patent suit to own the risk variant of BRCA1 which invalidated a lot of other gene patents.
Good side of gene patents
- 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
23andMe GWAS
GWAS using data from 23andMe on MDD identified a couple diff variants of significance (n= ~70 000)
Protecting your DNA
do you want:
- the government to have your DNA?
- the healthcare system?
- insurance companies?
all could pose issues - privacy, discriminatory practices
DNA testing and the law
- 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)
Genetic discrimination
Possibility that you would be treated differently if you have a gene mutation that increases the risk of a certain disorder.
GINA and Bill S-201
GINA - (USA) Genetic information non-discrimination act
Bill s-201 - same thing but Canadian
Problems with defining a certain gene as a ‘good’ gene
- 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
