Class Notes (834,986)
Canada (508,846)
Biology (6,794)

2-Differential Gene Expression.pdf

18 Pages
Unlock Document

Biology 3338A
Sashko Damjanovski

•  Genes aren’t destroyed but turned off •  Can regulate transcription at many levels •  Can regulate RNA processing (splice, adding tails etc.) •  Can selectively translate RNA 1 •  Use elements to study how things work •  Look where gene is expressed; if found in certain places, must be transcription factors that are specific to the area (ie. limb, brain) •  If think something is responsible for head, can clone enhancer sequences and hook them up to reporters 2 •  Have pancreatic precursor cellsà can make all parts of the pancreas •  Pdx1 is a pluripotent cell •  Know transcription factors that are needed to make B cell •  If pdx1 is fated to be a B cell, but if give if different transcription factors can end up being something else •  As cells are differentiating they are turning on different TF 3 •  Put a gene into a mouse that isn’t usually there; transgenic mouse usually are a KO of a gene (null) •  Easier to analyze what happens when u KO a gene, than making more of gene •  ICM cells are basically embryonic stem cells •  ICM can form an embryo but not extra embryonic tissue •  ICM cells are pluripotent to embryonic properà can form an entire embryo but not extra tissue •  ICM are not specified •  ICM can be cultured and can add thing to them; can then return them to the embryo without an cells noticing 4 •  Can place vector into cells; want to study bmp, place vector into cells, and grown mice with a lot of bmp •  Usually want to knock out •  Not all embryonic stem cells are going to take in the vectorà vector has a neo resistant gene; use neomycin to kill cells that didn’t take in vector •  Gene needs a promoter; when knocking it out its not an issue 5   •  Put neo r gene into coding of bmpà gene now is not going to code for bmp but for neo r (occurs in plasmid) •  Infect stem cell with plasmid •  Using a variety of enzymes induce homologous recombination with one of the copies of bmp gene •  Will now have one copy of neo r and one of bmp 6   •  Grow cells now in neo •  If don’t have trans gene will die •  Now eject them back into embryo •  Will form tissues that have transgene in them 7   •  Mice that are born contain transgene somewhere in them (random; don’t know where exactly they went) •  End up with chimeric mice (don’t all look the same) •  Some of the tissue in the mice contain transgene (called chimeric because not all tissues are the same) •  Want to knock out gene in every cell (right now they're heterozygous, need homozygous) •  Ideally want transgene to be in gametes so it can be passed onà know if its in gametes by isolating them (this destroys the gametes) •  Do southern blot with bits of the mice, isolate DNA and looking for confirmation that mouse is chimeric •  Neo gene is pre-set that tells us that transgene is present (mice don’t have neo gene) •  Want a homozygous mouse; take chimeric mouse (don’t know if its gametes, but hope) •  If in gametes, 2 possible genotypes (wt or KO) •  Know if its heterozygous (KO) run a blot •  If chimeric mate with other heterozygous sibling 8   •  Equivalent DNA but halve different transcription factors or regulatory factors 9 •  Three different ways of specificationà becomes autonomous •  If a cell is committed and determinedà know what is going to be an
More Less

Related notes for Biology 3338A

Log In


Join OneClass

Access over 10 million pages of study
documents for 1.3 million courses.

Sign up

Join to view


By registering, I agree to the Terms and Privacy Policies
Already have an account?
Just a few more details

So we can recommend you notes for your school.

Reset Password

Please enter below the email address you registered with and we will send you a link to reset your password.

Add your courses

Get notes from the top students in your class.