BHCM 307 Exam II

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Explain why the genetic code is degenerate
- because it is redundant
- most aa have more than one codon
- first 2 nucelotides are most important
Start and Stop codons
- Start is AUG (Met)
- 3 stop codons no aa listed
Wobble
- because third base pair isn't as important
- There for several tRNAs for each aa
- Inosine can bond with A, U, or C = wild card
Check pH question

:-)
aminoacyl- t RNA sythetases
- at least one sythetase for each aa
- catalyze reaction
- aa + ATP -> aminoacyl-AMP +PP_i
- aminoacyl-AMP + tRNA -> AMP + aminoacyl-tRNA
- they are specific to the aa and tRNA
- aa linked to 3' hydroxyl
Open Reading Frame
- ORF
- sequence starting with AT(U)G and ending with a stop codon
Translation In Eukaryotes
- ATG is usually the initiator codon
- 1 mRNA = 1 protein
Shine-Dalgarno Sequence
- sequence that binds to ribosomal RNA and initates translation by makin an initiation complex
- In prokaryotes
- can be anywhere in mRNA = can code for more than one protien
Chain initiation in Prokaryote

formation of initiation complex (mRNA, ribosomes subunits, initiation factors, fMet-tRNA)
Chain Elongation in Prokaryote
- Ribosome attaches to P site and A site
- Step 1: aminoacyl-tRNA binds to A site and anticodon loop binds to next codon
- Step 2: peptide chain moves from P to A site
- -catalyzed by peptidyltransferase
- -forms new peptide bond
- -proteins synthesized N to C
- Step 3: release tRNA at P site and translocation of peptidyl-tRNA from A to P
Chain termination in Prokaryotes
- Release factors bind to A site with a stop codon
- peptide is released from final tRNA hydrolysis
- mRNA is relased
- ribosomal subunits dissociate
cloning

isolating the gene in a form that can be replicated in abundance in another host
nucleases
- enzymes that degrade nucleic acids
- exonucleases: degrade from 5' to 3' end
- endonucleases: degrade nucleic acids by cleaving at specific internal sequences
- Developed by bacteria to kill virus DNA
Restriction endonucleases
- cut DNA at specific sequences
- usally palindromes
- hydrolyze the phosphodiester backbone
recognition sequence
- place were enzymes cleave
- usually even numbered
- size determines frequency
- longer more rare 6 bp = relatively rare
Sticky ends
- can rebind with H-base pairing
- weak and easily comes apart
- DNA ligase can can reform the phosiester covalent bond and make it strong
- It is like molecular glue
Vectors
- what DNA can be cloned into
- Used to use bacteriophages to make single strand DNA
- Now use plasmids circular DNA
Plasmids
- Circular DNA
- In bacteria
- Carry their own origin of replication
- carry antibiotic resistance genes
- many convenient restriction enzyme sites for cloning
- Engineered more sites
DNA RNA both neither
adenosine

both
DNA RNA both neither
ribose

RNA
Purines

hexose and pentose
pyrididines

hextose
DNA replication begins at the

origin
peptidyltransferase
- enzyme encoded by ribosomes
- makes new peptide bond
- fMet transfers
formation of intitation complex
- RNA
- small ribosomal sub units
- Initiation factor
- fMet-tRNA
- large ribosomal sub unit
cDNA
- copy DNA
- complementary DNA
- synthesized in vitro by reverse transcriptase using mRNA as a template
- represents genes expressed in a cell or tissue
- There are cDNA libraries
cloning DNA in Plasmid vectors
- digest wanted DNA and plasmid with same restriction enzyme
- mix DNA and allow sticky ends to anneal
- add DNA ligase and ATP
- add mixture to antibiotic sensitive bacteria that can take up the DNA
- grow in antibiotic
DNA electrophorese
- DNA is negatively charged
- smaller moves faster
- ethidum bromide- makes it floress
dideoxy nucleotides
- don't have the 3' or 2' OH
- therefore end the sequence
- if you know where it ends you can figure out the sequence
microray v. northern blot
- microray: does thousands at once
- attaches them to a chip
- tags and floresses them
- northern blot:puts it on a nylon sheet
- hybridizes it with DNA probe
PCR
- uses taq polymerase
- heat to 94 for 30 seconds
- cool to ~55 for 30 seconds
- heat to 74 for how ever long you want the bp

Author

MRK

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Card Set

BHCM 307 Exam II

Description

Translation

Updated

2013-10-26T20:14:51Z

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