100% satisfaction guarantee Immediately available after payment Both online and in PDF No strings attached
logo-home
Advanced tools in molecular biology notes $9.13
Add to cart

Class notes

Advanced tools in molecular biology notes

 4 views  0 purchase
  • Course
  • Institution

Detailed notes on all of the lectures, self-study assignments, and lab practicals including a lot of pictures

Preview 3 out of 29  pages

  • October 29, 2024
  • 29
  • 2024/2025
  • Class notes
  • K.a. wolf
  • All classes
avatar-seller
DESIGNING MOLECULAR PROBES
CELL BIOLOGICAL PROCESSES

− replication:
o templated polymerization of a new strand
o C+G&A+T
o DNA polymerases only extend in 5' to 3' direction
o Okazaki fragments on the 5' strand




− transcription:
o a promoter sequence before
the start site
o RNA polymerase produces
mRNA used later by
ribosomes
o prokaryotes = picture
o eukaryotes = exons and
introns (spliced out at mRNA
maturation), enhancers,
silencers, transcription
factors etc.
− translation:
o ORF:
o open reading frame
o a small subpart of
mRNA that will be translated
o always starts with AUG and stops with UAA, UAG, or UGA
o UTR = untranslated region that doesn't encode for proteins but also consists of exons
o in prokaryotes there are sometimes multiple ribosome-binding sites coding for multiple
proteins → multiple reading frames on one strand

,− protein routing:
o routing of mature proteins to different cellular compartments
o transport through nuclear pores, across membranes, and by vesicles
o NLS (nuclear localization sequence) and NES (nuclear export sequence)
o signal peptide, transmembrane domain, GPI anchors, lipid tails etc.
− RNA flavours:
o mRNAs = coding for proteins
o rRNAs = form ribosome core and catalyse protein synthesis
o microRNAs:
o regulate gene expression
o binding to mRNA to make it double-stranded which is then degraded by the cell =
RNAi (RNA interferences)
o miRNA – endogenous
o siRNA and shRNA – experimental
o tRNAs = adaptors between mRNA and AAs during protein synthesis
o RNA polymerase I = transcribes most of rRNA genes
o RNA polymerase II = transcribes protein-coding genes, miRNA genes, and genes for some
small RNAs
o RNA polymerase III = transcribes tRNA genes, 5S rRNA gene, and genes for many other small
RNAs
− using translation and transcription for research:




MOLECULAR BIOLOGICAL TECHNIQUES

− cloning:
o using nuclear DNA (or a cell) from one organism to create a second organism with the same
nuclear DNA
o cutting a piece of DNA from one organism and inserting it into a 'vector' where it can be
replicated by a host organism → subcloning
− DNA polymerase:
o PCR
o creates DNA molecules by assembling nucleotides
o Taq polymerase = thermostable DNA polymerase isolated from Thermus aquaticus, no
proofreading

, o Pfu polymerase = thermostable DNA polymerase isolated from Pyrococcus furiosus,
proofreading (3’ – 5’ exonuclease activity)
− ligases:
o joining two ends of DNA strands
o if they contain 5’ phosphate groups
o if they ‘fit’ → hard for blunt ends, much more efficient if there is overlap between the ends
− end-modification enzymes:
o Klenow = DNA Polymerase fragment that makes 5’ overhangs blunt by filling in the missing
nucleotides
o Shrimp Alkaline Phosphatase = removes phosphate group at the 5’ end of DNA strands
preventing ligation of these fragments (chemically synthesized oligonucleotides lack 5’
phosphate groups)
o T4 Polynucleotide Kinase = adds (labelled) phosphate group to the 5’ end of DNA strands
enabling detection or ligation
− restriction enzymes:
o molecular scissors that cut double stranded DNA molecules at specific points
o found naturally in a wide variety of prokaryotes
o recognition site = either end of the palindromic sequence (between the two strands)
consisting of around 6 bp
o bacteria use them as defence against bacteriophages → prevent the replication of phage by
cleaving its DNA at specific site, host DNA is protected by methylases adding methyl groups
to A or C within the recognition site
o type 1 = recognize DNA sequence but cut it at random sites as far as 1k bp away from the
recognition site
o type 2 = recognize and cut within the recognition site
o type 3 = enzymes recognize sequences but cut at a different location close to the recognition
site (within 25 bp)
o endonuclease = cuts in the middle of DNA sequence
o exonuclease = cuts only at the end of DNA sequence
o restriction products: 5’ sticky ends, blunt ends (rarely used since ligation is difficult), and 3’
sticky ends
o isoschizomers = restriction enzymes that have the same recognition sequence as well as the
same cleavage site
o neoschizomers = restriction enzymes that have the same recognition sequence but cleave
the DNA at a different site within that sequence
− agents for subcloning:
o not on the exam
o pieces of DNA that can be replicated in the lab
o plasmids:
o circular pieces of double-stranded DNA naturally found in bacteria
o naturally obtained plasmids can carry antibiotic resistance genes, genes for
receptors, toxins, or other proteins
o replicate separately from the genome of the organism
o using pilus encoded by F plasmid to transfer the plasmid to the recipient organism

The benefits of buying summaries with Stuvia:

Guaranteed quality through customer reviews

Guaranteed quality through customer reviews

Stuvia customers have reviewed more than 700,000 summaries. This how you know that you are buying the best documents.

Quick and easy check-out

Quick and easy check-out

You can quickly pay through credit card or Stuvia-credit for the summaries. There is no membership needed.

Focus on what matters

Focus on what matters

Your fellow students write the study notes themselves, which is why the documents are always reliable and up-to-date. This ensures you quickly get to the core!

Frequently asked questions

What do I get when I buy this document?

You get a PDF, available immediately after your purchase. The purchased document is accessible anytime, anywhere and indefinitely through your profile.

Satisfaction guarantee: how does it work?

Our satisfaction guarantee ensures that you always find a study document that suits you well. You fill out a form, and our customer service team takes care of the rest.

Who am I buying these notes from?

Stuvia is a marketplace, so you are not buying this document from us, but from seller ninajunakovi. Stuvia facilitates payment to the seller.

Will I be stuck with a subscription?

No, you only buy these notes for $9.13. You're not tied to anything after your purchase.

Can Stuvia be trusted?

4.6 stars on Google & Trustpilot (+1000 reviews)

52510 documents were sold in the last 30 days

Founded in 2010, the go-to place to buy study notes for 14 years now

Start selling
$9.13
  • (0)
Add to cart
Added