U3 AOS 1 Flashcards
Nucleic Acids
DNA: Deoxyribonucleic Acid
RNA: Ribonuic Acid
Nucleotide: composed of phosphate, 5-Carbone sugar, Nitrogenous Base
RNA types
a) messenger RNA (mRNA) - carries genetic info from nucleus to ribosome to direct protein synthesis
b) ribosomal RNA (rRNA) - makes up ribosomes
c) transfer RNA (tRNA) - transfer amino acids to ribosomes for protein synthesis
Genetic Code
Represents genetic information stored in DNA as a triplet code with gene selection
A gene
A section of DNA that codes for proteins
Gene Expression Steps: Transcription
**Initiation- ** proteins Bind to a promoter region when RNA breaks bonds and unwinds DNA
** Elongation-** RNA moves in a 3’ to 5’ direction reading the nucleotide sequence and adding complementary nucleotides to form pre-mRNA
** Termination -** A terminator sequence is reached. DNA rewinds and reforms.
Gene Expression Steps - RNA Processing
Turns pre-mRNA into mRNA
• Adds 5’ methyl cap and 3’ Poly-A-Tail
• (removes introns, splices exons)
Gene Expression Steps - Translation
Initiation - Ribosomes bind to mRNA and reads code to make a chain of amino acids
Elongation - tRNA binds a complementary anticodon to the mRNA codon.
Termination - stop codon reached. Polypeptide formed.
Degeneration
Several codons may code for the same amino acid
Exons
Coding regions of a gene
(Transcribed and Translated )
Introns
Non -coding regions of a gene
(Transcribed only)
Promoter Region
A region of DNA that contains a binding site for RNA polymerase
Operator Region
Regions of DNA that the repressor proteins binds to.
Regulatory + Structural Genes
Regulatory - produces proteins that turn structural proteins on and off.
**Structural - ** codes for proteins needed for the body to function.
Amino acid
R group
Cooh group
Central carbon
Hydrogen ion
Amine group
Role of Rough Endoplasmic Reticulum
Modify Polypeptide
Role of Ribosomes
Translate polypeptide chains
Role of Transport Vesicles
Transport modified polypeptide
Golgi Vesicle
Transport proteins to cell membrane
Plasma Membrane
Proteins are secreted to the extracellular fluid.
Enzymes
Speed up reactions
Amylase. Catalase.
Proteome
Set of proteins produced by a cell or organism
Non Competitive Inhibitors
A molecule that binds to an enzyme at any site other than the active site
Changes the shape so that the substrate can no longer bind to active site.
Competitive Inhibition
A substrate that competes with a substrate for an active site.
Lock and Key Mechanism
The substrate fits into the active site to enable a reaction. If it doesn’t fit then no reaction occurs.
Induced Fit Model
The substrate binds to the active site of an enzyme, changing the shape of site to fit.
Ligase
Enzyme that joines nucleic acids together
RNA Polymerase - manipulating DNA
• Unwinds in a 5’ to 3’ direction
• Forms RNA (and 3 types) by transcription of a gene
DNA Polymerase
Assembles DNA
Endonuclease (restriction enzyme)
**Cut up DNA into smaller fragments by cutting at recognition site. **
Resulting Fragments can be;
**1. Sticky Ends ** - can only join to sticky ends with complementary base sequences. Overhang
2. Blunt Ends - can join to blunt ends. No Overhang
Reverse Transcriptase Steps
- MRNA is isolated from cytosol
- Poly-A- tail is added to mRNA
- Primer is added and binds to tail
- Reverse enzyme added.
- When DNA is complete, mRNA removed.
- Polymerase enzyme added.
- Double Stranded DNA product.
**Cut twice.
If linear = 3pieces
If circular = 2 pieces **
Sticky Ends
- A restriction endonuclease cuts DNA at a recognition site.
- Cut produces sticky ends
- Two sticky ends cut by the same endonuclease join by base pairing
Ligation
**Joins the fragments together **
Steps: sticky end
1. Sticky ends are formed by endonuclease.
- Weak hydrogen bonds attract
- Other ends of foreign DNA are attached to the remaining sticky end of the plasmid with ligase
Steps: blunt.
1. Two fragments come into contact and using ligase, join together.
Crispr-Cas9
A genome editiong tool that allows the cutting and editing of DNA.
Steps:
- Create a complex with a short RNA sequence (sgRNA) and CRISPR. The RNA sequence is complementary to target DNA
- Using guide RNA, Cas 9 identifies the DNA in genome and cuts strands.
- Insert donor DNA at cut
- Cell repairs DNA integrating change.
PCR Process
Steps
- Denaturation- separate DNA by **heating ** to 95-98 degrees for 5 minutes to break hydrogen bonds.
- Annealing- mixture cools to 55 degrees to allow primers to bind to complementary strands.
- Elongation- Heat to 70 degrees, DNA polymerase moves along strands, adding nucleotides to 3’ end.
Gel Electrophoresis: Steps
- Gel is made
- DNA is extracted.
- Endonucleases break DNA into
fragments, or PCR makes copies.
- DNA loaded into wells
- One lane has a marked solution.
- Current applied to gel : negatively charged DNA attaches to positively charged gel.
- Small particles move faster than large
Transgenic Organism
Types of GMOs that contain genes from another species.
Transgenic Organism
A type of GMO that has genes from another species.
Used to improve crops and prevent disease.
Implications
- Biological: safety of consumption, cross pollination
- Social: equal access, labelling, pricing
-Ethical: intervention in evolutionary process.
Active site
The region of an enzyme where substrate molecules bind and undergo a chemical reaction
Allosteric site
Any part of an enzyme other than an active site
Exocytosis
Bulk transport of substances out of a cell
Endocytosis
Bulk transport of substances into a cell
Vector in recombinant DNA technology
Adds foreign DNA into a cell or organism
Recombinant plasmid
A plasmid containing foreign genetic material
PAM (Protospacer adjacent motif)
A short sequence of nucleotides downstream from a target sequence that allows the binding of Cas9.
- It separates the strands of DNA to allow the binding of single guide RNA (sgRNA) to the complementary DNA sequence.
Cofactor
a molecule that aids enzyme function
What are the three stages and their corresponding temperatures that are cycled through in PCR?
**heating ** to 95-98 degrees for 5 minutes for denaturation, 55°C for annealing and 70°C for extension
Which end of a nucleotide can new nucleotides be added to?
3’ end
Which base replaces thymine in RNA?
Uracil
Principles of the trp operon
The trp operon is found in E. coli bacteria. Tryptophan is an amino acid that is needed in small quantities, and so to save energy and resources in a cell, the genes that code for it are only expressed when it is in low levels. These genes are part of the trp operon, which consists of 5 structural genes (trp E to trp A) that share common promoter and operator regions upstream of the genes. When tryptophan is present, it binds to repressor molecules, which change their shape so that they are complementary in shape to the operator region, and bind to it, so that RNA polymerase cannot pass to transcribe the genes. However if tryptophan is present in low levels, it does not bind to the repressor molecule, and so the repressor molecule is not complementary to the operator region, and transcription continues.
Short tandem repeats
Short repeated DNA sequences that can be used to match a persons genetic material to a sample
CRISPR-Cas 9 principles
CRISPR-Cas9 in bacteria has two important features, nucleotide repeats and spacers. The spacers are segments of DNA cut from invading viruses (bacteriophages) stored in the bacteria’s genome that allow the bacteria to recognise the same virus in the event of subsequent invasions. Cas9 is an endonuclease associated with CRISPR and precisely cuts DNA. Cas9 will only cut out the target sequence if it recognises a very short nucleotide sequence adjacent to the target spacer called a protospacer adjacent motif (PAM) sequence. When utilised in genetic engineering, a single-guide RNA (sgRNA) sequence that joins the Cas9 complex will bind to a complementary DNA sequence and cut it at this sequence, acting as a spot where DNA can be added or removed, and the cell will detect and repair the broken sequence of DNA incorporating the changes made.
What is the purpose of gel electrophoresis?
To separate fragments of DNA based on their base length
How many hydrogen bonds form between adenine and thymine/uracil?
2
How many hydrogen bonds form between cytosine and guanine?
3
Terms for groups of three nucleotides
Base triplets (DNA) –> Codons (mRNA) –> Anticodons (tRNA)
Terminator region
A region of DNA found downstream of a gene that stops the transcription of the gene
Function of methyl cap and poly-A tail in post-transcriptional modification
Increase mRNA stability and prevent degradation
Reverse transcriptase
An enzyme that synthesises single-stranded DNA using RNA as a template.
Process to produce recombinant plasmids
1) Target DNA is cut out from a DNA sequence using restriction enzymes that leave sticky ends.
2) The bacterial plasmid is then cut by the same restriction enzyme to leave sticky ends that are complementary to that of the foreign DNA.
3) The plasmid and the DNA are exposed to each other with DNA ligase to catalyse the formation of phosphodiester bonds to join the fragments.
4) Some plasmids will accept the DNA.
Differences between RNA and DNA
- RNA contains uracil, while DNA contains thymine,
- RNA is double stranded while DNA is double stranded
- RNA contains ribose sugar while DNA contains deoxyribose sugar
Why is gene regulation important for an organism?
Proteins will only be produced when they are required, which saves resources and energy (in the form of ATP) that are required to produce proteins
What does the presence of uracil or thymine in a nucleic acid indicate respectively?
That the nucleic acid is RNA or DNA
steps to form bioethanol
- Find and deconstruct biomass
- Break down starch and cellulose into glucose
- Make ethanol via anaerobic fermentation
- Dehydrate and purify ethanol
repression steps
- Tryptophan binds to TRP repressor.
- Binds to operator region, blocking RNA Polymerase from binding.
Therefore stopping TRP production
Attenuation
- Leader segment of TRP is translated by RNA Polymerase
- Forms hairpin loop to stop translation
