DNA Flashcards
What is the light DNA?
Euchromatin
What is the dark DNA in nucleus?
Heterochromatin
What DNA is expressed?
Euchromatin
What is a nucleosome?
Chromatin core that’s positive
DNA linking which is negative
Strong interactions
Where are beads on a string located?
Euchromatin hence gene expressed
What and where are solenoids located?
Tightly packed chromatin and DNA hence located in Heterochromatin which is not expressed. 30nm
How is DNA tightly held together?
Scaffolding proteins
How many chromosomes in the human?
24
22 are autosomal and 2 are sex
What is a nucleic acid?
DNA & RNA
Polynucleotides - linear polymer of nucleotides
Difference in the sugars in RNA and DNA?
RNA has ribose which has an OH in C2
DNA has deoxyribose with just a H in C2
What are the 2 types of bases?
Purine and Pyramidine
What is a purine?
2 rings
Adenine & Guanine
What is a pyramidine?
1 ring
Cytosine, thymine and uracil
Base A and nucleotide name?
Adenine –> Adenosine/ Deoxyadenosine
Base G and nucleotide name?
Guanine – Guanosine /deoxy…
Base T and nucleotide name?
Thymine —> depxythymidine NO RNA
Base C and nucleotide name?
Cytosine — Cytidine — deoxy…
Base U and nucleotide name?
Uracil —- Uridine ONLY IN RNA
Full name for base?
Nitrogenous bases
Base pairs occur between…
One purine and one pyramidine
A to
T to 2 (h-bonds)
C to
G with 3 (h-bonds)
How are RNA and DNA labelled?
Top strand - 5’ to 3’
With 5’ being phosphate
What do you add nucleotides to?
Add to 5’. The phosphate
Duplex structure?
Complementary anti-parallel strands in DNA
What is at 3’?
-OH
How are nucleotides connected?
Covalent bonds between phosphates called phosphodiester bonds. also polarity in each nucleotide
What is RNA stem-loop structure?
Single strand loops back on itself forming h bonds on anti-parallel complementary sequences
How often are turns in DNA?
Every 10 bases or every 3.4nm
How far are bases away in DNA?
0.34nm
Describe structure of DNA?
Anti-parallel double stranded helix. Complimentary
Planar bases
Rotates right handed
Which groove has the baes exposed?
Major groove
What stage is DNA replicated?
S phase
When is chromatin present?
Interphase
Mitosis chromosomes are…?
Highly condensed fibres hence genome is unexpressed and can’t replicated as condensed
Interphase chromatin is…?
Decondensed hence long thin beads on a string hence can be expressed and replicated
What does DNA replication require to activate?
Activated precursors, dNTPs (deoxyribonucleoside triphosphate)
Why is the hydrolysis of ATP important for DNA replication?
Drives each reaction to extend existing chain by one unit
What direction is chain growth in DNA replication
5’ to 3’
What is semi-conservative replication?
The replication that occurs to human DNA.
Each daughter has one of the strands from original strand of DNA
What are the 3 stages of DNA rep?
Initiation - helicase + primase
Elongation - extend 5’ to 3’ and Okazaki Fragments
Termination
What is initiation of DNA rep?
The DNA helix unravelled by DNA helicase, which breaks the hydrogen bonds between bases. This there exposes the base to DNA polymerase. However DNA polymerase only extends from the 3’ end of pre-existing DNA.
Primase is also required to initiate the replication of each strand
What is elongation in DNA rep?
The DNA polymerase replicates the leading strand from 5’ to 3’.
The lagging strand is replicated discontinuously hence produces Okazaki Fragments. These fragments are joined together by DNA ligase which covalently bonds the P to OH
What is termination of DNA rep?
The DNA has been replicated and there is semi-conservative arrangements of old and new strands
What is transition point mutation?
Mutation of one base. Purine to purine. Or pyramidine to pyramidine
Transversion point mutation?
Purine to pyramidine or vice versa
If the mutation occurs in non-coding DNA of genome?
Little or no phenotypic effect
Mutation within or near genes?
Result in disease as affects gene expression or promoter regions eg TATA box
How can silent mutations still be bad?
Might not change the AA but can disrupt RNA splicing hence heritable disease
Missense mutations?
AA substituted for another - single base change
Nonsense mutation?
changes to AA hence affecting stop codon
Where else apart from the gene is it bad for a mutation?
Binding sites, promoter sequence, splice sites
Insertion of millions of nucleotides?
Tandem duplication
What is premature termination condons (PTCs)?
mRNA degraded as defence - nonsense mediated decay to not produce mutated protein
Frame shift mutation?
Reading frame of mRNA altered due to insertion or deletion not being a multiple of 3 hence shift ping the bases in each codon.
Conservative missense mutations?
Some AA substitutions better than others for example valine to alanine is better as similar AA hence less damage.
Spotneous mutations?
Not caused by exposure to a known mutagen causing errors in DNA replication hence slightly chemical instability
Types of errors in DNA replication?
Tautomeric forms
Slippage
What is tautomeric forms?
Rare. Altered base-pairing. Proton changes positions hence behaves like altered template ANOMALOUS base.
Tautomeric causes C to A pairing and T to G pairing.
If T in rare form, DNA polymerase sees T as C hence G in new strand.
What is slippage in terms of DNA replication error?
Looping out hence extra repeat as additional nucleotide added to new strand. The template strand loops out hence omission of 1 nucleotide on new strand
The rate of spontaneous maturation a depends of…?
Size
Sequence
What causes induced mutations?
Chemicals - nitrous acid replaces amino to keto acids C –> U hence pairs to A. A –> H (hypoxanthine) g –> X (xanthine) hence pairs to C
Radiation
Ethicist bromide intercalates DNA
Ethyl Methane Sulphate removes purine ring hence bind to any bases
Mutation causing called mutations
Cancer causing called carcinogens
What does Alkylating agents do?
Remove bases
What does Acridine agents do?
Add/ remove bases
What do x Rays do?
Break chromosomes/ delete few nucleotides
What does uv radiation do?
Create thymidine diners
Adjacent to T bases pair to each other by spontaneous PHOTO-REACTIVATION
What does IQ do?
Disrupts DNA base pairing by moving the, further apart therefore single base deletions at GC pairs as misreading by DNA polymerase
What is the mutation?
A change in a nucleic acid sequence, which can be the insertion or deletion of one or more nucleotides, or the rearrangement of several nucleotides
What is a wild type mutation?
Individual within a pop displays wild type trait, which is different to the trait most common in that population
Mutant phenotype
Mutant allele
Mutations in the germ line have possibility to be passed on hence GERM LINE MUTATIONS
DNA REPAIR- mismatch repair?
Enzyme detects nucleotide that doesn’t base pair in new DNA hence is incorrect base paired is excised or replaces. The detection of his is called PROOF READING
Post- replication
DNA REPAIR- excision repair?
Damaged DNA due to oxidation, alkalylated, uracil delaminated bases. DNA removed by excision of bass and replacement by DNA polymerase. The double strand is broken and chromosomes rearrangement occurs
DNA REPAIR- nucleotide excision?
Repair replaces up to 30 bases and used in uv damage repair and carcinogens.
Failure causes mismatch hence cancer.
Mismatch repair genes mutated - MLH1, MSH2 & MSH6
DNA REPAIR- base excision repair?
Replaces 1-5 bases and repairs oxidative damage
Caused by endogenous factors and ROS
What is promoted if the p53 monitor is damaged?
Apoptosis
What occurs if damaged DNA is not repaired?
PROPTOSIS
Senescence
Cancer
Disease
What are tumours?
Individual abnormal cells have lack of normal growth hence uncontrollable rapid cell growth
Generated by multi step process
Requires 6 mutations
All cells in tumour are same type
Behaviour of tumour depends on cell type
Chromosomal and microsatellite instability
What are oncogenes?
Genes that control cell division
Present in normal cells
Different classes
Stimulate or inhibit growth
What are tumour capabilities?
Ignore anti-growth signal
Avoid apoptosis
Divide without senescence
David independently of external growth signals
Stimulate sustained angiogenesis hence grow Invade tissue
What are tumour stressor genes?
Genes involved in protecting the steps against one step on the path to cancer
BRACA 1&2 are breast cancers regulating genes hence mutation would increase change of breast cancer so can screen
What is proto-oncogenes?
The genes normally present in cells, after mutation becomes oncogenes.
What type of pathogens can carry oncogenes?
Viruses can carry copies hence transform cells to cancerous
What does PCR do?
Amplifies DNA segments by repeated copying using thermo-stable DNA polymerase and pairs of primers to define a region. This allows the diagnosis of genetic diseases.
How is sickle cell detected?
Restriction site for enzyme MstII destroyed hence with gel electrophoresis the mutated genes will have one less DNA fragment as lacking site
Overview southern booting in mutation detection?
Used to analyse larger segments of DNA within or around a gene, used to analyse tucker repeat disorders such as huntingtons disease of fragile X syndrome
Talk about single strand confirmation polymorphism?
SSCO mutation scanning.
Identifies mutation
Targets DNA sequencing
Heterozygous for mutation hence PCR = mixture for normal and mutated DNA
Heated to denature DNA then cooled rapidly
Individual strands have sequence specfic partly double stranded
DNA electrophoresis in polyacrylamide gel and silver gel to detect SSCP
What is Array CGH?
Array Comparative Genomic Hybridisation
Used to screen sub-microscopic chromosomal deletions for which location can’t be deduced by patient phenotype
How does Array CGH work?
Array of DNA probes covering genome applied to surface of solid matrix
Patient DNA and normal control DNA labelled different coloured flourescent tags
Patient red and control green
Equal amounts of labelled DNA added then hybridised to probe array and hybridisation signals detected and compared
If normal DNA signal exceeds patients DNA, patient has deletion of chromosomal region. Green > red
What is MLPA used for?
Exon count
Ethical issues with genetic screening?
Used prenatally and in abortion cases
How is foetal DNA obtained?
Parental or sibling give sample of blood or saliva
Prenatal diagnosis uses fatal DNA from
Amniotic fluid cells but give 0.5% - 1% miscarriage risk
Chorion villus biopsy 2% risk
From maternal blood
