Cell biology theme 4 Flashcards
What is the genome ?
the total genetic information carried by a cell or an organism
How much of the human genome codes for proteins ?
2%
How long is the human genome ?
3.2 x 10^9
How long is the mitochondrial genome ?`
17,000 bp
How long is the bacterial genome ?
4.6 million bp
What can comparison of genome sequences tell us ?
there are exon sequences that are highly conserved
some intron sequences can be conserved - not known why
What is special about mammals and fish ?
they diverged millions of years ago
this would have obliterated similar sequences but there are conserved genes and pieces of regulatory sequences
why can’t genome sequencing predict the biology if an organism ?
we dont know which sequences are genes
What are open reading sequences ?
sequences that are over 100 codons that lack a stop codon
what is an example of a start codon ?
AUG
What are examples of stop codons ?
TAA TAG and TGA
How can you test whether a sequence is a gene ?
isolate the sequence
place into the bacterial plasmid
test if its functional
What are distinguishing features of genes ?
they have splice site sequences - at intron/exon boundaries
they have promoters
codon bias
what are pseudogenes ?
nucleotide sequences resemble genes but have a mutation that prevents their expression
How can we use mRNA to sequence genes ?
collect mRNA and then reverse transcribe into DNA
map this back to DNA to get info on extragenic sequences and introns
but this is hard as most mRNA is non coding
What is the significance of homologues ?
a homologue is an organism with a high degree of similarity
if we know the function of a protein in another organism we can infer that the organism has a similar sequence
What is an ortholog ?
genes in different species that have evolved from a common ancestral gene. Identification of orthologs is critical for reliable production of gene function.
Are genes evenly distributed or not ?
no they are not evenly distributed in the genome
what have mobile genetic elements done ?
transposons and retrotransposons have multiplied and replicated themselves adding new copies in differnet positions
What are unique sequences ?
gene regulatory sequences
RNA sequences
unknown sequences
What do non protein coding sequences code for ?
microRNA and long non coding RNAs
How does miRNA work ?
it controls gene expression
base pairing with specific mRNAs
reduces their stability and ability to translate into a protein. mRNA is destroyed into nucleases
What is the RNA induced silencing complex ?
Double stranded siRNAs are taken up to RISCs
one strand is discarded and the other is used to bind to target RNA
How do long non coding RNAs work ?
they coat the chromosome and attract enzymes in chromatin remodelling complexes this alters gene expression as highly condensed chromatin forms
What are regulatory DNA sequences ?
promoters
enhancers
transcriptional regulators
What are epigentic changes ?
changes in gene expression that do not involve changes in the DNA sequence
What is methylation of cytosine bases ?
this can lead to chromatin remodelling complexes
this stops transcriptional regulators from working
switches genes/off
What is histone modification of acetyl groups ?
acetyl groups are transferred to histone proteins
this leads to gene regulation
Are epigenetic changes heritable ?
yes they can be inherited through the germ line
What is in-vitro testing ?
in glass and plastic
add gene cloning and gene editing techniques
however might not be as accurate as cells are out of context
What are GWAS ?
Genome wide association studies
can help us identify polymorphisms and DNA variants associated with diseases
What was GWAS initially focussed on ?
SNPs to search for genes that predispose individuals to common diseases such as Diabetes
What has been associated with LOF mutations ?
schizophrenia and autism - they arose in the germ line which is why they are still common
What was the study with age related macular degeneration ?
comparison of SNPs in populations found one SNP was more present. It was located in the Cfh gene but in an intron. therefore there was no effect on the protein. further research found 3 more variants. one caused a sub of amino acids,
What is empirical medicine ?
has developed from experience and observation eg.vaccines
What is individualised medicine ?
cancer treatment and vaccines which are made fro specific genotypes
What is stratified medicine ?
dividing the population into groups depending on which SNPs they have - some individuals might have SNPs for drug metabolising enzymes - cant metabolise the drug. Some might benefit some might not
What is herceptin ?
a monoclonal antibody against HER2 a receptor tyrosine kinase
What is the link between the ERBB2 gene and HER2 ?
amplification of the ERBB2 gene that encodes HER2 is found in some early stage breast cancers - therefore herceptin is only effective in cancers with the amplification
What is performed to ensure the herceptin will be effective ?
an immunohistochemical test for HER2 is performed on a biopsied tissue sample - makes sure it only works on patients.
What immunohistochemical tests can be performed to see the presence of The gene amplification ?
the RTK is stained in a protein test in IHC
in FISH the gene that produces mRNA for the RTK can be identified.
What are biomarkers ?
proteins/genes that give us an idea of the risk for or aid in the diagnosis of a disease - they allow the personalisation of medicine.
What are diagnostic biomarkers ?
diagnose a specific disease as early as possible
What are susceptibility/risk biomarkers ?
tells us about the personal risk an individual has of developing a disease - the ERBB2 mutation can be detected in parents/siblings
What is a prognostic biomarker ?
tellus about the evolution of a disease , how far and advanced it is
What is a predictive biomarker ?
.help us predict the response to a treatment based on toxicity
What is genetic testing based on and what os it used for ?
it is used for the targeted amplification of a specific mutation in a gene and detection and this is done by PCR.
What else can be genetically tested for ?
testing for any mutation in a specific gene - sequence the gene and make comparisons
What would require whole genome analysis ?
looking for any mutation in any gene
What is pyrosequencing ?
a method of sequencing based on ‘sequencing by synthesis’ - relies on the detection of pyrophosphate release on nucleotide incorporation,
What is ARMS ?
amplification refractory mutation system
allele specific primers are used in PCR to discriminate between normal/immune sequences.
What is gene therapy ?
direct genetic modification to achieve a therapeutic goal
What is germ line therapy ?
alters all the cells in an individual
passed on to the next generation
involves the modification of a gamete , zygote or an early embryo
What is somatic gene therapy ?
affects only some cells and is not transmitted on to offspring
Which cells are targeted in somatic gene therapy ?
cells that are often directly involved in pathogenic processes
in cancer gene therapy - normal immune cells are altered to invoke a powerful immune response
What are the 4 ways of modifying somatic cells ?
gene augmentation
gene mutation correction
gene expression inhibition
killing of diseased cells
What is gene augmentation ?
supplying a functional gene copy to supplement a defective gene
they can be used in diseases where there is gene not working
can be used in cancer therapy to replace defective tumour suppressor genes
What is gene mutation correction ?
when a mutated gene is restored with a normal sequence
required in GOF mutations
can induce splicing skipping of a harmful exon
What is gene expression inhibition ?
used to target essential genes in mutant cells
for example silence activated oncogenes in cancer cells
or silencing a GOF mutant allele
this can be done with RNA silencing
How can we kill specific diseased cells ?
using genes can encode toxins
a prodrug metabolising gene can be added to a cell to make it vulnerable to the effects of a prodrug added later.
What is transduction ?
using viral based vectors (viruses are more effective in infecting cells , inserting their genomes and getting their genes expressed for a long time)
What is transfection ?
using non viral vectors to transfer RNA and DNA
does not involve genome integration and have low efficiencies
examples of cells used in transfection ?
liposomes which bind to cell membranes and enter by endocytosis
What is ex-vivo ?
cells that are taken , grown in culture and modified by transfer of nucleic acid
cells that have taken up the modification are amplified and injected into the patient
this is appropriate where the target cells are accessible fro removal
What is in-vivo ?
gene transfer is carried in situ
this is done when the target cells arent accessible , cannot be reimplanted successfully or cant be cultured in great numbers.
