From Genotype to Phenotype Flashcards
Define
Single nucleotide polymorphism (SNP)
a DNA sequence variation occurring when a single nucleotide adenine (A), thymine (T), cytosine (C), or guanine (G]) in the genome (or other shared sequence) differs between members of a species or paired chromosomes in an individual
Definition
a prokaryotic immune system that confers resistance to foreign genetic elements such as those present within plasmids and phages that provides a form of acquired immunity
CRISPR/Cas
How do SNP arrays work?
- A “gene chip” contains thousands of spots, each containing single-stranded 25 base reference DNA molecules (oligonucleotides)
- Each reference DNA is complementary to an SNP allele
- Oligonucleotides are printed onto the chip, or synthesised directly on it
- Genomic DNA to be tested is fragmented, amplified as a single strand, labelled, then put on the chip
- Binding conditinos favour perfet matching between probe DNA and chip DNA
What three major classes of RNAs make up the eukaryotic transcriptome?
- Ribosomal RNAs (rRNA)
- Protein-encoding RNAs (mRNA)
- Small RNAs (including tRNA)
Define
Polymorphism
a DNA variation present in more than 1% of people
Definition
a molecular biology term for an insertion or deletion of bases in the genome of an organism
INDELs
Define
Haplotype
the unique combination of alleles that makes up an individual
Define
Orthologues
Any of two or more homologous gene sequences found in different species related by linear descent
Define
Complementary DNA (cDNA)
DNA synthesized from a single-stranded RNA (e.g., messenger RNA (mRNA) or microRNA) template in a reaction catalyzed by the enzyme reverse transcriptase
How does silencing gene expression through RNAi work?
- dsRNA is formed in the cytoplasm
- dsRNA is cleaved into 21-23 nt short interfering (si)RNA duplexes by Dicer enzyme
- siRNAs are incorporated into multiprotein RNA-inducing silencing complex (RISC)
- siRNA is unwound - antisense strand remains associated with RISC and base-pairs with target mRNA
- RISC cleaves and inactivates target mRNA. Expression is reduced but not completely abolished
Define
Allele
an alternative form of a genetic locus
Why is gene identification in eukaryotic genomes more difficult than gene identification in prokaryotes?
- Density of genes varies
- Genes are concentrated in G/C rich areas of the genome
- < 2% of human genome encodes protein or ncRNA
- ~25% of human genome is regulatory/introns
- Rest of the genome is scaffolding or “junk DNA”
- Promotor sequences are not easily recognisable
- Regulatory factors bind enhancers far form the transcription initiation site
- Most genes are interrupted (introns)
- Exons (ORFs) can be very small (< 600 codons)
- Particular exons do not always appear in mRNA (alternative splicing)
- Some genes encode microRNAs (small and not translated)
True or False:
Each EST only contains exons
True
Define
Isoforms
any of two or more functionally similar proteins that have a similar but not identical amino acid sequence and are either encoded by different genes or by RNA transcripts from the same gene which have had different exons removed
Most RNA is highly susceptible to _____________, so it is very difficult to purify and maintain an RNA molecule in an intact state
Most RNA is highly susceptible to nucleases, so it is very difficult to purify and maintain an RNA molecule in an intact state
Definition
Any of two or more homologous gene sequences found in different species related by linear descent
Orthologues
What do we need to know about each gene? How do we obtain this information?
For each gene we need to know:
- Where and when it is transcribed into RNA
- How it is spliced, and how many spliceoforms there are
- Whether particular spliceoforms are restricted to particular cells or growth stage
This information cannot be directly deduced from genomic sequence with confidence. We rely on analysis of complementary DNA (cDNA) and Expression Sequence Tages (ESTs), derived from RNA.
Definition
an approach used to identify genes (or set of genes) responsible for a particular phenotype of an organism
Forward genetics
Definition
the polymerase that only transcribes ribosomal RNA (but not 5S rRNA), a type of RNA that accounts for over 50% of the total RNA synthesized in a cell
RNA polymerase I
________ genetic analysis can be applied to specifically mutate the gene in question
Reverse genetic analysis can be applied to specifically mutate the gene in question
Definition
the complete collection of RNA produced from a genome
Transcriptome
True or False:
Each EST is a small segment of mRNA generated by reverse transcriptase
False
What can be concluded if a DNA sequence from a genome matches exactly to a specific EST?
It can be concluded that the genomic DNA is transcribed and that it represents a gene (or gene fragment)
What is the first step to understanding gene function?
Determine where and when the gene is expressed
Definition
short sub-sequence of a cDNA sequence. They may be used to identify gene transcripts, and are instrumental in gene discovery and in gene-sequence determination
Expression sequence tags (ESTs)
True or False:
Each EST has a 5’ end matching the transciptional start point of its gene
False
Definition
the study of thousands of RNAs, simultaneously
Transcriptomics
Definition
the general concept of aiming for high number of unique reads of each region of a sequence
Deep sequencing
Definition
the occurrence in members of a population of combinations of linked genes in non-random proportions
Linkage disequilibrium
90% of human genome variation is in the form of _____
90% of human genome variation is in the form of SNPs
_________ are cDNAs made from mRNAs originating from a specific cell or tissue
ESTs are cDNAs made from mRNAs originating from a specific cell or tissue
What RNA polymerase transcribes rRNA?
RNA polymerase I
Definition
a multiprotein complex. It is one of the three RNAP enzymes found in the nucleus of eukaryotic cells. It catalyzes the transcription of DNA to synthesize precursors of mRNA and most snRNA and microRNA
RNA polymerase II
Inter-species comparisons reveal segment boundaries, and _________ regions where orthologous genes are likely to be located
Inter-species comparisons reveal segment boundaries, and syntenic regions where orthologous genes are likely to be located
How is protein expression detected?
- Antibodies or other binding reagent (cell/tissue structure can be maintained)
- Enzyme activity (usually in cell or fluid extracts)
- Mass spectrometry/proteomics (usually in cll or fluid extracts)
What do all mRNAs have at their 3’ ends?
A stretch of adenines
Define
RNA polymerase II
a multiprotein complex. It is one of the three RNAP enzymes found in the nucleus of eukaryotic cells. It catalyzes the transcription of DNA to synthesize precursors of mRNA and most snRNA and microRNA
Define
Forward genetics
an approach used to identify genes (or set of genes) responsible for a particular phenotype of an organism
What are the advantages or CRISPR/Cas editing?
- Can be used in any organism where IVF technology exists
- CRISPR knockout mice can be generated in ~ 3-6 months, compared to 18 months for conventional mouse knockout
- Both alleles can be mutated simultaneously to make homozygous deficient animals
- Precise changes can be made
- Up to five genes have been edited at once in mice
- Can be adapted to global functioning genomic/network studies
Definition
transcribes DNA to synthesize ribosomal 5S rRNA, tRNA and other small RNAs
RNA polymerase III
How is rRNA modified after transcription?
rRNAs are cut out from a longer precursor (pre-rRNA)
Definition
a DNA variation present in more than 1% of people
Polymorphism
Define
Expression sequence tags (ESTs)
short sub-sequence of a cDNA sequence. They may be used to identify gene transcripts, and are instrumental in gene discovery and in gene-sequence determination
Genomes from different species can be compared to identify _______ genes that lie in _________chromosomal regions
Genomes from different species can be compared to identify orthologous genes that lie in syntenic chromosomal regions
How many different mRNAs can be produced from this precursor if any splice donor can interact with any splice acceptor?
4
A-B-C-D
A-C-D
A-B-D
A-D
Predicting susceptibility profiles for broad range of disease or treatments for a particular individual requires what?
- A reference map of SNPs
- Developing rapid (and cheap) screening methods to map at least 10,000 of these SNPs in a patint
What genetic information does SNP analysis not provide about an individual?
Variations not represented in SNP arrays, like INDELs and mobile elements
Theoretically, a SNP could have ___ possible alleles, but most SNPs have only __ alleles
Theoretically, a SNP could have 4 possible alleles, but most SNPs have only 2 alleles
What RNA polymerase transcribes mRNA?
RNA polymerase II
How are false positives reduced in SNP arrays?
Each SNP position is represented by up to forty different but overlapping DNA sequences
Define
RNA interference (RNAi)
a biological process in which RNA molecules inhibit gene expression or translation, by neutralizing targeted mRNA molecules
The number of clones containing the same EST in one library is proportional to what?
The transcriptional activity of the gene
True or False:
The order of orthologous genes is conserved in syntenic blocks
Definition
the branch of pharmacology concerned with the effect of genetic factors on reactions to drugs
Pharmacogenetics
True or False:
ESTs will only match exons
True
Definition
a laboratory mouse in which researchers have inactivated, or “knocked out,” an existing gene by replacing it or disrupting it with an artificial piece of DNA.
Knockout mice
Define
RNA polymerase I
the polymerase that only transcribes ribosomal RNA (but not 5S rRNA), a type of RNA that accounts for over 50% of the total RNA synthesized in a cell
Definition
a genomic technique for sequencing all of the protein-coding region of genes in a genome (known as the exome)
Whole exome sequencing (WES)
The point at which an EST to genomic DNA match ends can indicate what?
An intro/exon boundary or transcriptional start/end point
Define
Mutation
a sequence present in less than 0.1% of people
True or False:
Each EST represents genes active in every cell
False
Define
Whole exome sequencing (WES)
a genomic technique for sequencing all of the protein-coding region of genes in a genome (known as the exome)
How does in situ hybridsation work?
Labelled DNA or RNA complementary to target mRNA is soaked into cell or tissue, treated to minimise disruption to structure.
Probe will bind specifically to the target mRNA and identify where it is being produced
What enzyme allows DNA to be produced from RNA?
Definition
the unique combination of alleles that makes up an individual
Haplotype
Define
SNP microarrays
a type of DNA microarray which is used to detect polymorphisms within a population
Definition
analyzes the phenotype of an organism following the disruption of a known gene
Reverse genetics
Definition
any of two or more functionally similar proteins that have a similar but not identical amino acid sequence and are either encoded by different genes or by RNA transcripts from the same gene which have had different exons removed
Isoforms
How can transcripts from a single gene be assessed?
in situ hybridisation on intact cells or tissue
Reverse transcriptase PCR, and real time quantitative PCR
Why are there different numbers of read for different parts of mRNA?
Alternative splicing removes exon 4 from some RNAS
What accounts for the majority of human sequence variation?
SNPs
Chromosomes recombine segments called ________ blocks
Chromosomes recombine segments called haplotype blocks
Define
Reverse genetics
analyzes the phenotype of an organism following the disruption of a known gene
Define
RNA polymerase III
transcribes DNA to synthesize ribosomal 5S rRNA, tRNA and other small RNAs
Definition
a biological process in which RNA molecules inhibit gene expression or translation, by neutralizing targeted mRNA molecules
RNA interference (RNAi)
Definition
either of a pair of genes that derives from the same ancestral gene. (genomics) a pair of genes that derives from the same ancestral gene and now reside at different locations within the same genome
Paralogues
Definition
a sequence present in less than 0.1% of people
Mutation
How does reverse transcriptase PCR work?
What RNA polymerase transcribes small RNAs?
RNA polymerase III
________ of the gene can be assessed in specific cells, most recently via deep sequencing analyisis
Transcription of the gene can be assessed in specific cells, most recently via deep sequencing analyisis
Definition
a DNA sequence variation occurring when a single nucleotide adenine (A), thymine (T), cytosine (C), or guanine (G]) in the genome (or other shared sequence) differs between members of a species or paired chromosomes in an individual
Single nucleotide polymorphism (SNP)
Definition
DNA synthesized from a single-stranded RNA (e.g., messenger RNA (mRNA) or microRNA) template in a reaction catalyzed by the enzyme reverse transcriptase
Complementary DNA (cDNA)
Define
Paralogues
either of a pair of genes that derives from the same ancestral gene. (genomics) a pair of genes that derives from the same ancestral gene and now reside at different locations within the same genome
Define
CRISPR/Cas
a prokaryotic immune system that confers resistance to foreign genetic elements such as those present within plasmids and phages that provides a form of acquired immunity
In the older knockout mouse technology ________ are manipulated in the laboratory to mutate one ____ of the gene of interest, then the mutant cell are injected into a blastocyst, which is transferred into a foster mother. Resulting chimeric mice that can transmit the mutation to offspring become the founders of a knockout line, which can be examined for changes to ________
In the older knockout mouse technology embryonic stem cells are manipulated in the laboratory to mutate one allele of the gene of interest, then the mutant cell are injected into a blastocyst, which is transferred into a foster mother. Resulting chimeric mice that can transmit the mutation to offspring become the founders of a knockout line, which can be examined for changes to phenotype
Define
Alternative splicing
A mechanism by which different forms of mature mRNAs (messengers RNAs) are generated from the same gene
Why can relatively few genes in the genome give rise to a vastly greater number of proteins?
Alternative splicing
Define
Pharmacogenetics
the branch of pharmacology concerned with the effect of genetic factors on reactions to drugs
Definition
DNA sequences that can move around the genome, changing their number of copies or simply changing their location, often affecting the activity of nearby genes
Mobile genetic elements
How can transcripts from a many genes be assessed?
Microarray analysis
RNA deep sequencing
Define
Transcriptomics
the study of thousands of RNAs, simultaneously
Definition
an alternative form of a genetic locus
Allele
Related but different polypetides can be generated from the same primary transcript through which process?
Alternative splicing
Define
INDELs
a molecular biology term for an insertion or deletion of bases in the genome of an organism
Define
Linkage disequilibrium
the occurrence in members of a population of combinations of linked genes in non-random proportions
Define
Mobile genetic elements
DNA sequences that can move around the genome, changing their number of copies or simply changing their location, often affecting the activity of nearby genes
What can you conclude about the abundance of this mRNA in the liver?
About 50% is alternatively spliced
Define
Transcriptome
the complete collection of RNA produced from a genome
What can make identifying orthologs difficult?
- Orthologous genes may be on different chromosomes in different species
- There may be a number of similar (paralogues) in the genome
Definition
A mechanism by which different forms of mature mRNAs (messengers RNAs) are generated from the same gene
Alternative splicing
Any two people picked at random show > 97% nucleotide identity in their genomes. How do variations arise?
- Large-scale or small-scale INDELs
- Differing numbers or positions of mobile genetic elements
- Single nucleotide polymorphisms (SNPs)
Define
Knockout mice
a laboratory mouse in which researchers have inactivated, or “knocked out,” an existing gene by replacing it or disrupting it with an artificial piece of DNA.
Splicing errors are a common result of mutation. How can they cause disease?
- Mutations can occur in splice donor or acceptor sequences, or can generate new (cryptic) splice sequences
- Exons can be ommited, deleting a section of protein, severly affecting structure
- Use of false (cryptic) acceptor or donor sites can truncate or lengthen exons, severly affecting protein structure
Define
Deep sequencing
the general concept of aiming for high number of unique reads of each region of a sequence
Recently the CRISPR/Cas gene _______ system developed from bacteria has revolutionized _______ genetic analysis. This involved short _________ sequences that bind both the target DNA and ________ which makes a double strand break in the DNA. Repair of the break by the cell’s machinery often introduces a short __________ that mutates the gene
Recently the CRISPR/Cas gene editing system developed from bacteria has revolutionized reverse genetic analysis. This involved short RNA sequences that bind both the target DNA and endonuclease which makes a double strand break in the DNA. Repair of the break by the cell’s machinery often introduces a short deletion that mutates the gene
Definition
a type of DNA microarray which is used to detect polymorphisms within a population
SNP microarrays
Which of the following SNPs is most likely to alter gene expression?
Select one:
Synonymous SNPs
Nonsense SNPs
Missense SNPs
Non-coding region SNPs
Which of the following SNPs is most likely to alter gene expression?
Select one:
Synonymous SNPs
Nonsense SNPs
Missense SNPs
Non-coding region SNPs
Forward genetics describes a process of discovery of the relationship between gene and phenotype…
Select one:
by comparing the function of corresponding genes in two species
by identifying the gene responsible for a particular phenotype
by specifically mutating a gene and analysing the resulting phenotype
by introducing a gene into cultured cells and analysing the resulting phenotype
Forward genetics describes a process of discovery of the relationship between gene and phenotype…
Select one:
by comparing the function of corresponding genes in two species
by identifying the gene responsible for a particular phenotype
by specifically mutating a gene and analysing the resulting phenotype
by introducing a gene into cultured cells and analysing the resulting phenotype
Whole exome sequencing differs from whole genome sequencing in that…
Select one:
Whole exon sequencing focuses mostly on the protein coding regions of the genome
Whole exon sequencing focuses mostly on the non-coding regions of the genome
Whole exon sequencing focuses specifically on identifying indels and SNPs in individuals
Whole exon sequencing focuses specifically on identifying indels and SNPs in populations
Whole exome sequencing differs from whole genome sequencing in that…
Select one:
Whole exon sequencing focuses mostly on the protein coding regions of the genome
Whole exon sequencing focuses mostly on the non-coding regions of the genome
Whole exon sequencing focuses specifically on identifying indels and SNPs in individuals
Whole exon sequencing focuses specifically on identifying indels and SNPs in populations
The transcriptome of a cell…
Select one:
represents the total nuclear and mitochondrial DNA content of the cell
represents all genes ever expressed in that cell
represents only the genes transcribed at the time of sampling
represents only mRNA
The transcriptome of a cell…
Select one:
represents the total nuclear and mitochondrial DNA content of the cell
represents all genes ever expressed in that cell
represents only the genes transcribed at the time of sampling
represents only mRNA
One of the greatest advantages of RNA-sequencing over microarrays is that…
Select one:
RNA sequencing can identify SNPs
RNA sequencing can measure expression across every known gene
RNA sequencing is much faster
RNA sequencing can measure when genes are expressed
One of the greatest advantages of RNA-sequencing over microarrays is that…
Select one:
RNA sequencing can identify SNPs
RNA sequencing can measure expression across every known gene
RNA sequencing is much faster
RNA sequencing can measure when genes are expressed
CpG islands usually mark regions of the genome where…
Select one:
transcription is occurring
transcription is silenced
replication is initiated
replication is terminated
CpG islands usually mark regions of the genome where…
Select one:
transcription is occurring
transcription is silenced
replication is initiated
replication is terminated
Which of the following statements about single nucleotide polymorphisms (SNPs) is FALSE?
Select one:
SNPs are found in all sequenced mammalian genomes
SNPs are concentrated in CpG islands
SNPs are the greatest source of variation between individual genomes
SNPs may contribute to disease
Which of the following statements about single nucleotide polymorphisms (SNPs) is FALSE?
Select one:
SNPs are found in all sequenced mammalian genomes
SNPs are concentrated in CpG islands
SNPs are the greatest source of variation between individual genomes
SNPs may contribute to disease
Sickle cell anaemia is an inherited blood disorder associated with a single nucleotide polymorphism (SNP) designated rs334 in the human beta globin (HBB) gene. Individuals with sickle cell anaemia are about the 10-fold more resistant to life-threatening forms of malaria. The most common allele is rs334(A), encoding the normal form of haemoglobin. Individuals homozygous for the rs334(T) allele exhibit the sickling form of hemoglobin, Hb-S.
Shown below are the maternal and paternal HBB gene sequences from 4 individuals being assessed for sickle cell anaemia with the SNP indicated in bold. The coding sequence commences at nucleotide position 25.
In terms of its effect on the HBB coding sequence the rs334(T) allele can be best described as…
Select one:
sense
nonsense
missense
antisense
In terms of its effect on the HBB coding sequence the rs334(T) allele can be best described as…
Select one:
sense
nonsense
missense
antisense
Sickle cell anaemia is an inherited blood disorder associated with a single nucleotide polymorphism (SNP) designated rs334 in the human beta globin (HBB) gene. Individuals with sickle cell anaemia are about the 10-fold more resistant to life-threatening forms of malaria. The most common allele is rs334(A), encoding the normal form of haemoglobin. Individuals homozygous for the rs334(T) allele exhibit the sickling form of hemoglobin, Hb-S.
Shown below are the maternal and paternal HBB gene sequences from 4 individuals being assessed for sickle cell anaemia with the SNP indicated in bold. The coding sequence commences at nucleotide position 25.
A new rs334 (G) allele was recently discovered, which would vary the codon to…
Select one:
glutamic acid
glycine
valine
glutamine
A new rs334 (G) allele was recently discovered, which would vary the codon to…
Select one:
glutamic acid
glycine
valine
glutamine
