A. DNA and RNA Flashcards
How to produce a vaccine? (4)
1) Sequence the viral RNA.
2) Identify important parts of the virus for the immune system response.
3) Isolate those genes.
4) Use them as either mRNA which our cells will use to make viral proteins which we can then develop
an immune response to or put them into a new harmless virus to act as a vaccine.
What is Chronic Myeloid Leukaemia? (4)
- Cancer characterised by puss-filled blood.
- Caused by a translocation between chromosomes 9 and 22 written as t(9;22).
- The ABL gene breaks from chromosome 9 and the BCR gene breaks from chromosome 22 and they
fuse. - Chromosome 22, now containing BCR and ABL is known as the Philadelphia (Ph) chromosome.
- ABL is a tyrosine kinase:
an enzyme involved in cell signalling.
What is BCR-ABL?
- BCR-ABL is a functional but dysregulated tyrosine kinase and causes CML, and thus inhibiting this
tyrosine kinase (BCR-ABL) is the treatment for CML.
What is Imatinib? (2)
- Imatinib is a drug that inhibits BCR-ABL.
- Normally patients would die within 5 years of being diagnosed, but taking imatinib allows the CML
to be treated as a chronic disease. CML was changed from a fatal disease to a chronic one.
DNA:
- There are more than 10 million species on Earth, of which most are single-celled.
- In both single and multicellular organisms, the single cell contains all the ______ information
(the entire genome can be found in a single cell) of that species.
- This hereditary information is found in a _____ chemical called DNA (Deoxyribonucleic acid).
- DNA performs the same function regardless of ______.
hereditary
linear
What is the genome?
The genome comprises of all the genetic material that an organism possesses. It includes the DNA sequence of each chromosome, as well as any DNA in organelles.
- Remember that the entire genome can be found in just a single cell of the organism, except for mature red blood cells as these do not have a nucleus.
What is the function of the genome?
- The genome provides instructions to the body on how to live (maintain normal bodily processes - homeostasis) through RNA and protein synthesis and for reproduction.
What is the transcriptome?
The transcriptome is the complete set of RNAs present in a cell, tissue, or organism. Its complexity is mostly due to mRNAs, but it also includes noncoding RNAs. Note that not all DNA is transcribed.
What is the proteome?
The proteome is the complete set of proteins expressed by the entire genome. Some genes code for multiple proteins, and thus the size of the proteome is greater than the number of genes.
What is The Central Dogma of Gene Expression?
- Through the production of mRNA (transcription) and the synthesis of proteins (translation), the information contained in DNA is expressed.
What are Prokaryotes?
Have no distinct nucleus or specialized organelles.
What are Eukaryotes?
Have membrane-bound organelles – most notably a nucleus.
What is the origin of the eukaryotic cell?
The origin of the eukaryotic cell is the single most profound change in cellular organization during the evolution of life on Earth. We do not know how this happened, but the most probable reasoning is the Invagination theory. The cell membrane folded in on itself (invaginate) into the cytoplasm and established itself around the nuclear material of the cell, forming a nucleus and later an endoplasmic reticulum. A nucleus allows for greater specialization and diversity.
Prokaryotes vs Eukaryotes: (6)
What is the tree of life?
Bacterial genome arrangement: (2)
- Bacterial DNA is a circular molecule that is a few million nucleotides in length.
- To fit in the bacterial cell, it must be compacted 1000 times (1000-fold):
Firstly through formation of loop domains – compacts 10 times. Secondly through DNA super coiling within loops.
Bacterial genome arrangement: (2)
- Bacterial DNA is a circular molecule that is a few million nucleotides in length.
- To fit in the bacterial cell, it must be compacted 1000 times (1000-fold):
Firstly through formation of loop domains – compacts 10 times. Secondly through DNA super coiling within loops.
Eukaryotic genome arrangement: (5)
- To gain complexity, eukaryotic organisms gained more and more DNA, and circular DNA become linear to accommodate for more DNA.
- Eukaryotic species can contain one set of DNA (haploid) but usually contain two (haploid). Can contain more.
- All chromosomes are linear and are located in the nucleus and must thus be highly compacted to fit there.
- Each diploid cell contains about 2 meters of DNA.
- It is a problem to fit that much of DNA into a small space, and thus must be coiled 10 000 times.
What are histone proteins?
Histone Proteins – 8 histone proteins (two of each H2A, H2B, H3 and H4) come together and form a scaffold (backbone) around which DNA will coil. Histones are small, positively charged proteins.
How are histone proteins used to pair DNA?
- Since DNA is negatively charged (due to phosphate groups in its phosphate-sugar backbone), histones bind with DNA very tightly. DNA wraps twice around the 8 histones.
What are Nucleosomes?
- Nucleosomes- The basic unit of DNA packaging in eukaryotes. The unit of histone proteins plus its DNA wrapped around it is a nucleosome.
- A histone is a protein, which is a length of amino acids. The bulk of these amino acids from the
spherical protein, the amino terminal consisting of amino acids “waving around” are not part of the spherical core and can be called histone tails. The N-terminal tails of core histones protrude from the nucleosomes and can be modified enzymatically by biochemicals through: (3)
- Acetylation
- Phosphorylation
- Methylation
- The histone proteins are in intimate contact with the DNA and thus modifications on the histone
tails will have _____ effects on the DNA.
profound
How are adjacent nucleosomes joined by?
- Adjacent nucleosomes are joined by linker DNA which can be 8-114 base pairs (bps) long. This
makes it look like “beads on a string”. Nucleosomes are dynamic and thus can move its position along the DNA strand – it does not have a fixed position. The length of linker DNA between nucleosomes can change as the nucleosome moves. A given length of DNA anywhere in your genome may contain more or less nucleosomes at different points in time. The chromatin structure might be different due to the movement of the nucleosomes.
Packaging DNA into nucleosomes shortens it 7 times. 1 meter of DNA will become a 14 cm “string of beads”. Much more compaction is required and occurs as shown:
What is chromatin? (3)
- Eukaryotic DNA does not exist as naked DNA. It is always associated with histone proteins and other non-histone proteins.
- Chromatin refers to DNA and its associated proteins (histone proteins and non-histone proteins).
- Prokaryotes do not have chromatin as they do not have histones.
- There are two types of chromatin:
1) Heterochromatin
2) Euchromatin
What is Euchromatin?
2) Euchromatin – Stains lightly as it is less condensed. Nucleosomes are further apart from each
other, and the DNA is said to be in a relaxed conformation.
What is Heterochromatin?
1) Heterochromatin – Stains darkly as it is highly condensed. It is dispersed throughout nucleus but concentrated at nuclear envelope and only found in Eukaryotes. Nucleosomes are closer to each other and the length of linker DNA between nucleosomes is much shorter and the DNA is tightly compacted.
- _____ regions are called heterochromatic regions and _____ regions are called euchromatic regions.
Dark
Light
How do cells control access to their DNA? (2)
- Cells can control access to their DNA by modifying the structure of their chromatin. Highly compacted chromatin (heterochromatin) is not accessible to enzymes and transcription factors involved in DNA transcription. Genes occurring in these heterochromatic regions will be silenced.
- The opposite is true for euchromatic regions. Genes in euchromatic regions can be accessed by transcription factors and will thus be transcribed.
- Chromosome arrangement in the nucleus is not random.
How are chromosomes arranged in the 3-D nucleus? (3)
- Interphase chromosomes occupy a distinct part of the nuclear space known as chromosome
territories. - Inter and intra-chromosome interactions are specific and affect nuclear functioning, including gene
expression. - DNA-associated proteins (notably, histones) maintain order in the nucleus of eukaryotes.
What is a gene? (4)
- Made up of DNA.
- Basic physical unit of inheritance.
- Contain information needed to specify traits.
- Humans have approximately 22 000 genes.
DNA consists of two chains of polynucleotide chains in an antiparallel configuration (one strand is in the 5’ to 3’ direction and the other strand is in the 3’ to 5’ direction).
- 5’ =
- 3’ =
- 5’ = upstream (beginning of gene)
- 3’ = downstream (end of gene)
Components of a gene:
- Exons =
- Exons – Portion of a gene that codes for amino acids (protein). Usually short (100-200 bp in size).
Components of a gene:
- Introns =
- Introns – Non-coding regions of the gene between introns. Larger than exons (About 3000 bp long but length varies). Intron positions are usually conserved..
We call our genes “______ genes” because of introns and exons.
interrupted
- Regulatory sequences/ regions are found upstream of the gene and include ______ and promoters. They regulate transcription of a gene to mRNA.
enhancers
What are enhancers?
Enhancers – Activate the use of a promoter and by doing so control the efficiency and rate of transcription for that gene. They are usually found upstream but can be found downstream or anywhere along the gene. They must be present in the same strand of DNA as the gene being transcribed. They can also inhibit transcription by acting as Silencers. Due to their ability to either allow a gene to be transcribed or to block transcription, they are referred to the control region of a gene.
What are promoters?
Promoters – Special sequence that signals the start of the gene (TATA box). They are located upstream at the 5’ end and they bind transcription factors. They facilitate recruitment and binding of RNA Polymerase II and facilitate the start of transcription. It is a region recognised by the enhancer.
How do promoters and enhancers help start transcription?
How do promoters and enhancers help start transcription?
Transcription:
Initiation -
- Initiation – The signalling given by the promoter region to say that the transcription factors and RNA Polymerase should bind.
Transcription:
Elongation -
- Elongation – RNA Polymerase begins to add RNA nucleotides in the sequence the DNA specifies, therefore synthesizing the RNA molecule in the 5’ to 3’ direction. Transcription begins at the beginning of the first exon.
Transcription:
Termination -
- Termination – Terminator sequence signals for end of transcription. It is reached when the end of the last exon is transcribed. The RNA sequence formed is called pre-mRNA.
Transcription:
Termination -
- Termination – Terminator sequence signals for end of transcription. It is reached when the end of the last exon is transcribed. The RNA sequence formed is called pre-mRNA.
How is RNA processed? (4)
- The pre-mRNA formed is now processed and introns are removed (spliced out).
- Splicing is a post-translational modification that results in the removal of introns/intronic
sequences. Mature mRNA is formed, and it is ready for translation. - Conserved regions (regions that are usually always the same) of intron sequences are found
flanking the exon/intron boundaries and are called splice sites or splice junctions. 5’ splice sites are
found at the beginning of each intron and 3’ splice sites are found at the end of each intron. - 5’ splice sites begin with GT (eg GTA or GTG) and 3’ splice sites end with AG (eg CAG). Recognition
of these bases allow for the removal of introns. We say they are conserved because they remain the same and are thus easily recognisable. If there are changes to these sites, the intron may not be spliced out and this may have consequences in the process of translation and further on in the functioning of the protein that is supposed to be produced.
What is the process of translation?
- This is the decoding of mRNA and using its information to build a polypeptide (string of amino acids).
- mRNA is made up of codons (3 nucleotides).
- Specific start and end codons mark the beginning and end of the amino acid sequence.
- Start codons mark the initiation of translation (such as AUG/ATG).
- Stop codons mark the termination of translation (such as TAA, TAG, TGA or UAA, UAG, UGA). Stop
codons have no corresponding tRNA’s. - The start codon is not usually located on the first base of the first exon and the stop codon is not
usually located on the last base of the last codon. Thus, we have regions that have been transcribed
but will not be translated. - The region at the beginning of the first exon (before the start codon) that is transcribed but will not
be translated is called the 5’ UTR (untranslated region). - The region at the end of the last exon (after the stop codon) that has been transcribed but will not
be translated is called the 3’ UTR (untranslated region).
- Promoter –
- Start transcription at beginning of _____ and end at end of ______.
- Start translation at start codon ____ and end at stop codon _____.
- Exons in bold.
- Introns start with ___ and end with ____.
- 5’ UTR and 3’ UTR are regions translated but not transcribed.
TATA box
orange orange
ATG and TAA
GT and AG
When are nitrogenous bases called nucleosides?
- When nitrogenous bases are attached to the 1’ carbon, they are called Nucleosides. A Nucleoside combined with one or more phosphates is called
a Nucleotide. Nucleotides could also termed depending on the number of Phosphate groups they are attached to, such as Adenosine monophosphate (AMP), Adenosine diphosphate (ADP), Adenosine triphosphate (ATP) and so o
What is the process of semi-discontinuous and bi-directional replication?
- Recall that DNA is anti-parallel – the two strands are parallel to each other but in the opposite direction. One strand is in the 5’ to 3’ direction and the other is in the 3’ to 5’ direction.
- The enzyme responsible for DNA replication is DNA Polymerase (DNA is a polymer and Polymerase builds the polymer).
- DNA Polymerase can only read DNA in the 3’ to 5’ direction, and thus builds the new strand in the
5’ to 3’ direction (because the new DNA strand must be anti-parallel). Since the top strand is in the correct direction (3’ to 5’), the DNA Polymerase can build continuously, and this is called Continuous Replication. - The problem now arises at the bottom strand, which is in the 5’ to 3’ direction. DNA Polymerase must read it in reverse, but can only do so in short fragments, and these are known as Okazaki fragments, and this is called Discontinuous Replication.
- Hence, in both the top strand (leading strand) and the bottom strand (lagging strand), DNA is still being read in the 3’ to 5’ direction and synthesized in the 5’ to 3’ direction, but because they are being synthesized in opposite directions, we say it is bi-directional.
What is DNA Polymerisation? (3)
- The joining of two nucleotides is a condensation reaction that releases a stable molecule called Pyrophosphate (PPi – the i stands for inorganic). A proton (H+) is also released.
- A Phosphodiester bond is formed between the 5’ Phosphate group of one nucleotide and
3’ Hydroxyl group of another. - DNA is always synthesized in the 5’ to 3’ direction.
Replication Bubble:
How does the replication fork work? (5)
- In the leading strand, Primase synthesizes an RNA primer and DNA Polymerase binds and builds the
new strand from there. - In the lagging strand, Primase
synthesizes an RNA primer and DNA Polymerase binds and builds the first Okazaki Fragment. Primase then synthesizes a new RNA primer and DNA Polymerase builds the next Okazaki Fragment. The process is
repeated several times. - DNA ligase joins (ligates) the Okazaki Fragments
together to form the lagging strand. Phosphodiester bonds are formed. - Everything happens simultaneously. Primase synthesizes RNA Primer to start DNA replication, DNA Polymerase extends the strands forming the Okazaki fragment, and DNA Ligase joins the fragments.
- When the bond between the Phosphates is broken, the energy released is used to form the Phosphodiester between the incoming nucleotides and the existing chain.
How is DNA proofread during DNA synthesis?
- Transcription produces a sequence of ribonucleotides _______ to the sequence of
deoxyribonucleotides in one strand of DNA. - In DNA, the 5’ to 3’ strand is known as the coding/sense strand. This is the gene sequence used by geneticists or given in journals. The 3’ to 5’ strand
Is called the ______/_______ strand. - Only the template/antisense strand (3’ to 5’) is
used for______ (in DNA replication the template/antisense strand provides continuous replication, and the coding/sense strand provides discontinuous replication). - Therefore, the RNA produced will be complementary (have opposite base pairs) to the 3’ to 5’ strand and will be exactly the same as the 5’ to 3’ strand, except it will have _____ instead of Thymine.
Complementary
Template/Antisense strand
Transcription
Uracil
- The section of the gene between the
transcription start site and the transcription stop site is known as a transcription _____.
unit
Process of Transcription:
- Elongation = (4)
1) RNA Polymerase moves along the template/antisense DNA strand and helicase unwinds it, forming an elongation bubble. As with DNA replication, topoisomerase prevents the supercoiling of the DNA as the strands unwind.
2) Nucleotides are added to the 3’ OH of the growing RNA chain in a specific sequence (A pairs with U and C pairs with G) dictated by the template/antisense strand, and this is known as Watson-Crick Pairing.
3) The new RNA molecule (primary transcript) forms a short RNA-DNA hybrid molecule with the DNA template in the elongation bubble.
4) The RNA strand (like DNA in DNA replication) is synthesized in the 5’ to 3’ direction until a termination sequence is reached.
How does the process of transcription itself regulate transcription?
- Transcription is regulated through transcription factors – if they are not present, transcription will not occur.
- Control elements are present in the promoter (TATA and CAAT).
- Control regions called enhancers and silencers are present in the DNA sequence, and if bound, enhancers would increase transcription and silencers would inhibit transcription.
- Cis acting (they are part of the same DNA strand) enhancer regions are binding sites for activator proteins (which are proteins like transcription factors). The enhancer regions are usually far upstream or downstream of the transcription start site.
- Activators bind to both the enhancer and RNA Polymerase II, forming a DNA loop in the process.
- The binding of activators recruits general transcription factors, which help from a transcription
initiation complex with RNA Polymerase II, and we get increased transcription/gene expression of that gene. - Cis acting (they are part of the same DNA strand) silencer regions can be found far upstream or downstream of the transcription start site. Silencer regions are binding sites for repressor proteins (which are proteins like transcription factors).
- The repressor proteins then binds to both transcription factors and RNA Polymerase II at the promoter forming a DNA loop.
- The repressor proteins then inhibit the RNA Polymerase II and inhibits the binding transcription factors and hence inhibit or silence transcription from that gene.
How is mRNA processed? (6)
- In mammalian cells, mRNA is transcribed as a precursor molecule called pre-mRNA. This is simply mRNA with both introns and exons.
- The pre-mRNA is modified.
- The 5’ end is capped. 7methylguanosine triphosphate is attached to it. This allows recognition of
the ribosome, processing of the pre-mRNA and for it to be exported out of the nucleus. - The 3’ hydroxyl end is polyadenylated (20-250 ATP’s are attached to it). This helps to regulate the
half-life of mRNA. The longer the polyadenylated tail, the longer the mRNA will survive. - mRNA is then cleaved and spliced to remove introns. This occurs inside the nucleus on particles
called spliceosomes by snRNA (small nuclear RNA) and associated proteins forming snRNPS (small
nuclear ribonuclear proteins). Spicing occurs at spice junctions (intron/exon boundaries). - We have sequence conservation (sequences that are usually the same and thus easily recognised)
at intron/exon boundaries – The end of an intron may have an AG and the beginning of the following intron may have GU.
How is mRNA processed? (6)
- In mammalian cells, mRNA is transcribed as a precursor molecule called pre-mRNA. This is simply mRNA with both introns and exons.
- The pre-mRNA is modified.
- The 5’ end is capped. 7methylguanosine triphosphate is attached to it. This allows recognition of
the ribosome, processing of the pre-mRNA and for it to be exported out of the nucleus. - The 3’ hydroxyl end is polyadenylated (20-250 ATP’s are attached to it). This helps to regulate the
half-life of mRNA. The longer the polyadenylated tail, the longer the mRNA will survive. - mRNA is then cleaved and spliced to remove introns. This occurs inside the nucleus on particles
called spliceosomes by snRNA (small nuclear RNA) and associated proteins forming snRNPS (small
nuclear ribonuclear proteins). Spicing occurs at spice junctions (intron/exon boundaries). - We have sequence conservation (sequences that are usually the same and thus easily recognised)
at intron/exon boundaries – The end of an intron may have an AG and the beginning of the following intron may have GU.
What is alternative splicing? (2)
- By splicing at different points instead of all the intron/exon boundaries, we can derive numerous mature mRNA’s from a single pre-mRNA.
- This is how humans only have about 22 000 genes but are the most complex organisms. One gene has many functions due to alternative splicing. Other organisms have far more genes than humans but are far less complex. The number of genes in an organism is not an indication of complexity.
- tRNA has 4 arms:
- The ribosome is made up of two major subunits: A ___S and a ___S subunit.
40
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