Molecular Biology: DNA, RNA, and Protein Flashcards
What is a genome?
- Every organism has a genome that contains the information needed to make and maintain itself
- Most genomes are made of DNA but some viruses have an RNA genome
- The human genome has 3,235,000,000 base pairs of DNA
- We also have a mitochondrial genome that is 16,569 base pairs of DNA
- Can be over 1000 mitochondria per human cell and each mitochondria can have 5 to 10 copies of mitochondrial DNA
What is a chromosome?
- The human genome has 3,235,000,000 base pairs of DNA
- In eukaryotic organisms, the DNA is not a single long chain
- Instead the DNA is split up into different sizes of linear molecules that condense to form a chromosome
This is a karyotype of which organism?
Diploid organism (in this case a bull)
29 autosomal chromosomes x 2 copies
2 sex chromosomes (X and Y)
Total = 60 chromosomes
How many chromosomes does each organism have?
39 (1n); 78 (2n) = chicken
23 (1n); 46 (2n) = human
52 (1n); 104 (2n) = catfish
1n = 1 set (copy) of chromosomes (haploid)
2n = 2 sets (copies) of chromosomes (diploid)
All organisms have DNA made from the same molecules.
True or False?
True.
DNA =
Deoxyribonucleic acid
Genetic material is a long double stranded DNA polymer.
The sequence of units in DNA are called […].
deoxyribonucleotides
The chemical composition of deoxyribonucleotides is the same in all organisms.
True or False?
True.
This is one reason why molecular biologists can transfer genes
amongst organisms
Describe the structure of nucleotide.
Pyrimidines [2].
- Cytosine
- Thymine (uracil in RNA)
Purines [2].
- Adenine
- Guanine
- Cytosine
- Thymine (Uracil in RNA)
Pyrimidines [2].
- Adenine
- Guanine
Purines [2].
How are nucleotides abbreviated?
- Abbreviations of the four nucleotides are dATP, dCTP, dGTP, dTTP
- For example, dATP = 2ʹ-deoxyadenosine 5ʹ-triphosphate
Nucleotide subunits are joined by […].
- Nucleotide subunits are joined by phosphodiester bonds.
Describe the chemical structure of a single strand of DNA.
- The phosphate group of the 5’-carbon of one nucleotide is linked to the 3’-OH group of the deoxyribose of the adjacent nucleotide.
- The polynucleotide strand has a 3’-OH group at one end and a 5’-phosphate group at the other end.
- The nucleotide subunits are joined by phosphodiester bonds.
How do nucleotides join together to make DNA?
- DNA synthesis occurs in the 5’ to 3’ direction.
- An enzyme called DNA polymerase adds nucleotides to the DNA strand.
- The alpha-phosphate of the nucleotide gets incorporated into the DNA strand and the beta-gamma phosphates (called pyrophosphates) are removed.
Describe the structure of a double strand of DNA.
- Antiparallel
- C pairs with G (3 h-bonds)
- T pairs with A (2 h-bonds)
Adenine and thymine bond via 2 hydrogen bonds.
True or False?
True.
Adenine and thymine bond via 3 hydrogen bonds.
True or False?
False.
They bond by only 2.
Cytosine and guanine bond via 2 hydrogen bonds.
True or False?
False.
They bond via 3.
Cytosine and guanine bond via 3 hydrogen bonds.
True or False?
True.
Think C3PO. (3 bonds goes with cytosine)
How much DNA do organisms have?
- A thousand base pairs = kilobase pairs or Kb
- A million base pairs = megabase pairs or Mb
- A billion base pairs = gigabase pairs or Gb
What are the major functions of DNA? [4]
- Encodes the information for the production of proteins
- It is reproduced (replicated) with a high degree of accuracy to pass the encoded information onto new cells.
- DNA single-strand can act as a template for the production of a new complementary strand.
- Sequence of nucleotides in a gene provides the code for the production of a protein.
Watson-Crick-Franklin model of DNA fully meets these requirements.
What is the complementary DNA strand?
5’-TAGGCAT-3’
3’-ATCCGTA-5’
How do RNA molecules differ from DNA? [3]
- Sugar is ribose instead of deoxyribose
- Instead of thymine (T), the base is uracil in RNA
- Most RNA is single stranded
List the different types of RNA in the cell.
- sncRNA
- lncRNA
- hnRNA
- mRNA
- rRNA
- tRNA
What is sncRNA?
short non-coding RNA < 200 nucleotides
What is lncRNA?
long noncoding RNA
What is hnRNA?
heterogenous nuclear RNA (term for unprocessed mRNA)
What is mRNA?
messenger RNA - codes for a protein, goes to the ribosome for translation
What is rRNA?
ribosomal RNA - becomes part of the ribosome (large complex that translates proteins)
What is tRNA?
transfer RNA - attaches to an amino acid and carries the amino acid to incorporation into a protein (translation)
Which types of RNA are only found in eukaryotes? [2]
hnRNA (pre-mRNA)
lncRNA
Of total RNA, how much is coding and how much is non-coding?
- Coding RNA = 4% of total
- Non-coding RNA = 96% of total
Which RNA is coding?
- mRNA
Which RNA is non-coding?
- rRNA
- tRNA
- sncRNA
- lncRNA
Is RNA inherited like DNA?
No.
How is RNA synthesized?
- One strand of DNA is the template for RNA synthesis
- The template strand (a.k.a. anti-sense strand), is in the 3’ to 5’ direction
- mRNA transcription proceeds in the 5’ to 3’ direction
- An enzyme called RNA polymerase joins the ribonucleotides together via phosphodiester linkages to make the RNA strand.
- No primer necessary in RNA synthesis
The anti-sense strand in RNA synthesis is in the […] direction.
- 3’ to 5’ direction
- mRNA transcription proceeds in the 5’ to 3’ direction
What defines a gene?
- Only specific regions of the genomic DNA are transcribed
- mRNA, tRNA, and rRNA all need to be transcribed from the genomic DNA
- Sequences that control the initiation of transcription occur before the gene coding sequence
Describe the lay-out of a gene.
- Promoter - sequence of this region is where RNA polymerase binds DNA
- Terminator - sequence where RNA polymerase falls off the DNA
- Initiation (start) and termination (stop) codons for protein translation
DNA synthesis occurs in the 5’ to 3’ direction.
True or False?
True.
DNA synthesis occurs in the 3’ to 5’ direction.
True or False?
False.
DNA synthesis occurs in the 5’ to 3’ direction.
mRNA transcription proceeds in the 5’ to 3’ direction.
True or False?
True.
mRNA transcription proceeds in the 3’ to 5’ direction.
True or False?
False.
mRNA transcription proceeds in the 5’ to 3’ direction.
Describe the prokaryotic gene structure.
- Shine-Dalgarno sequence - ribosome binding site in prokaryotic mRNA
- No separation between DNA and cytoplasm
RNAs are synthesized as precursor RNAs that need to be processed to their final form.
True or False?
True.
Eukaryotic mRNA molecules often require extensive processing and transport, while prokaryotic mRNA molecules do not.
However, the primary transcripts of both rRNAs and tRNAs must undergo a series of processing steps in prokaryotic as well as eukaryotic cells.
Where does the splicing occur?
- Splicing occurs in the nucleus
- Introns are cut out of the pre-mRNA and the exons are rejoined to give the functional mRNA.
- mRNA exits the nucleus and goes into the cytoplasm for translation
- Introns can also be present in rRNA and tRNA genes
Describe modifications to the 5’ end of mRNA in eukaryotes.
- 5’ cap added to the 5’ end of the mRNA (modified guanine nucleotide: 7-methyl guanine)
In prokaryotic cells, the triphosphate is present at the 5’ end of the mRNA but no modified guanine nucleotide.
Why is the cap structure (the modified guanine nucleotide) added to the 5’ end of mRNA in eukaryotes?
- Required to help initiate translation of the mRNA into a protein by the ribosome in the cytosol
Describe modifications to the 3’ end of mRNA in eukaryotes.
- The poly(A) tail is a series of up to 250 adenine nucleotides that is added to the 3ʹ-end of the mRNA – only occurs in eukaryotic cells.
- Long poly(A) tail needed to protect against mRNA degradation
The 5’-cap and the poly(A) tail added to the ends of mRNA in eukaryotes help to [3]:
- Increase the stability of the mRNA - protects mRNA from ribonucleases and degradation
- Transport the mature mRNA out of the nucleus and into the cytoplasm
- Help the ribosome to bind to the mRNA to begin translation.
What is the main purpose of the poly(A) tail?
Long poly(A) tail needed to protect against mRNA degradation
mRNA is cleaved downstream of the polyA cleavage signal (AAUAAA in human cells), then PolyA polymerase
adds the adenine nucleotides
Discuss the total RNA in a cell.
- The thousands of mRNAs in a cell represent only about 3 to 5% of the cellular RNA
- Greater than 90% of the RNA in a metabolically active cell is rRNA which is found in ribosomes
- tRNA represents ~4% of the cellular RNA
- rRNA combines with ribosomal proteins to form ribonucleoprotein complexes that make up the large and small subunit of the ribosome
- During protein synthesis, one large ribosomal subunit and one small ribosomal subunit combine to form a ribosome
- The ribosome translates mRNA into proteins
What are ribosomes?
- Proteins are synthesized by the large RNA–protein complexes called ribosomes
- An E. coli cell contains approximately 20,000 ribosomes in the cytoplasm
- Human cells contains more than a million ribosomes, some free in the cytoplasm and some attached to the outer surface of the rough endoplasmic reticulum
Compare the composition of eukaryotic and bacterial ribosomes.
- Eukaryotic ribosomes are 80S - composed of a 40S small subunit and a 60S large subunit.
- Bacterial ribosomes are 70S, composed of a 50S large subunit and a 30S small subunit.
What is the function of tRNAs and how many types are there?
- Transfer RNAs carry amino acids to the ribosome for incorporation into proteins
Describe the detailed structure of a bacterial ribosome.
Which strand codes for the protein: sense or anti-sense?
- The sense strand codes for the protein
- The anti-sense strand acts as the template for mRNA synthesis
What occurs at the E site of a bacterial ribosome?
tRNA departs to go and pick up another amino acid
What happens at the P site of a bacterial ribosome?
tRNA with amino acid that is being attached to the protein
What happens at the A site of a bacterial ribosome?
- tRNA carrying the next amino acid to be attached to the protein
Describe translation initiation in prokaryotes.
- Translation is initiated by the binding of a small ribosomal subunit to an mRNA by base pairing between a sequence of ~7 nucleotides called the Shine-Dalgarno (SD) sequence
- SD sequence is near the 5’ end of the mRNA and is complementary to the 3’end of the rRNA of the small ribosomal subunit
- The anticodon (UAC) of the initiator tRNA- Methionine base pairs with the start codon (AUG) of the mRNA
- The large subunit then comes in to form the initiation complex
What is the start codon and what does it code for?
AUG - methionine
Describe translation initiation in eukaryotes.
- Translation is initiated by the binding of an initiator Methionine-tRNA and translation initiation factors (not shown here) to a small ribosomal subunit
- Next the 5’capped end of the mRNA, which is combined with specific proteins, associates with the initiator Met-tRNA- small ribosomal complex
- The complex moves along the mRNA until a start codon (AUG) is found
- Once the UAC anticodon sequence of the initiator Met-tRNA base pairs with the AUG sequence of the mRNA, the large ribosomal subunit joins the complex to form the initiation complex
Describe translation elongation in prokaryotes and eukarotes.
- Elongation and termination of translation are similar in prokaryotes and eukaryotes
- Elongation is the formation of peptide bonds between adjacent amino acids
- After the initiation complex is formed, the second set of three nucleotides (triplet or codon) in the mRNA that follows the start codon will be bound by a tRNA carrying the complementary anticodon
- In this example the second codon in the mRNA is “CUG” which is recognized by a tRNA carrying Leucine and the ‘GAC’ anticodon
- A peptidyl bond is formed in the ‘P’ position by a peptidyl transferase (RNA enzyme that is part of the large ribosomal subunit)
- Once the amino acid from a tRNA has been incorporated into the peptide chain, the tRNA exits the ribosome (E site)
Describe translation termination in prokaryotes and eukaryotes.
- The elongation process continues until a UAA, UAG or UGA stop codon (or termination codon) is encountered
- There are no naturally occurring tRNAs with anticodons that are complementary to these codons (this is mostly true…)
- A protein called a termination factor or release factor recognizes the stop codon and binds to the ribosome
- The uncharged tRNA, completed protein, and the mRNA dissociate from the ribosome
- A ribosome releasing factor separates the ribosomal subunits
What is the stop codon?
There are 3: UAA; UAG; UGA
Describe the genetic code.
- The codons are read in the 5’ to 3’ direction in an mRNA.
- Amino acids are designated by the standard three-letter abbreviations.
The genetic code is universal.
True or False?
False.
What is the Shine-Dalgarno sequence?
- The Shine-Dalgarno sequence is a ~7 nucleotide sequence is near the 5’ end of prokaryotic mRNA and is complementary to the 3’ end of the rRNA of the small ribosomal subunit.
- It is located ~8 nucleotides upstream of the AUG start codon and does not get translated into the protein.
In prokaryotes
How does the initiator transfer RNA differ in prokaryotes and eukaryotes?
- In prokaryotes, the initiator Methionine is Formylated at the amino group (fMet)
- In eukaryotes, the initiator Methionine is NOT formylated.
What happens to the DNA products during the second and third cycles of PCR?
- From the first-cycle products shown at the top of the diagram, the next cycle leads to four products, two of which are identical to the first-cycle products, and two of which are made entirely of new DNA.
- During the third cycle, the latter give rise to short products that, in subsequent cycles, accumulate in an exponential fashion.
Typically PCR reactions are 25 to 30 cycles.
There is more than one restriction site for each
enzyme in the pUC8 plasmid.
True or False?
False.
There is only one restriction site for each enzyme in the plasmid
There is only one restriction site for each enzyme in the pUC8 plasmid
True or False?
True.
