2.7 DNA replication, transcription and translation

Question 1

Analysis of Meselson and Stahl’s results

Answer 1

• Grew E. coli bacteria in medium containing N15, a heavy isotope of N14
• After 15 generations, they transferred the bacteria to a medium containing only N14
• The bacteria replicated every 20 min, then extracted
• spun it in a centrifuge, DNA created a layer based on its density

Question 2

results of Meselson and Stahl’s experiment

Answer 2

Their results showed that N14 was incorporated into the new DNA molecules with each new replication.

Question 3

semi-conservative replication

Answer 3

• Two strands of the double helix separate
• Each original strand acts as a guide or template for the new strand
• New strands are created by adding nucleotides following complementary base-pairing
• The two new strands formed will be identical to the original strand.

Question 4

helicase

Answer 4

• Unwinds and unzips the DNA Helix
• Separates the two polynucleotide strands by breaking the hydrogen bonds between complementary base pairs
• ATP is needed by helicase to both move along the DNA molecule and to break the hydrogen bonds
• The two separated strands become parent/template strands for the replication process

Question 5

DNA polymerase

Answer 5

• creates complementary strands from template strands
• DNA polymerase brings nucleotides into position where they can form H-bonds with the template strand
• DNA polymerase always builds in a 5’ to 3’ direction
• DNA polymerase bonds the 5’ (phosphate) end of the free nucleotide to the 3’ (hydroxyl) end of the new strand in a condensation reaction
• DNA polymerase catalyses the covalent phosphodiester bonds between sugars and phosphate groups
• DNA Polymerase proof-reads the complementary base pairing.

Question 6

Energy is needed to…

Answer 6

link new nucleotides to the growing DNA strand

Question 7

deoxynucleoside triphosphates…

Answer 7

• align opposite their matching base partner
• DNA plolymerase cleaves the 2 excess phosphates, energy released is used to link phosphate to 3’ end of sugar covalently

Question 8

Polymerase Chain Reaction (PCR)

Answer 8

1. Denaturation: DNA sample is heated to separate it into two strands
2. Annealing: DNA primers attach to opposite ends of the target sequence
3. Elongation: A heat-tolerant DNA polymerase (Taq) copies the strands

Question 9

Transcription

Answer 9

the synthesis of RNA, using DNA as a template
-happens in the nucleus of eukaryotic cells
- happens in the cytoplasm of prokaryotic cells

Question 10

Translation

Answer 10

when a ribosome reads the mRNA and builds the corresponding polypeptide chain
- happens in the cytoplasm for both eukaryotic and prokaryotic cells

Question 11

Three main types of RNA

Answer 11

Messenger RNA (mRNA): A transcript copy of a gene used to encode a polypeptide
Transfer RNA (tRNA): A clover leaf shaped sequence that carries an amino acid
Ribosomal RNA (rRNA): A primary component of ribosomes

Question 12

transcription process

Answer 12

• enzyme RNA polymerase binds to site on DNA at start of a gene
• RNA polymerase separates the DNA strands and synthesises a complementary RNA copy from the antisense DNA strand
• RNA polymerase forms cvalent bonds between RNA nucleotides
• Eventually, RNA polymerase comes across a termination sequence in the DNA, RNA polymerase will detach, double helix reforms

Question 13

Introns

Answer 13

do not code for proteins and must be removed before the mRNA is considered mature

Question 14

exons

Answer 14

(the coding part) are spliced back together, the completed mRNA strand leaves the nucleus.

Question 15

A ribosome is composed of…

Answer 15

two halves, a large and a small subunit. During translation, ribosomal subunits assemble together like a sandwich on the strand of mRNA:

Question 16

Only certain genes in a genome need to be expressed depending on:

Answer 16

Cell specialism
Environment

Question 17

The genetic code

Answer 17

the set of rules by which information encoded in mRNA sequences is converted into proteins (amino acid (AA) sequences) by living cells

Question 18

Codons are a triplet of bases which…

Answer 18

encodes a particular AA
- The codons can translate for 20 amino acids

Question 19

The coding region always starts with a START codon

Answer 19

(AUG)
Methionine

Question 20

The coding region of mRNA terminates with a STOP codon

Answer 20

the STOP codon does not add an amino acid – instead it causes the release of the polypeptide

Question 21

Amino acids are carried by

Answer 21

transfer RNA (tRNA)
The anti-codons on tRNA are complementary to the codons on mRNA

Question 22

tRNA molecules

Answer 22

• have an anticodon of three bases that binds to a complementary codon on mRNA
• carry the amino acid corresponding to their codon

Question 23

mRNA has a sequence of…

Answer 23

codons that specifies the amino acid sequence of the polypeptide

Question 24

Ribosomes:

Answer 24

act as the binding site for mRNA and tRNA
catalyse the peptide bonds of the polypeptide

Question 25

An outline of translation and polypeptide synthesis:

Answer 25

1. An mRNA binds to the small subunit of the ribosome
2. A molecule of tRNA binds to the ribosome. This tRNA has an anti-codon that is complementary to the first codon on mRNA → That codon is always AUG, which codes for the amino acid “met” (methionine)
3. A second tRNA with an anticodon complementary to the second codon binds. Only 2 tRNA molecules can bind to the mRNA at the same time.
4. A peptide bond is created between the two amino acids – a dipeptide forms
5. The ribosome moves along the mRNA like a roller coaster on a track… it shifts over one codon, and the next tRNA carrying the appropriate amino acid comes in and binds to that codon with its anti-codon.
6. This process repeats over and over again until a STOP codon is reached. This doesn’t code for an amino acid, but rather, initiates the detachment of the polypeptide.
7. This polypeptide (amino acid chain) can now go on to fold and produce a functional protein

Question 26

produce human insulin in bacteria

Answer 26

• The gene responsible for insulin production is extracted from a human cell
• It is spliced into a plasmid vector (for autonomous replication and expression) before being inserted into a bacterial cell
• The transgenic bacteria (typically E. coli) are then selected and cultured in a fermentation tank (to increase bacterial numbers)
• The bacteria now produce human insulin, which is harvested, purified and packaged for human use (i.e. by diabetics)