2.7: DNA Replication, Transcription and Translation

Describe the meaning of "semi-conservative" in relation to DNA replication.

Understanding: The replication of DNA is semi-conservative and depends on complimentary base pairing.

Describe the meaning of "semi-conservative" in relation to DNA replication.

Understanding: The replication of DNA is semi-conservative and depends on complimentary base pairing.

Semi-conservative means the products of DNA replication each contain one of the original DNA strands and one new strand.

Explain the role of complementary base pairing in DNA replication.​

Understanding: The replication of DNA is semi-conservative and depends on complimentary base pairing.

Complementary base pairing (A-T, C-G) ensures that the DNA sequence remains consistent after DNA replication. This ensures that the genetic code remains intact between generations.

State why DNA strands must be separated prior to replication.

Understanding: Helicase unwinds the double helix and separates the two strands by breaking hydrogen bonds.

The two strands of the parent DNA molecule must separate so that each can serve as a template for the new DNA strands that are being built.

Outline the function of helicase.

Understanding: Helicase unwinds the double helix and separates the two strands by breaking hydrogen bonds.

Helicase is an enzyme that attaches to the DNA and moves along the molecule separating unwinding the helix and separating the two strands by breaking hydrogen bonds.

State the role of the origin of replication in DNA replication.

Understanding: Helicase unwinds the double helix and separates the two strands by breaking hydrogen bonds.

The origin of replication is the sequence of DNA nucleotides at which replication is initiated.

Contrast the number of origins in prokaryotic cells to the number in eukaryotic cells.​

Understanding: Helicase unwinds the double helix and separates the two strands by breaking hydrogen bonds.

Prokaryotic cells have 1 origin of replication (therefor one replication bubble with two replication forks)

Eukaryotic cells have many origins of replication (therefor multiple replication bubbles that eventually fuse)

Describe the movement of DNA polymerase along the DNA template strand.

Understanding: DNA polymerase links nucleotides together to form a new strand, using a pre-existing strand as a template.

DNA polymerase moves along the parent DNA strand from the 3' end to the 5' end of the parent strand. The parent/template is read from 3' to 5'.

DNA polymerase builds a complementary strand of DNA from the parent/template strand. The daughter strand is built from 5' to 3'.

Describe the action of DNA polymerase III in DNA replication.​

Understanding: DNA polymerase links nucleotides together to form a new strand, using a pre-existing strand as a template.

DNA polymerase "reads" a parent/template DNA strand and adds complementary nucleotides to build a new strand of DNA.

DNA polymerase can only add new nucleotides to the 3' end of the growing daughter strand.

Define "transcription."

Understanding: Transcription is the synthesis of mRNA copied from the DNA base sequences by RNA polymerase.

During transcription, a section of the cell's DNA serves as a template for creation of an RNA molecule.

In some cases, the newly created RNA molecule is itself a finished product, and it serves an important function within the cell. In other cases, the RNA molecule carries messages from the DNA to other parts of the cell for processing. Most often, the RNA is used to manufacture proteins (during translation).

Outline the role of RNA polymerase in transcription.

Understanding: Transcription is the synthesis of mRNA copied from the DNA base sequences by RNA polymerase.

RNA polymerase is an enzyme that is responsible for transcribing a DNA sequence into an RNA sequence.

Summarize the steps of transcription.

Understanding: Transcription is the synthesis of mRNA copied from the DNA base sequences by RNA polymerase.

Initiation

RNA polymerase, together with one or more general transcription factors, binds to a sequence of DNA called the "promoter".

RNA polymerase separates the two strands of the DNA helix by breaking the hydrogen bonds between complementary DNA nucleotides.

Elongation

RNA polymerase adds RNA nucleotides (which are complementary to the nucleotides of one DNA strand).

RNA polymerase catalyzes the formation of the RNA sugar-phosphate backbone to form an RNA strand.

Termination

At a DNA sequence called the "terminator", RNA polymerase breaks the hydrogen bonds of the RNA-DNA helix, freeing the newly synthesized RNA strand.

Outline the base pairing in transcription.

Understanding: Transcription is the synthesis of mRNA copied from the DNA base sequences by RNA polymerase.

A complementary strand of RNA may be constructed from a DNA template sequence following the base pairing rules: A=U and G≡C.

Define "sense" and "antisense" in relation to DNA transcription.

Understanding: Transcription is the synthesis of mRNA copied from the DNA base sequences by RNA polymerase.

Two strands of complementary sequence are referred to as sense and antisense. The DNA sense strand has the same sequence as the mRNA being transcribed (substituting U for T) however the DNA sense strand is itself is not transcribed.

The DNA antisense strand, with bases complementary to the DNA sense strand, is used as a template for the RNA during transcription.

Identify the sense and antisense strands of DNA given a diagram of translation.​

Understanding: Transcription is the synthesis of mRNA copied from the DNA base sequences by RNA polymerase.

On a diagram of transcription, the DNA strand that is serving as the template for making RNA is the antisense strand. The strand of DNA that not being transcribed is the sense strand.

Define "translation"

Understanding: Translation is the synthesis of polypeptides on ribosomes.

Translation is the process in which ribosomes in the cytoplasm or ER synthesize a specific amino acid chain, or polypeptide, based on the mRNA sequence (that had been transcribed from DNA).

State the location of translation in a prokaryotic cell.​

Understanding: Translation is the synthesis of polypeptides on ribosomes.

Because there is no nucleus to separate the processes of transcription and translation, when bacterial genes are transcribed, their transcripts can immediately be translated by a ribosome in the cytoplasm.

State the location of translation in a eukaryotic cell.​

Understanding: Translation is the synthesis of polypeptides on ribosomes.

Transcription and translation are spatially and temporally separated in eukaryotic cells; transcription occurs in the nucleus to produce a mRNA which exits the nucleus and is translated in the cytoplasm on a free ribosome or at the endoplasmic reticulum (ER) on a bound ribosome.

Summarize the steps of translation.

Understanding: Translation is the synthesis of polypeptides on ribosomes.

Initiation

The ribosome subunits binds to mRNA at a specific area (called the start codon).

Elongation

The ribosome repeatedly matches tRNA anticodon sequences to the mRNA codon sequence. Each time a new tRNA comes into the ribosome, the amino acid that it was carrying gets added to the elongating polypeptide chain.

Termination

The ribosome hits a stop sequence of mRNA (called the stop codon), then it releases the polypeptide and the mRNA.

Define "genetic code."

Understanding: The amino acid sequence of polypeptides is determined by mRNA according to the genetic code.

The genetic code is the rules used by all cells to translate information encoded within genetic material (DNA) into proteins. The code defines how sequences of three mRNA nucleotides, called codons, specify which amino acid will be added next during protein synthesis.

Outline the role of messenger RNA (mRNA) in translation.

Understanding: The amino acid sequence of polypeptides is determined by mRNA according to the genetic code.

Messenger RNA (mRNA) carries the genetic information transcribed from DNA in the form of a series of three-nucleotide code ("codon"), each of which specifies a particular amino acid.

Outline the role of transfer RNA (tRNA) in translation.

Understanding: The amino acid sequence of polypeptides is determined by mRNA according to the genetic code.

Each type of amino acid has its own type of tRNA, which binds it and carries it to the growing end of a polypeptide chain if the next codon of mRNA calls for it. The correct tRNA with its attached amino acid is selected at each step because each specific tRNA molecule contains a three-base sequence ("anticodon") that can base-pair with its complementary code on the mRNA.

Outline the role of ribosomal RNA (rRNA) in translation.

Understanding: The amino acid sequence of polypeptides is determined by mRNA according to the genetic code.

Ribosomal RNA (rRNA) associates with a set of proteins to form ribosomes. These complex structures, which physically move along an mRNA molecule, catalyze the assembly of amino acids into polypeptide chains.

Define "codon" as related to translation.

Understanding: Codons of three bases on mRNA correspond to one amino acid in a polypeptide.

A codon is a three-base sequence (three nitrogenous bases in a row) on mRNA. It codes for a specific amino acid to be brought to the growing polypeptide during translation.

Define "redundant" as related to the genetic code.

Understanding: Codons of three bases on mRNA correspond to one amino acid in a polypeptide.

The genetic code is redundant, meaning more than one codon may specify a particular amino acid. For example, the codons CCC and CCG both code for the same amino acid, proline.

Explain how using a 4 letters nucleic acid "language" can code for a "language" of 20 amino acid letters in proteins.

Understanding: Codons of three bases on mRNA correspond to one amino acid in a polypeptide.

There are 64 possible permutations, or combinations, of three-letter nucleotide sequences that can be made from the four nucleotides. Although each codon is specific for only one amino acid (or one stop signal), the genetic code is described as redundant, because a single amino acid may be coded for by more than one codon.

Define "anticodon" as related to translation.

Understanding: Translation depends on complementary base-pairing between codons on mRNA and anticodons on tRNA.

An anticodon is a three-base sequence on tRNA that is complementary to the mRNA codon to which it forms hydrogen bonds.

Outline the role of complementary base pairing between mRNA and tRNA in translation.

Understanding: Translation depends on complementary base-pairing between codons on mRNA and anticodons on tRNA.

The incorporation of the correct amino acid into the polypeptide chain depends on a tRNA bringing the correct amino acid to the ribosome.

In order to ensure the correct amino acid, the mRNA codons are recognized by the tRNA anticodon which binds to the appropriate codon by complementary base pairing via hydrogen bonds. Each time a new tRNA comes into the ribosome, the amino acid that it was carrying gets added to the elongating polypeptide chain by the ribosome.

State the purpose of PCR.

Application: Use of Taq DNA polymerase to produce multiple copies of DNA rapidly by the polymerase chain reaction (PCR).

Polymerase chain reaction (PCR) is a technique used to amplify, or make many copies of, a specific target region of DNA in a test tube.

Outline the process of the PCR.

Application: Use of Taq DNA polymerase to produce multiple copies of DNA rapidly by the polymerase chain reaction (PCR).

Denaturation

Heat is used to denature a sequence of DNA

Annealing

The temperature is lowered and a primer sequence is added to the reaction mixture. The primer is a short sequence of nucleotides that binds ("anneals") to the DNA and provides a starting point for DNA synthesis.

Extension

Raise the reaction temperatures again so that Taq polymerase can synthesize new strands of DNA, starting from the primer.

This cycle repeats multiple times to create many copies of the original DNA strand.

Explain the use of Taq DNA polymerase in the PCR.

Application: Use of Taq DNA polymerase to produce multiple copies of DNA rapidly by the polymerase chain reaction (PCR).

Taq DNA polymerase is an enzyme that makes new strands of DNA, using existing strands as templates. It is named after the heat-tolerant bacterium from which it was isolated (Termus aquaticus) which lives in hot springs and hydrothermal vents. Taq polymerase is very heat-stable, meaning it can withstand the high temperatures used in the PCR.

Outline uses of the PCR.

Application: Use of Taq DNA polymerase to produce multiple copies of DNA rapidly by the polymerase chain reaction (PCR).

The goal of PCR is to make enough copies of the target DNA sequence that it can be analyzed or used in some other way. For instance, DNA amplified by PCR may be sent for sequencing, visualized by gel electrophoresis, or cloned into a plasmid for further experiments. PCR is used in many areas of biology, forensics and medicine.

Outline the benefits of using gene transfer technology in the production of pharmaceutical insulin.

Application: Production of human insulin in bacteria as an example of the universality of the genetic code allowing gene transfer between species.

Prior to widespread use of gene transfer technology, insulin for diabetes was isolated from pig pancreases. Some benefits of using gene transfer technology to produce insulin are:

Biotechnologically produced insulin is indistinguishable from human insulin produced in the pancreas and is therefore less likely to cause allergic reactions in diabetics.

Large quantities of insulin can be produced at the same time.

The ethical issues for diabetics who could not use pig's insulin because of religious beliefs or vegetarianism are overcome.

This form of insulin is absorbed more rapidly than animal derived insulin thus showing its effectiveness in a shorter duration.

Define "universal" as related to the genetic code.

Application: Production of human insulin in bacteria as an example of the universality of the genetic code allowing gene transfer between species.

With minor exceptions, all life uses the same genetic code.

Summarize the process for production of human insulin in bacteria.

Application: Production of human insulin in bacteria as an example of the universality of the genetic code allowing gene transfer between species.

Recombinant DNA technology has made it possible to insert a human gene into the DNA of a common bacterium. This "recombinant" bacteria will produce the protein encoded by the human gene because the genetic code is universal.

Scientists build the human insulin gene in the laboratory. Then they remove a plasmid from the bacteria insert the human insulin gene into the plasmid.

The "recombinant" plasmid is returned to the bacteria. The "recombinant" bacteria are put in large fermentation tanks where they repeatedly divides and use the insulin gene to begin producing human insulin.

The insulin is then harvested from the bacteria, purified and used as a medicine for people.

Use a genetic code table to deduce the mRNA codon(s) given the name of an amino acid.

Skill: Use a table of the genetic code to deduce which codons corresponds to which amino acids.

If given the amino acid name and asked to find the mRNA codon(s) by which it is coded:

Find the amino acid name in the genetic code table and the corresponding mRNA codons.

For example: the amino acid PHE is coded for by UUU and UUC.

Use a genetic code table to deduce amino acid given an mRNA codon sequence.

Skill: Use a table of mRNA codons and their corresponding amino acids to deduce the sequence of amino acids coded by a short mRNA strand of known base sequence.

If given the mRNA codon and asked to find the amino acid for which it codes:

Find the mRNA codon in the genetic code table and the corresponding amino acid.

For example: The mRNA codon CCG codes for the amino acid PRO.

Use a genetic code table to deduce amino acid given an tRNA anticodon sequence.

Skill: Use a table of mRNA codons and their corresponding amino acids to deduce the sequence of amino acids coded by a short mRNA strand of known base sequence.

If given the tRNA anticodon, first determine the complementary mRNA codon and then find the mRNA codon in the genetic code table and the corresponding amino acid. Unless otherwise noted, the genetic code table uses mRNA codons!

For example: The tRNA anticodon CCG is the complement to the mRNA codon GGC which codes for the amino acid GLY.

Use a genetic code table to deduce amino acid given an DNA antisense sequence.

Skill: Use a table of mRNA codons and their corresponding amino acids to deduce the sequence of amino acids coded by a short mRNA strand of known base sequence.

The antisense DNA is complementary to the mRNA. So if given the antisense sequence, first determine the complementary mRNA codon and then find the mRNA codon in the genetic code table and the corresponding amino acid. Unless otherwise noted, the genetic code table uses mRNA codons!

For example: The antisense DNA TTT is the complement to the mRNA codon AAA which codes for the amino acid LYS.

Use a genetic code table to deduce amino acid given an DNA sense sequence.

Skill: Use a table of mRNA codons and their corresponding amino acids to deduce the sequence of amino acids coded by a short mRNA strand of known base sequence.

The sense DNA is the same code as the mRNA, substituting T and U. So if given the sense sequence, determine the mRNA codon and then find the mRNA codon in the genetic code table and the corresponding amino acid. Unless otherwise noted, the genetic code table uses mRNA codons!

For example: The if the sense DNA is ATG then the mRNA codon is AUG which codes for the amino acid MET.

Compare dispersive, conservative and semi-conservative replication.

Skill: Analysis of Meselson and Stahl's results to obtain support for the theory of semi-conservative replication of DNA.

Semi-conservative model

Two parental strands separate and each makes a copy of itself. After one round of replication, the two daughter molecules each comprises one old and one new strand.

Conservative model

The parental molecule directs synthesis of an entirely new double-stranded molecule, such that after one round of replication, one molecule is conserved as two old strands.

Dispersive model

Material in the two parental strands is distributed more or less randomly between two daughter molecules.

Outline the method used in the Meselson and Stahl experiment.

Nature of Science: Obtaining of evidence for scientific theories- Meselson and Stahl obtained evidence for the semi-conservative replication of DNA.

1. Grow bacteria in a medium with "heavy" nitrogen (N15) so all the DNA contains heavy nitrogen to start.

2. Transfer some bacteria to "light" nitrogen (N14), bacterial growth continues and with each cell division cycle the light nitrogen incorporates into the DNA.

3. Take samples of bacteria after each round of replication (about 20 minutes).

4. Centrifuge the samples. The DNA with heavy and/or light nitrogen separate into different bands within the tube.

Explain how the Meselson and Stahl experiments suggested DNA replication was semiconservative.

Skill: Analysis of Meselson and Stahl's results to obtain support for the theory of semi-conservative replication of DNA.

After two generations of replication, half of the DNA was of intermediate density and have of the DNA was light only. There was no heavy-only DNA.

This pattern could only have been observed if each DNA molecule contains a template strand from the parental DNA; thus DNA replication is semiconservative.