2.7 DNA Replication, Transcription, and

TranslationIB Biology Year 2

Fall 2015

Thanks to Ms. Stuckey, modified by V. Azuree 2015

DNA replication

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● Post replication, DNA is semi-conservative○ 1 strand old○ 1 strand new

● Original used as a “template” for the 2 new strands○ Complementary base pairing

■ A-T■ C-G

Semi-Conservative Replication2.7 U1 The replication of DNA is semi-conservative and depends on complementary base pairing.

Even if it seems right, we need to back up theories!

1958 - Meselson and Stahl15N DNA vs. 14N DNACaesium chloride and a centrifugeMultiple generations of E. coli in 15N then

transfer to 14NMeasured density after several generations.

HUH?!?

Evidence for Semi-Conservative Replication2.7 NOS1 Obtaining evidence for scientific theories: Meselson and Stahl obtained evidence for the semi-conservative replication of DNA.

Meselson and Stahl

Link

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See also: Bozeman Bio

p.113 -114 in your book(see answer key if we don’t get to it in class!)

Pause to Analyze2.7 S2 Analysis of Meselson and Stahl’s results to obtain support for the theory of semi-conservative replication of DNA.

Step #1Twisty double-helix in your way? Here comes

helicase!Enzyme that uses ATP to unwind DNA by

breaking H-bondsActually a protein complexLooks like a donut (yum)Untwist AND unzip

Helicase (everyone’s fave enzyme)2.7 U2 Helicase unwinds the double helix and separates the two strands by breaking hydrogen bonds.

DNA REPLICATION !

Step #2Moves from 5’ → 3’ - NO EXCEPTIONS

Leading strand vs. lagging strandDNA polymerase adds in complementary

nucleotides by positioning them properly to form H bonds with the template

Covalently bonds phosphate group to sugar.Pretty fool-proof

DNA polymerase2.7 U3 DNA polymerase links nucleotides together to form a new strand, using the pre-existing strand as a template

https://www.youtube.com/watch?v=8kK2zwjRV0M#t=535

Make MANY copies of DNA using change in temp.

High temp (~95°C) breaks H bondsCool down (~54°C) for re-annealing of primers

Extra primers prevent parents from coming back together

Temp to 72°C to allow Taq DNA polymerase to replicate DNA

Taq DNA polymerasewithstand 95°C, still works at 54°C,

optimum 72°CRepeat over and over

30 cycles - DNA amplified by 1 billion!!

PCR - Polymerase Chain Reaction HUMAN MADE2.7 A1 Use of Taq DNA polymerase to produce multiple copies of DNA rapidly by the polymerase chain reaction (PCR).

PCR - Polymerase Chain Reaction2.7 A1 Use of Taq DNA polymerase to produce multiple copies of DNA rapidly by the polymerase chain reaction (PCR).

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Transcription

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DNA is pretty much useless without being “converted” to readable polypeptides.First step between DNA → Protein = Transcription

Transcription2.7 U4 Transcription is the synthesis of mRNA copied from the DNA base sequences by RNA polymerase.

TranscriptionDNA (template) → mRNA (end result)occurs in the NUCLEUS

1. RNA polymerase binds at the start of a gene2. Similar to DNA replication - RNA polymerase

forms H-bonds between complementary base pairs and covalent bonds between nucleotidesa. Still moves 5’ → 3’

3. RNA separates and double helix reforms4. Transcription stops at the end of the gene.

Transcription2.7 U4 Transcription is the synthesis of mRNA copied from the DNA base sequences by RNA polymerase.

Sense vs. Antisense strandsDoes it matter which strand DNA

copies?YES!!!

Template strand = ANTISENSE strand

is transcribed from 5’ → 3’has the complementary DNA

sequenceSense strand - not transcribed

Sequence matches mRNA (but T for U!)

Transcription2.7 U4 Transcription is the synthesis of mRNA copied from the DNA base sequences by RNA polymerase.

Translation

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Step #2mRNA → polypeptide chainRibosome “reads” and translates mRNA codons into amino acid chains

2 subunits - large and smallcomposed of rRNA, proteins

Translation2.7 U5 Translation is the synthesis of polypeptides on ribosomes.

mRNA - all different lengths (av. 2,000 nucleotides)depends on the gene

How much mRNA and which mRNA is produced?depends on the polypeptides needed by the cell

Pancreaslots of mRNA that carries code for insulin

Don’t forget about tRNA and rRNA, too!

mRNA and the Genetic Code2.7 U6 The amino acid sequence of polypeptides is determined by mRNA according to the genetic code.

How does the tRNA know which amino acids to bring?

Codons!mRNA read in sets of 3

nucleotides that code for a specific amino acid.

64 possibilities (4x4x4)20 amino acidsMemorize them all?

Heck no! CODON CHART!

Codons (or, Yes, Jessica, you have to use the stupid chart thingie)2.7 U7 Codons of three bases on mRNA correspond to one amino acid in a polypeptide.

1. Deduce the codons for:a. Trpb. Tyrc. Arg

2. Deduce amino acid sequence that corresponds to these mRNA sequences:a. ACG b. CACGGGc. CGCGCGAGG

3. If the mRNA contains the base sequence CUC AUC GAA UAA CCCa. deduce the AA sequence of the polypeptide translated from

the mRNA

b. deduce the base sequence of the ANTISENSE strand of DNA

Decoding Base Sequences2.7 S1, S3, S4 Use a table of the genetic code to deduce which codon(s) correspond to which amino acids; 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; Deducing the DNA base sequence for the mRNA strand.

UGGUAU, UACCGU, CGC, CGA, AGA, AGG

ThrHis Gly

Arg Ala Arg

Leu Ile Glu STOP Pro

GAG TAG CTT ATT GGG

1. mRNA has a 3 nucleotide codon

2. tRNA has a 3 nucleotide complementary anticodon

3. Ribosomes are binding sites for both and catalyse polypeptide assembly

Codons and Anticodons2.7 U8 Translation depends on complementary base pairing between codons on mRNA and anticodons on tRNA.

Main events:1. mRNA binds to small subunit of ribosome2. tRNA brings the first AA3. 2nd tRNA brings the next AA. Only 2 tRNAs at a

time!4. Ribosome catalyzes the addition of the 2 AAs5. Ribosome slides down, releasing tRNA 1 and shifting

tRNA 26. Next tRNA comes in7. Ribsome makes next peptide bond

Repeat over and over!Mistakes are rare!

Codons and Anticodons2.7 U8 Translation depends on complementary base pairing between codons on mRNA and anticodons on tRNA.

Diabetes patients need insulinbovine, porcine, shark used in pasthas some differences!can cause allergies

Genetically transfer the human insulin gene into bacteria

Bacteria then mass produce the human version of insulin which we can harvest!

Production of Human Insulin in Bacteria2.7 A2 Production of human insulin in bacteria as an example of the universality of the genetic code allowing gene transfer between species.

More review:Learn Genetics Transcribe and Translate a GeneDNA, Hot Pockets, & The Longest Word Ever: Crash Course Biology #11

Bozeman Science DNA Replication

Bozeman Science Transcription and Translation