Biological Sciences 210 - Genetics and Molecular Biology

Fall, 2006

Sequence of the CF Gene

The 4 Nucleotides in DNA

DNA Structure

The DNA Double Helix

E.M. of Prokaryotic Genome

Electron Micrograph of Plasmids

Genes and Intergenic Regions

Chromatin Structure

Chromatin Structure

Chromatin Structure

Human Karyotype

Comparative Genomes

Gene Structure

One Strand of a Gene is the Transcribed (Template) Strand

Figure 3-4, p. 60

Protein Structure

The Genetic Code

Missense Point Mutation

Wild-Type Missense Mutant

• DNA 3'-TACTTTGTCGGTCCA…-5' 3’-TACTTTGTAGGTCCA…-5'

• mRNA 5'-AUGAAACAGCCAGGU…-3' 5’-AUGAAACAUCCAGGU…-3'

• Protein fMet--Lys-Gln--Pro-Gly… fMet--Lys-His--Pro-Gly…

Nonsense Point Mutation

Wild-Type Nonsense Mutant

• DNA 3'-TACTTTGTCGGTCCA…-5' 3'-TACATTGTAGGTCCA…-5'

• mRNA 5'-AUGAAACAGCCAGGU…-3’ 5'-AUGUAACAUCCAGGU…-3'

• Protein fMet--Lys-Gln--Pro-Gly… fMet

Silent Point Mutation

Wild-Type Silent Mutant

• DNA 3'-TACTTTGTCGGTCCA…-5' 3'-TACTTTGTTGGTCCA…-5'

• mRNA 5'-AUGAAACAGCCAGGU…-3’ 5'-AUGAAACAACCAGGU…-3'

• Protein fMet--Lys-Gln--Pro-Gly… fMet--Lys-Gln--Pro-Gly…

Frameshift Mutations

• READING FRAME - Translation of an mRNA must start at the appropriate AUG of the mRNA and work along the mRNA in a 5' ----> 3' direction, reading the mRNA in units of 3. It must read in the exactly correct sequence of 3-base codons.

• Demonstration:

• THE FAT CAT ATE THE BIG RAT

• HEF ATC ATA TET HEB IGR AT

Frameshift Mutation• Frameshift mutations change the reading frame of the messenger RNA.

• For example: Template strand of DNA

• 3'-TACAGAGTGCTGCAT…-5'

• 5'-AUGUCUCACGACGUA…-3' mRNA

• AUG UCU CAC GAC GUA… codons

• fMet -- Ser -- His -- Asp -- Val…

• Through a deletion, the DNA strand loses the fifth base in the sequence above:

• 3'-TACAAGTGCTGCAT…-5'

• 5'-AUGUUCACGACGUA…-3' mRNA

• AUG UUC ACG ACG UA…

• fMet-- Phe -- Thr -- Thr…

Structure of the CFTR Protein

Automated DNA Sequencing Data

GGCTTACGGTTACTCGAATTCTACCGCAAGAATTCTGCTATTT

*Cuts at 1st EcoR I site, but not at 2nd EcoR I site (Partial digest):

GGCTTACGGTTACTCG + AATTCTACCGCAAGAATTCTGCTATTT

GGCTTACGGTTACTCGAATTCTACCGCAAGAATTCTGCTATTT

*Does not Cut at 1st EcoR I site, but cuts at 2nd EcoR I site (Partial digest):

GGCTTACGGTTACTCGAATTCTACCGCAAG + AATTCTGCTATTT

Now, you can align into Contigs!

GGCTTACGGTTACTCGAATTCTACCGCAAG

AATTCTACCGCAAGAATTCTGCTATTT

GGCTTACGGTTACTCGAATTCTACCGCAAGAATTCTGCTATTT

Constructing cDNA