Molecular Biology LU3L2

• Human Genome Project (HGP)


  The international, collaborative research program whose goal was the complete mapping and understanding of all the genes of humans
• Human Genome Project


  • Coordinated by the National Institutes of Health and the U.S. Department of Energy
  • Additional contributors included universities across the United States and international partners in the United Kingdom, France, Germany, Japan, and China
  • Formally began in 1990 and was completed in 2003, 2 years ahead of its original deadline
• The human genome is made up of 3 billion (3 000 000 000) bases of DNA, split into 23 chromosome pairs
• 1 single gram of DNA is capable of holding an amazing 700 terabytes of data
• It will take 50 years to type the entire human genome if someone types at a speed of 60 wpm (words per minute) and works 8 hours a day

  
• All the DNA in your cells could stretch from the earth to the sun 600 times
• Primary structure of nucleic acids


  • Repeating polymers formed by nucleotides
  • Nucleotides consist of nucleosides + phosphate group
  • Nucleosides consist of: pentose carbon sugar and nitrogen base

  
• Carbon pentose sugars


  • Ribose
  • Deoxyribose
• Nucleoside


  Pentose 5 carbon sugar + nitrogen base
• Nucleotide


  Pentose 5 carbon sugar + nitrogen base + phosphate
• Nucleotides


  • Held together by 5' to 3' phosphodiester bonds

  
• DNA


  Two polynucleotide strands

  
• Nucleic acid sequence


  • Written from 5' end to 3' end
  • Use capital letters
• Secondary structure of nucleic acids

  • DNA is in the shape of a right-handed helix
  • Sequence of the bases are complementary

  
• DNA strands


  • Sense/Plus/Coding strand: 5'->3' DNA strand
  • Antisense/Minus/Not coding strand: Complementary 3'->5' strand
• Practice Exercise 1


  1. Write down the DNA sequence (left) of the above macro-molecule from the 5' prime to the 3' prime end

  2. Write down the RNA sequence which will be complementary to the DNA in question 1
• Practice Exercise 3


  Write down the sequence of nucleotides as presented in the diagram

  
• Different forms of DNA

  • B-form - most common
  • A and C form - laboratory conditions
  • Z form - alternating purine and pyrimidine nucleotides

  
• B form DNA


  Most common form
• Single strand DNA


  • Self complementary which leads to folding of molecule
  • tRNA is clover shaped

  
• Separation of double stranded DNA

  • Anti-parallel strands are held together by hydrogen bonds and base stacking (hydrophobic interactions)
  • DNA denature at 90 °C and above
  • Melting temperature (Tm): 50 % of DNA is denatured

  
• Factors influencing DNA denaturation

  • Nature of DNA
  • Percentage of G+C in DNA sample
  • Environmental conditions
• Renaturation


  Separated strands re-associate
• Gene


  Region of DNA codes for a single RNA or protein
• Genome


  Entire set of genes in a cell, organelle or virus
• Chromosomal DNA


  Genomic DNA complexed with protein (Proteins which DNA is wrapped around)

  
• Locus


  Particular position along the chromosome
• Allele


  • Particular form of gene
  • Homozygous: identical allelic forms
  • Heterozygous: different allelic forms
• Polymorphism


  Different alleles at the same site in the genome
• Single nucleotide polymorphisms (SNPs)


  • Single nucleotide substitution at a precise location
  • Occur normally throughout a person's DNA
  • SNPs within a regulatory gene they may affect the gene's function

  
• Chromosomes


  Consist of centromere, chromatids and telomeres
• Karyotype


  Organised profile of an organism's chromosomes
• Renaturation kinetics and genome complexity

  • Applications of denaturation and renaturation kinetics
  • Understanding genome size and complexity
  • Understanding genetic relatedness
  • Understanding relative proportions of single copy and repetitive sequences
• Nature of Genetic code

  • Codon: sets of three nucleotides to encode a particular amino acid
  • Degeneracy of genetic code: more than one codon may encode a single amino acid
  • Specific codons indicate start and stop of a gene
  • Open reading frame: sequence with number of codons encoding for a continuous protein read in frame flanked by start and stop codons
• Location and packaging of nucleic acids

  • Eukaryotic cells: DNA in nucleus, and organelles like mitochondria
  • RNA normally in cytoplasm
  • DNA direct synthesis of RNA molecules: mRNA, rRNA, tRNA

  
• Packaging of DNA


  • Prokaryotes: tightly coiled in the cytoplasm
  • Eukaryotes: first order packaging = DNA around histones = nucleosome is DNA wound around structure twice
  • Second order packaging = nucleosome form chromatin fibre = chromatin fibre fold and loop to form chromosome structures