2 Nearest Neighbor Analysis (Kornberg 1956): In vitro DNA synthesis•Template DNA•DNA polymerase•dNTPs*dATP*dCTP*dGTP*dTTP•Allow DNA replication to take place•Denature DNA•Treat with micrococcal nuclease-yields mononucleotides with 3’ terminal phosphates
3 ssDNA:ssDNA treated with micrococcal nuclease:ACGT*ACGT*
4 Exercise:List all possible dinucleotides (analysis of nearest 5’ neighbor)Which must be present in equal frequencies if the DNA strands are:Parallel ? Antiparallel?
5 Exercise:List all possible dinucleotides (analysis of nearest 5’ neighbor)ApAApCApGApTGpAGpCGpGGpTCpACpCCpGCpTTpATpCTpGTpTWhich must be present in equal frequencies if the DNA strands are:Parallel ? Antiparallel?ApC = TpG CpA = GpTApG = TpC CpG = GpCApT = TpA CpT = GpAApC = GpT CpA = TpGApG = CpT GpA = TpC
8 DNA Melting*Temperature at which DNA strands are ½ denatured is the melting temperature or Tm*GC content of DNA has a significant effect on Tm
9 Comparison of Typical Prokaryotic and Eukaryotic Cells
10 Chromosome Sequence and Diversity Species# of chromosome(s)Form of chromosome(s)Genome size (Mb)PROKAYOTES:Escherichia coli1Circular4.6Agrobacterium tumifaciens43 Circular Linear5.67EUKARYOTES:Sacharomyces cerevisiae16Linear12.1Arabidopsis thaliana5125Drosophila melanogaster180Homo sapiens232,900
11 Gene density (genes/Mb) Gene Density in Different OrganismsSpeciesGenome SizeApprox. # of genesGene density (genes/Mb)PROKAYOTES:Escherichia coli4.64,400950Agrobacterium tumifaciens5.75,400960EUKARYOTES:Sacharomyces cerevisiae125,800480Arabidopsis thaliana12525,500200Drosophila melanogaster18013,70080Homo sapiens2,90027,0009.3