Genomics

Complete Genomes in The Public DataBases >100 Non-Eukaryotes Eukaryotes: Leishmania 257 Kb 79 orfs Plasmodium falciparum I 947 Kb 205 orfs Plasmodium falciparum I 1,060 Kb 220 orfs Guillardia theta 551 Kb 464 orfs Arabidopsis thaliana 115,428 Kb25,498 orfs Oryza (sativa L. ssp. Indica) 420,000 Kb 50,000 orfs Oryza (sativa ssp. Japonica) 420,000 Kb50,000 orfs Saccharomyces cerevisiae 12,069 Kb 6,294 orfs Schizosaccharomyces pombe 14,000 Kb 6,824 orfs Caenorhabditis elegans 97,000 Kb19,099 orfs Drosophila melanogaster137,000 Kb14,100 orfs Mouse Musculus ~ 3,000,000Kb ~50,000 orfs Homo sapiens ~ 3,000,000Kb ~50,000 orfs

SEQUENCING (according to Sanger et al.) A.Annealing and elongation B.Termination

Manual Sequencing 4 lanes / sample 10 samples / gel bases long a g c t a g c t a g c t 1 2 3

Dye Terminator Sequencing

Sequencing DNA templateprimers dNTPs A C T G A C A G T T G T ddNTPs C G T A G G C GC T GCT A CGATCCGTAAA CGATCCGTAAA CGATCCGTAAA CGATCCGTAAA

Dye Terminator Cycle Sequencing

Primer Walking

Shotgun Sequencing

Single Nucleotide Polymorphism (SNP) Single nucleotide polymorphisms (SNPs), i.e., single nucleotide base substitutions in gene sequences, account for 85% of the genetic variability in humans. This variability quite often accounts for differences in individuals' susceptibility to diseases and varying response to therapeutics. SNPs also have high utility in discovery of novel candidate genes through high resolution mapping

2. Purify PCR products/ Eliminate primers and dNTPs SNaPshot TM Multiplex Kit Work Flow 1. Target Amplification C G 3. SNaPshot™ Reaction Kit

SNaPshot TM Multiplex Kit: Single Base Extension ddGTP ddCTP ddUTP ddATP CCATGACTGATTCC NNNNNNAGCCTGGTACTGACTAAGG C NNNNNNN Interrogation target AmpliTaq® FS G

SNaPshot TM Multiplex Kit: Single Base Extension ddCTP ddUTP ddATP CCATGACTGATTCC NNNNNNAGCCTGGTACTGACTAAGG C NNNNNNN Interrogation target G AmpliTaq® FS Extends and Terminates Primer

CGATCCGAAAA PCR product ACGCGGT CGATCCGAAAA CGATCCGTAAA primer C G T A A C T G ddNTPs CGATCCGTAAA ACGCGGT ACGCGGT ACGCGGT A A T T SNaPshot reaction heterozygote

Sub

DNA micro-arrays

DNA Microarray (Filter) Oncogenes/Cell cycleSignal Transduction Apoptosis/DNA Repair House Keeping Transcrption FactorsReceptors Growth Factors

Glass DNA Chip

Photolithography