Expression profiling using microarrays

1
Expression profiling using micro-arrays

• Using chip technology, make micro-arrays of gene
  specific oligonucleotides, printed onto glass
  slides
• Use to compare gene expression profiles of the
  whole genome between different samples.
• Use two different conditions, eg non-growing and
  growing cells, or normal and cancer cells
• Extract RNA, reverse-transcribe into cDNA, using
  two different fluorescent nucleotides, eg dUTP
  labelled with Cy3 (green) for control, Cy5 (red)
  for test sample.
• Mix samples and hybridise to slides. Gives red,
  green or yellow hybridisation patterns. Analyse
  by fluorescent microscopy.
• Induction gives red dots, repression gives green
  dots, no change gives yellow dots.

2
Expression profiling of the yeast genome
From DeRisi et al., Science, 278, 680-686, 1997
4.1
3
Expression profiling of the yeast genome
time-course
4.2
From DeRisi et al., Science, 278, 680-686, 1997
4
Expression profiling in human cells effect of
serum
From Iyer et al., Science, 283, 83-87, 1999
4.3
5
Expression profiling in human cells cluster
analysis
From Iyer et al., Science, 283, 83-87, 1999
4.4
6
Expression profiling in human cells cluster
analysis
4.5
From Iyer et al., Science, 283, 83-87, 1999
7
Expression profiling and breast cancer
70 Genes
Black - No Metastasis
White - Metastasis
4.6
8
SNPs
Paternal allele CCCGCCTTCTTGGCTTTACA Maternal
allele CCCGCCTTCTCGGCTTTACA
Paternal allele CCCGCCTTCTTGGCTTTACA Maternal
allele CCCGCCTTCTTGGCTTTACA
4.7
9
From SNPs to disease?
4.8
10
Human-mouse chromosome conservation
Mouse chromosomes
2.13
11
Targeted recombination
Targeting vector
neo
neo
neo
Cells die in selection for neo
neo
neo
neo
From Recombinant DNA
4.9
12
Generation of mutant mice
From Human Molecular Genetics 2 (Strachan and
Read)
4.10