The human MitoChip: A high-throughput sequencing microarray for mitochondrial mutation detection

Anirban Maitra, Yoram Cohen, Susannah E.D. Gillespie, Elizabeth Mambo, Noriyoshi Fukushima, Mohammad O. Hoque, Nila Shah, Michael Goggins, Joseph Califano, David Sidransky, Aravinda Chakravarti

Research output: Contribution to journalArticlepeer-review

Abstract

Somatic mitochondrial mutations are common in human cancers, and can be used as a tool for early detection of cancer. We have developed a mitochondrial Custom Reseq™ microarray as an array-based sequencing platform for rapid and high-throughput analysis of mitochondrial DNA. The MitoChip contains oligonucleotide probes synthesized using standard photolithography and solid-phase synthesis, and is able to sequence > 29 kb of double-stranded DNA in a single assay. Both strands of the entire human mitochondrial coding sequence (15,451 bp) are arrayed on the MitoChip; both strands of an additional 12,935 bp (84% of coding DNA) are arrayed in duplicate. We used 300 ng of genomic DNA to amplify the mitochondrial coding sequence in three overlapping long PCR fragments. We then sequenced >2 million base pairs of mitochondrial DNA, and successfully assigned base calls at 96.0% of nucleotide positions. Replicate experiments demonstrated >99.99% reproducibility. In matched fluid samples (Urine and pancreatic juice, respectively) obtained from five patients with bladder cancer and four with pancreatic cancer, the MitoChip detected at least one cancer-associated mitochondrial mutation in six (66%) of nine samples. The MitoChip is a high-throughput sequencing tool for the reliable identification of mitochondrial DNA Mutations from primary tumors in clinical samples.

Original languageEnglish (US)
Pages (from-to)812-819
Number of pages8
JournalGenome research
Volume14
Issue number5
DOIs
StatePublished - May 2004

ASJC Scopus subject areas

  • Genetics
  • Genetics(clinical)

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