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Dana-Farber Cancer Institute, Oxford University


Dana-Farber Team Said ORFs Can Help Sharpen Focus of Gene Maps

Researchers at Dana-Farber Cancer Institute have used a new gene-mapping technique to produce what may be “the clearest picture yet” of all the genes of an organism.

They said that the work on the model organism Caenorhabditis elegans may eventually help improve the way scientists see human and other genomes.

“The fact is, our current picture of the ‘parts list’ of the human genome is rather fuzzy,” said the study’s senior author, Marc Vidal, of Dana-Farber. “Computer programs have been used to predict the position and structure of genes. However, we don’t know exactly where most genes begin and end, and there are literally thousands of gaps in our picture of how the building blocks of genes are arranged.”

In their research, published in the April 7 Nature Genetics, Vidal and colleagues relied on open reading frames to check the accuracy of the existing map of the C. elegans genome.

When a cell creates a protein, the ORF for it is transferred from a gene to messenger RNA, which then carries it to the ribosome. By capturing the RNA within C. elegans cells and converting it into cDNA, the investigators were able to gather the worm’s full set of ORF instructions. The team then compared the segments of cDNAs with the stretches of sequence thought to contain those genes.

Using this technique to examine all 19,000 predicted genes in C. elegans. The researchers found that in more than half the cases the predicted genes did not completely match the actual genes isolated in their study.

“This demonstrates that even in C. elegans … the genome map needs a great deal of correcting and refining,” Vidal said in a statement last week. “We’re still a long way from having a perfect picture of the parts list encoded by the worm genome, let alone the human one.

“The success of this technique with C. elegans suggests that it can be equally successful with the genomes of other creatures, including humans,” said Vidal.

Labs that contributed to the Dana-Farner research included the Unite de Recherche en Biologie Moleculaire, in Belgium; Research Genetics, in Huntsville, Ala.; Life Technologies, of Rockville, Md.; Protedyne, of Windsor, Conn.; McGill University, in Montreal; the Public Health Research Institute, in Newark, NJ; Yale University, in New Haven, Conn.; Albert Einstein College of Medicine, in New York; and Genome Therapeutics, of Waltham, Mass.

Helping to fund the research were the National Cancer Institute, the National Human Genome Research Institute, the National Institute of General Medical Sciences, and the Merck Genome Research Institute.

Oxford Said to Develop Method For Evaluating Non-Coding Genes

Researchers at Oxford University’s Wellcome Trust Center for Human Genetics last week said they have developed a way to evaluate the functional significance of non-coding polymorphisms in the human genome.

Oxford said the discovery may eventually improve the way in which scientists are able to determine genetic susceptibility to disease.

Most SNPs in non-coding regions “may be of little functional significance, but some have important effects” on gene regulation, according to Isis Innovation, an Oxford University spin-off. Today, researchers can screen for SNPs that affect transcriptional regulation by examining DNA in an in vitro setting like reporter genes or electromobility-shift assays. But these are known to produce misleading results.

In vivo evaluation of the effects of SNPs on transcriptional regulation is preferable, though allele-specific real-time PCR of a single heterozygous individual requires the SNP to be linked to a genetic marker that appears on the RNA transcript. “This approach has limited applicability since many non-coding region SNPs may not produce a transcribed marker SNP to discriminate between the alleles,” Oxford said.

In response to these difficulties, Oxford researchers have developed a sensitive method for detecting SNPs that affect gene expression in heterozygous cells, even if there are no suitable genetic markers on the RNA transcript. Indeed, the new approach allows scientists to analyze SNPs in vivo within their natural chromosomal environment, and is subject to all the natural regulatory processes that operate in the specific cell type in which it is investigated.

“If you have a gene that is heterozygous and there’s also a SNP in the promoter sequence that could be affecting whether one gets transcribed or … the heterozygous [gene] gets promoted,” said a spokesman for Isis Innovation, Oxford University’s technology transfer company. “The idea is that if the SNP in the promoter region is having an effect on the binding of the transcription unit, they’ll be able to detect this difference in the binding by using this antibody-detecting technique.

Isis has filed for a patent on this discovery and welcomes contact from potential commercial partners.

— KL

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