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C. elegans Full ORF Entry Clones (DFCI - Vidal)

C. elegans Full ORF Entry Clones 1.1 (DFCI - Vidal)

C. elegans has proved invaluable as a worldwide model system for studies in developmental genetics and neurobiology. In a collaborative effort based at the Dana-Farber Cancer Institute and Harvard Medical School, Dr Marc Vidal and colleagues (http://vidal.dfci.harvard.edu/) have cloned a first version (version 1.1) of all predicted protein-coding open reading frames (ORFs), or the "ORFeome" of Caenorhabditis elegans. Their effort has resulted in the generation of over 12,000 clones (ORFeome 1.1), of which ~4,000 correspond to genes that have remained untouched by any cDNA or expressed sequence tag (EST).

These clones have been produced using InVitrogen Gateway™ recombinational vector system pDONR201. As a starting point, the researchers designed 19,000 primer pairs by computational methods, based on gene predictions from one of first drafts of the worm genome annotation made public in 1999. Gene-specific primer pairs would amplify all 'splice variants' between the two primers designed for any gene. Pools of ~50-1,000 transformants for each set of amplification products were partially sequenced to produce OSTs (ORF sequence tags). This confirmed gene identity and permitted identification of pools where at least one processed mRNA was represented. All the data from this project has been made available in WorfDB, which integrates with other mapping data ( http://worfdb.dfci.harvard.edu/ ). 

 

We offer, for research purposes only, over 10,500 ORF pools from this collection which can be obtained as individual pooled clones from a single well, a subset of pools or the complete set of 114 plates (96-well format).

We can also make subsets of your chosen clones from the collection. Please contact us to discuss your requirements.

Protocols / Clone Handling

Please note that the clone(s) are tested as negative in our phage contamination assay prior to shipping. However, these clones should still be handled with care as no phage assay can be guaranteed to be 100% accurate.

Plates: contain LB broth with 8% glycerol and kanamycin, are sent on Dry Ice. These should be stored at -70°C.

Individual clone pools: have been streaked onto LB agar containing kanamycin (100µg/ml). Please store them at 4°C (not in a freezer). As soon as possible, transfer the culture to LB broth containing kanamycin + 8% glycerol and incubate overnight at 37°C (do not try to transfer single colonies, instead scoop out the whole bacterial culture using a disposable inoculation loop). Incubate overnight at 37°C for subsequent freezing at -70°C. These frozen stocks can then be used as templates for PCR amplification of ORFs, either directly in colony PCRs or as plasmid DNA.

These clones are for research purposes only.

References

We are grateful to Dr. Marc Vidal (Dana-Farber Cancer Institute, Harvard Medical School, Boston) for the gift of this library and associated database and Dr Troy Moore (Open Biosystems) for preparing a copy of the library.

Please acknowledge the Dana-Farber Cancer Institute and Source BioScience LifeSciences in any resulting publications, clones should be described in the paper in their ID form eg T22D1.3

Please also cite the following paper:

C. elegans ORFeome version 1.1: experimental verification of the genome annotation and resource for proteome-scale protein expression: Reboul J, Vaglio P, Rual JF, Lamesch P, Martinez M, Armstrong CM, Li S, Jacotot L, Bertin N, Janky R, Moore T, Hudson JR Jr, Hartley JL, Brasch MA, Vandenhaute J, Boulton S, Endress GA, Jenna S, Chevet E, Papasotiropoulos V, Tolias PP, Ptacek J, Snyder M, Huang R, Chance MR, Lee H, Doucette-Stamm L, Hill DE, Vidal M.Nat Genet. 2003 May;34(1):35-41.


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