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URL: http://www.addgene.org/33031
Proper Citation: RRID:Addgene_33031
Insert Name: Toca-1
Organism: Homo sapiens
Bacterial Resistance: Ampicillin
Defining Citation: PMID:15260990
Vector Backbone Description: Backbone Size:4350; Vector Backbone:pCS2+MT; Vector Types:Mammalian Expression; Bacterial Resistance:Ampicillin
Comments: Molecular cloning of Toca-1 was difficult, because the cDNA encoding human Toca-1 is lethal to E. coli. The toxicity problem was not resolved by using vectors with low copy numbers or with minimal transcriptional activity, suggesting that the source of toxicity was not the translated protein product but rather the DNA sequence itself. The toxic region of the Toca-1 cDNA was localized to codon 127. The Xenopus tropicalis homolog of Toca-1 is completely nontoxic when cloned using E. coli and is identical in amino acid sequence to human Toca-1 between amino acids 124 and 127, but these amino acids are encoded by different codons. Replacing codons 126 and 127 in the human Toca-1 with the X. Tropicalis sequence (which constitute silent mutations) completely ameliorated the toxicity problem and allowed us to propagate the human Toca-1 clone in E. coli without difficulty.
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Source: Addgene (RRID:SCR_002037)
Description: Non-profit plasmid repository dedicated to helping scientists around the world share high-quality plasmids. Facilitates archiving and distributing DNA-based research reagents and associated data to scientists worldwide. Repository contains over 65,000 plasmids, including special collections on CRISPR, fluorescent proteins, and ready-to-use viral preparations. There is no cost for scientists to deposit plasmids, which saves time and money associated with shipping plasmids themselves. All plasmids are fully sequenced for validation and sequencing data is openly available. We handle the appropriate Material Transfer Agreements (MTA) with institutions, facilitating open exchange and offering intellectual property and liability protection for depositing scientists. Furthermore, we curate free educational resources for the scientific community including a blog, eBooks, video protocols, and detailed molecular biology resources.
URL: http://www.addgene.org
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