Subcloning PCR Products the TOPO Way

"Aarrrggghhhh!" My cry of frustration echoed down the hall. My latest attempt to subclone five important PCR products had failed yet again. After two months of fruitless effort, I was at my wit's end until a colleague suggested I try the TOPO TA Cloning® Kit from Invitrogen. "Look," he said, "all you need to do is to combine 1 microliter of your PCR reaction and 1 microliter of the pCR2.1-TOPO® vector, add 3 microliters of water, and wait 5 minutes. Then transform your competent cells

Written byMichael Brush

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My cry of frustration echoed down the hall. My latest attempt to subclone five important PCR products had failed yet again. After two months of fruitless effort, I was at my wit's end until a colleague suggested I try the TOPO TA Cloning^® Kit from Invitrogen.

"Look," he said, "all you need to do is to combine 1 microliter of your PCR reaction and 1 microliter of the pCR2.1-TOPO^® vector, add 3 microliters of water, and wait 5 minutes. Then transform your competent cells. In the morning, you will have more white colonies than you will know what to do with." As proof, he showed me a petri dish with six blue colonies surrounded by hundreds of white ones.

I was sold on the spot. I promptly borrowed his TOPO TA Cloning kit and set about subcloning those pesky PCR products.

pCR2.1 Topo Map A glorious sight awaited me the ...

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