Davidson:pSB1A7 Cloning Solution

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Our new pSB1A7 cloning vector blocks read-through transcription

To prevent uncontrolled expression of parts within our device, we built insulated cloning vector pSB1A7 (<partinfo>BBa_J31009</partinfo>). pSB1A7 was built using the pre-existing <partinfo>pSB1A3</partinfo> vector. The terminators in pSB1A3 are designed to prevent read-through from inside the device out into the vector (purple arrows) but the vector is not designed to stop read-through into the device (red arrows). We added double terminators (<partinfo>BBa_B0015</partinfo> shown as solid red and hatched hexagons) in opposite orientations at each end of the BioBrick cloning site region to shield the insert from transcription read-through. When the pancakes are cloned into pSB1A7, tetracycline resistance is dependent on the proper orientation of the two DNA pancakes.

See the part design page for details on how pSB1A7 was built.

pSB1A7 is not compatible with parts carrying B0015

Our new cloning vector pSB1A7 insulates our parts from read-through transcription, but this vector does not accept parts that carry the B0015 double terminator. Our flipping device was originally designed with a B0015 near the end. Several attempts to clone parts carrying B0015 into pSB1A7 failed. This problem prompted a redesign of our system.

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