I am using the Gibson HiFi Assembly Kit to assemble a DNA fragment. Both the DNA fragment and the vector are approximately 1000 bp in length. Note- I am not performing a PCR step. After completing the Gibson assembly reaction, I can observe the desired band corresponding to the insert. However, after performing E. coli transformation, selecting specific colonies, extracting plasmids from them, and running them on a gel, I observe bands that are not of the desired length.
What steps should I take to ensure I obtain the correct bands or vector size of the desired length?
Hello
The vector DNA fragment looks smaller than typical. 1kb looks barely enough to have e.g. AmpR coding sequence (but I do not know details of your design, of course). I suppose that your construction might be non-viable in your host organism and you select any other non-specific plasmid (lack information on any of controls).
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