Dear all,

I have been trying to use a pgr106 vector for cloning work. However I am facing many issues when I am trying to digest it in the MCS region. The sites I have selected for cloning is cla1 and sal1, however when I am trying to digest the sites, I am getting multiple fragments on gel visualisation. I have tried single digestion and double digestion in all possible combinations, using ethanol precipitation and changing the buffer and enzyme vials, streaking and re-streaking the vector from glycerol and DMSO stocks and also using various vector isolation protocols. I am attaching a representative image where I have digested the vector using not1 and sal1 run along with a 1kb ladder (Thermo fisher scientific). Suggestions please!

Thanks in advanced.

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