I have made a construct with a target gene and the sequence result was good. is there any chance of mutation in the vector during further plating or subculture what are the precautions should i take to avoid mutation?
1. For the vector, they can be stored frozen and stay for a long time. Even large BAC vectors under appropriate conditions (TE pH 7.5-8.0, -20C) can be stored up to several years and remain viability (see attached article). You can even aliquot the vectors into a few tubes, so you can avoid thaw-and-freeze cycles.
2. For the bacteria itself:
According to American Type Culture Collection (ATCC): To insurance against phenotypic drift associated with prolonged passage (subcultures), due to genetic instability and/ or selective pressures, microorganisms for standard protocols should be used within FIVE passages (subcultures) of the ATCC reference culture.
See attached article from ATCC: How many passages are too many?
Just an information--
There are also services to help researches store their vectors, but only you have a cloning service with them.
For example: 'VectorArk Storage' from GenScript ( http://www.genscript.com/vector-archive.html ). It says "5 years of free storage for any plasmid vector." See more info below.
"....Whenever you ship a vector to us for custom cloning, we perform quality control by sequencing the MCS region of your vector and analyzing the restriction sites; this allows us to confirm the integrity of the reagents and the appropriateness of the cloning strategy before we start your cloning project. After delivering your final clone to you, we then securely store both the physical reagent and the relevant digital information for 5 years, free of charge to you for your convenience........"
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