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welcome to nikolais genetics lessons tips and tricks and today we are going to talk why your gel missing a band or have faint bend in many cases this is not a failure of your jaw electrophoresis but of the pcr and the reason number one to few cycles were used using too few pcr cycles can lead to insufficient amplification use 20 35 cycles use fewer cycles when template concentration is higher and use more cycles when template concentration is low and the reason number two extension time was too short if the extension time is too short there will be sufficient time for complete replication of the target generally use extension time one minute per kilobase and the reason number three annealing time was too short if the annealing time is too short primers do not have enough time to bind to the template use annealing time for at least 30 seconds