Using the kit LowCell# ChIP I got very high background; even with SAT2 I got the band performing end-point PCR. What can be the problem?
There are several important points: • You performed end-point PCR – 45 cycles. This is too much; it is not possible to correctly quantify the intensity of bands as PCR is saturated. If you want to continue with end-point PCR, you should try to find semi-quantitative conditions by decreasing the number of amplification cycles. It is better to use qPCR for the analyse of ChIP results. • Be aware that there is not cross-contamination between samples: pay attention working with strips and separate caps always putting the caps to corresponding tubes. If possible, work with barrier tips. • SAT2 primers amplify repeated genomic sequences that make them very sensitive for contamination. Even small traces of human DNA are enough to provoke this contamination. DNA free barrier tips, sterile hood and gloves are recommended for working with this primers. We are aware of this problem that’s why we are looking for a new control which is not based on repeated sequences. • Magnetic beads are known to
