Dear Bhawana Ghimire

contaminations are omnipresent in molecular labs and become visible when using amplification methods or methods with high sensitivity. Investigations on the source is often tidious and without sufficient result. There is some actions you can take for PCR. First, use small aliquots for all ingredients including primer dilutions. If you see contaminations you can easily dispose and prepare new. Always keep one primer stock that is only touched once for first dilution. For preparation, use pipettes, diluents (water) and tips (best filter tips) only dedicated to this purpose. If possible try to separete pre and post PCR sections in the lab. Keep in mind that autoclavation does not destroy plasmids/vectors and these are often a source of conamination. Hence, glaswear used for PCR water etc should also be heated/ baked before use. Last but not least, use negative PCR controls (as you did). In your case you should be able to freshly prepare all components used for your PCR and you should be fine. However, some "contaminations" are primer dimers and you should check amplicon size and the gardient you use.

Good luck

Boris