Abstract: Molecular-based diagnostic kits and methods for detecting nucleotide sequences of DNA, DNA copy and RNA of infectious agents, transgenes, alleles or non-encoding sequences, with no initial isolation are provided. The kit comprises at least a first calibrated dropper bottle with an anti-contamination system containing a solution of polymerase enzyme, chaotropic agents, salts and deoxyribonucleotides; a second calibrated dropper bottle with an anti-contamination system containing a set of at least 4 primers, part of the sequences being preserved in the nucleic acid fragment to be detected; a dropper bottle containing a mixture of developing reagents; a carrier to hold the sample; at least one LFD developing system, colorimetric or fluorogenic reaction; positive and negative controls. The polymerase enzyme is selected from Bst enzymes formed by DNA polymerase/helicase, with no exonuclease activity.