Were collected 28 soil samples in 14 municipalities in Colombia. Sporulated bacillus were isolated, characterized microscopically by quantifying crystals. Isolates positive for crystals were subjected to biochemical characterization by electrophoresis of total proteins (SDS-PAGE). The strains positive for the presence of cry1 genes were subjected to two rounds of M-PCR with two oligonucleotide mixtures, recognizing specific genes 6. 99 bacillus were isolated forms filed with amorphous crystals, bipiramidal, square, round and triangular. Bacillus were observed with 1, 2, 3 and 4 forms of glass, with 18 different profiles. For SDS, showed protein bands of 28 to 150 kDa in 7 classified by their potential biological activity, resulting in 28 different profiles. 35 sporulated bacillus showed cry1 genes, and these were detected by M-PCR gene cry1Aa, cry1Ab, cry1Ac, cry1B, cry1C and cry1D in 76, 26, 21, 35, 32, and 8.8% respectively. According to these characterizations 10 isolates were selected as promising for the control of Tuta absoluta and challenged against the 2nd instar larvae of this insect pest. Strain ZBUJTL39 and introduced ZCUJTL11 better biological activity that the reference strain Bt. var kurstaki HD1. strain ZCUJTL11 presented a LC50 of 2.4 mg / ml (P <0.05). The methodology selected strains according to their potential biological activity, as a preliminary step to biological tests, this result is promising for further research into genetic engineering of tomato cultivars to obtain resistant to Tuta absoluta.