1. A composition for amplification of DNA sequences of Mycobacterium tuberculosis
complex from different biological samples comprising a master mix, a primer mix
comprising forward and reverse oligonucleotide primers for binding to specific genes
5 consisting of IS6110 (genbank id X17348.1, SEQ.ID.NO.18), rpoB (genbank id
Rv0667, SEQ.ID.NO.19) and gyrB (genbank id Rv0005, SEQ.ID.NO.20) of
Mycobacterium tuberculosis complex and enhancer; wherein the master mix is in a
range between 70% -75% v/v, the primer mix is in a range between 19% - 25% v/v, the
enhancer is in a range between 5% - 6% v/v of the composition.
10 2. The composition of claim 1, wherein the primer mix comprises:
forward oligonucleotide primers consisting of SEQ.ID.Nos: 1, 3 which bind to IS6110
of SEQ.ID.NO.18 ;
reverse oligonucleotide primers consisting of SEQ.ID.Nos: 2, 4, 5 which bind to IS6110
of SEQ.ID.NO.18;
15 forward oligonucleotide primers consisting of SEQ.ID.Nos: 6, 8, 10 which bind to rpoB
of SEQ.ID.NO.19;
reverse oligonucleotide primers consisting of SEQ.ID.Nos: 7,9,11 which bind to rpoB
of SEQ.ID.NO.19;
forward oligonucleotide primers consisting of SEQ.ID.Nos: 12,14,16 which bind to
20 gyrB of SEQ.ID.NO.20;
reverse oligonucleotide primers consisting of SEQ.ID.Nos: 13,15,17 which bind to
gyrB of SEQ.ID.NO.20.
3. The composition of claim 1, wherein the primer mix comprises forward and reverse
oligonucleotide primers for binding to IS6110 (genbank id X17348.1, SEQ.ID.NO.18),
25 rpoB (genbank id Rv0667, SEQ.ID.NO.19), gyrB (genbank id Rv0005,
SEQ.ID.NO.20) of Mycobacterium tuberculosis complex in the ratio of 45:50:5 v/v.
4. The composition of claim 1, wherein the master mix comprises DNA polymerase,
preferably Bst polymerase, deoxynucleotide triphosphates (dNTPs), and reaction
buffer.
30 5. The composition of claim 4, wherein the reaction buffer comprises Tris-Hydrochloride,
Ammonium sulfate, Potassium Chloride, Magnesium Sulfate, Tween 20.
6. The composition of claim 4, wherein the deoxynucleotide triphosphates (dNTPs),
consist of deoxyadenosine triphosphate, dATP; deoxythymidine triphosphate, dTTP;
22
deoxycytosine triphosphate, dCTP; and deoxyguanosine triphosphate, dGTP in equal
proportion.
7. The composition of claim 1, wherein the Mycobacterium tuberculosis complex is one
of genetically related Mycobacterium species that cause tuberculosis, comprising
5 Mycobacterium tuberculosis, Mycobacterium africanum, Mycobacterium orygis,
Mycobacterium bovis, Mycobacterium microti, Mycobacterium canetti,
Mycobacterium caprae, Mycobacterium pinnipedii, Mycobacterium suricattae,
Mycobacterium mungi or a combination thereof.
8. The composition of claim 1, wherein the enhancer comprises Guanidine Hydrochloride.
10 9. The composition of claim 1, wherein the biological sample comprises human sputum
samples or human tissue samples.
10. A method for identification of Mycobacterium tuberculosis complex from different
biological samples using the naked eye, comprising:
 extraction of DNA from biological samples;
15  amplification of said extracted DNA by mixing the extracted DNA with the
composition for amplification of DNA sequences of Mycobacterium
tuberculosis complex to form a reaction mixture and incubating the reaction
mixture at 65°C for 35-45 minutes; wherein the composition for amplification
of DNA sequences of Mycobacterium tuberculosis complex comprises a master
20 mix, a primer mix comprising forward and reverse oligonucleotide primers for
binding to specific genes consisting of IS6110 (genbank id X17348.1,
SEQ.ID.NO.18 ), rpoB (genbank id Rv0667, SEQ.ID.NO.19) and gyrB
(genbank id Rv0005, SEQ.ID.NO.20) of Mycobacterium tuberculosis complex
and enhancer; wherein the master mix is in a range between 70% -75% v/v, the
25 primer mix is in a range between 19% -25% v/v, the enhancer is in a range
between 5% - 6% v/v;
 cooling the reaction mixture to 25℃ to 28℃;
 visualization of amplified DNA using pH sensitive indicator dye comprising
Phenol Red wherein the yellow color indicates a positive result, while the pink
30 color indicates a negative result for Mycobacterium tuberculosis complex