TY - JOUR KW - Animals KW - Base Sequence KW - Biopsy KW - Culture Media KW - DNA, Bacterial KW - DNA, Ribosomal KW - Endopeptidase K KW - Freezing KW - Hot Temperature KW - Humans KW - leprosy KW - Mice KW - Molecular Sequence Data KW - Mycobacterium leprae KW - polymerase chain reaction KW - Preservation, Biological KW - RNA, Bacterial KW - RNA, Ribosomal, 16S KW - Sensitivity and Specificity KW - Serine Endopeptidases KW - Skin KW - Species Specificity AU - Pattyn S R AU - Ursi D AU - Ieven M AU - Raes V AU - Jamet P AB -

The sensitivity of the polymerase chain reaction (PCR) on the DNA coding for the species-specific fragment of 16S rRNA of Mycobacterium leprae studied on mouse foot pad harvests and human skin biopsies varies widely between 1 and 3 x 10(4) organisms. This is probably the result of variations in the proportions of organisms with sufficiently intact DNA suitable for PCR. Preserving human skin biopsies for 3 weeks at an ambient temperature even after boiling for 6 minutes gives rise to a 10-fold decrease in sensitivity. Fixation of tissues in formol 10% or Lowy fixative or preserving in Dubos OAA broth is very harmful to the PCR, mainly due to the enhancement of an inhibitory effect on the PCR reaction. For preservation, the best choice at the moment seems to be alcohol 70%. Sample preparation of five cycles of freeze-boiling is simple and generally more efficient than proteinase K treatment and DNA extraction.

BT - International journal of leprosy and other mycobacterial diseases : official organ of the International Leprosy Association C1 -

http://www.ncbi.nlm.nih.gov/pubmed/1381740?dopt=Abstract

DA - 1992 Jun IS - 2 J2 - Int. J. Lepr. Other Mycobact. Dis. LA - eng N2 -

The sensitivity of the polymerase chain reaction (PCR) on the DNA coding for the species-specific fragment of 16S rRNA of Mycobacterium leprae studied on mouse foot pad harvests and human skin biopsies varies widely between 1 and 3 x 10(4) organisms. This is probably the result of variations in the proportions of organisms with sufficiently intact DNA suitable for PCR. Preserving human skin biopsies for 3 weeks at an ambient temperature even after boiling for 6 minutes gives rise to a 10-fold decrease in sensitivity. Fixation of tissues in formol 10% or Lowy fixative or preserving in Dubos OAA broth is very harmful to the PCR, mainly due to the enhancement of an inhibitory effect on the PCR reaction. For preservation, the best choice at the moment seems to be alcohol 70%. Sample preparation of five cycles of freeze-boiling is simple and generally more efficient than proteinase K treatment and DNA extraction.

PY - 1992 SP - 234 EP - 43 T2 - International journal of leprosy and other mycobacterial diseases : official organ of the International Leprosy Association TI - Polymerase chain reaction amplifying DNA coding for species-specific rRNA of Mycobacterium leprae. UR - http://ila.ilsl.br/pdfs/v60n2a09.pdf VL - 60 SN - 0148-916X ER -