The vertical electrophoresis apparatus used was P8DS™ Emperor Pen

The vertical electrophoresis apparatus used was P8DS™ Emperor Penguin (Owl, Thermo Scientific) with an adaptor for Lonza precast gels. The run was performed at 100 V in TBE 1X. Diagnostic key A dichotomous key was developed comparing in silico digestion results and the evaluation of visible bands with the use of ImageLab™ 2.0 software (Bio-Rad Laboratories, Inc.). Results and discussion In silico analysis The analysis and comparison of restriction profiles

obtained with in silico digestion of bifidobacterial hsp60 sequences selleck products allowed the identification of a set of appropriate frequent-cutter endonucleases that recognize non degenerated sequences. The restriction enzyme HaeIII was found to give the clearest and most discriminatory profiles in theoretical PCR-RFLP patterns, discriminating the majority of Bifidobacterium type-strains

selleck chemical tested (Table  SRT1720 research buy 3). Furthermore, the profiles of other strains, belonging to the investigated species, have been analyzed to confirm the conservation of RFLP profiles within species. Table 3 Expected fragment sizes obtained with in silico digestion of the hsp60 gene sequences Bifidobacterium species GenBank entry Predicted fragment sizes Profile B. adolescentis AF210319 31-36-81-103-339   B. angulatum AF240568 42-54-59-139-296   B. animalis subsp. animalis AY004273 17-53-86-97-114-223   B. animalis subsp. lactis AY004282 71-86-96-114-223   B. asteroides AF240570 30-38-75-97-109-242   B. bifidum AY004280 22-31-59-181-297   B. boum AY004285 22-117-200-251   B. breve AF240566 106-139-139-200   B. catenulatum AY004272 53-198-338   B. choerinum AY013247 36-42-51-52-54-59-97-200   B. coryneforme AY004275 PFKL 16-32-54-158-338   B. cuniculi AY004283 16-42-53-70-128-281   B. dentium AF240572 22-31-42-68-130-139-158   B. gallicum AF240575 42-253-297   B. gallinarum AY004279 16-31-42-81-139-281   B. indicum AF240574 16-32-36-42-45-123-296   B. longum subsp. longum AF240578 42-113-138-139-158 * B. longum subsp. infantis AF240577 42-113-138-139-158 * B. longum subsp. suis AY013248 42-113-138-139-158 * B. merycicum

AY004277 22-31-42-59-139-297   B. minimum AY004284 16-51-60-66-70-327   B. pseudocatenulatum AY004274 42-53-198-297   B. pseudolongum subsp pseudolongum AY004282 17-22-30-32-42-42-109-297   B. pseudolongum subsp. globosum AF286736 16-17-22-30-32-42-109-323   B. pullorum AY004278 16-31-36-42-81-87-297   B. ruminantium AF240571 31-106-114-339   B. subtile Not available Not avaiable + B. thermacidophilum subsp porcinum AY004276 20-42-53-59-97-139-180 *† B. thermacidophilum subsp thermacidophilum AY004276 20-42-53-59-97-139-180 *† B. thermophilum AF240567 54-59-117-139-222   + hsp60 sequence of B. subtile type strain was not available in the press-time. † the available sequences at GeneBank and cpnDB belonged to B. thermacidophilum (with no distinction in subspecies). *subspecies not discernable.

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