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56 PROCEEDINGS OF INTERNATIONAL SCIENTIFIC CONFERENCE ON APPLIED BIOTECHNOLOGYcorresponding to herboxidiene (Fig. 6; Fig. 7) and further analysis of the sample with HR-QTOF mass analysis showed the fragmentation pattern which matches with standard herboxidiene (Fig. 7). While looking for the possible reasons we encountered many instances in PKS biosynthetic clusters that has been studied with extra DH domain present in the pathway. This kind of phenomenon happens due to duplication and insertion of a full or partial module cassette to fulfill the role of missing domains. Few of the examples are FK 506 (2 DH), rifamycin (6, 7, and 8 DH) and epothilone (7, and 8 DH) [21,22]. To study herboxidiene PKS in such detail, the DH domain should be compared with other modules from different PKS of S. chromofuscus or the substrate-dehydratase interrogation must be done as suggested for studying the DH activity of curacin PKS [21].Another possible reason could be that double crossover does not occur in the mutant strain, even though it loses viability on the apramycin plate. The single-crossover strain may produce herboxidiene and new products. Detailed genetic analysis of the mutant strain should be conducted through genome sequencing in future study. 5. CONCLUSIONSThe functional inactivation of module 8 DH domain was confirmed. The functional inactivation of module 8 DH was found not to inhibit the THP ring formation as expected. We found the corresponding mass number of one expected compound in case of module 8 DH domain, however the structure needs further analysis and elucidation. Conflicts of Interest: The authors declare no conflict of interest.