Computational protocol: Rhodomycin analogues from Streptomyces purpurascens: isolation, characterization and biological activities

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Protocol publication

[…] A confirmatory taxonomic identification was done by the nucleotide sequencing of the 16S rRNA gene. Genomic DNA extraction, amplification and sequencing of the 16S rRNA gene were performed as described earlier (Mayilraj et al. ). The 16S rRNA gene was amplified with primers 8-27f (5′-AGAGTTTGATCCTGGCTCAG-3') and 1500r (5′AGAAAGGAGGTGATCCAGCCA-3′). The amplified DNA fragment was separated on 1% agarose gel, eluted from the gel and purified using QIAquick gel extraction kit (Qiagen). The purified PCR product was sequenced with 27f, 519r (5-GWATTACCGCGGCKGCTG-3), 1087r (5-CTCGTTGCGGGACTTAACCC-3), 530f (5-TTCGTGCCAGCAGCCGCGG-3), 945f (5-GGGCCCGCACAAGCGGTGG-3) and 1492r respectively (Escherichia coli numbering system). The rDNA sequence was determined by the dideoxy chain-termination method using the Big-Dye terminator kit using ABI 310 Genetic Analyzer (Applied Biosystems, USA). Almost complete sequence (1458 bp) of 16S rRNA of strain MTCC 8547 was determined and was compared with those of other closely related taxa retrieved from the GenBank database. A phylogenetic tree was constructed by Neighbour-Joining plot (Perrière & Gouy ). A sequence similarity search was done using GenBank BLASTN (Altschul et al. ). Sequences of closely related taxa were retrieved; aligned using Clustal X programme (Thompson et al. ) and the alignment was manually corrected. For the Neighbour-joining analysis (Saitou & Nei ), the distances between the sequences were calculated using Kimura’s two-parameter model (Kimura ). Bootstrap analysis was performed to assess the confidence limits of the branching (Felsenstein ). […]

Pipeline specifications

Software tools BLASTN, Clustal W
Application Phylogenetics
Organisms Bacillus subtilis
Diseases Drug-Related Side Effects and Adverse Reactions
Chemicals Carbohydrates