Computational protocol: Streptomyces luridus So3.2 from Antarctic soil as a novel producer of compounds with bioemulsification potential

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Protocol publication

[…] Genomic DNA was extracted using the UltraClean® Microbial DNA Isolation Kit (MOBIO, CA, USA) according to the manufacturer’s instructions. 16S rDNA was selectively amplified from genomic DNA by polymerase chain reaction (PCR) using universal primers 27f (5’-AGAGTTTGATCCTGGCTCAG-3’) and 1492r (5’-GGTTACCTTGTTACGACTT-3’), enabling the amplification of approximately 1.500bp of the 16S rRNA gene. PCR amplification was performed in a Multigene Optimal Thermal Cycler (Labnet, USA) in 50 μl of PCR mix comprising 25 μl mix reaction buffer 2x (SapphireAmp Fast PCR Master Mix, Takara), 22 μl ultra-pure water, 1 μl of each primer (10 μM) and 1 μl of DNA. The temperature and cycling conditions were as follows: preheating at 94°C for 2 min; 30 cycles at 94°C for 1 min; 55°C for 1 min; 72°C for 1.5 min; and incubation at 72°C for 10 min. The presence of PCR products were assessed by electrophoresis on a 1% agarose gel stained with gel red. Sequencing was done with a dye Terminator Cycle Sequencing Kit and an ABI 3730XL DNA Sequencer (Applied Biosystems) by Macrogen (Korea). The nearest taxonomic group was identified by 16S rDNA nucleotide sequence BLASTN (http://www.ncbi.nlm.nih.gov/blast) using DDBJ/EMBL/GenBank nucleotide sequence databases.A phylogenetic tree was constructed in MEGA7 [] using the neighbor-joining method []. The bootstrap consensus inferred from 1000 replicates [] was taken to represent the evolutionary history of the taxa analyzed []. The evolutionary distances were computed using the maximum composite likelihood method []. The nucleotide sequence identified in this study was deposited in the NCBI nucleotide sequence database (GenBank/NCBI) under accession number MH070262. […]

Pipeline specifications

Software tools BLASTN, MEGA
Databases DDBJ
Application Phylogenetics
Chemicals Carbon, Glucose, Glycerol, Hydrocarbons