Polymerase chain reaction (PCR) was performed with Amplitaq Gold mastermix (Thermofisher Willington, MA) using the following parameters; 95 °C, 10 min, 35 cycles of 95 °C, 30 s; 58 °C, 30 s; 72 °C, 60 s. PCR primers targeting the SSU rRNA were: 18f, 5′-CACCAGGTTGATTCTGCC-3′ and 1537r, 5′-TTATGATCCTGCTAATGGTTC-3′46 (link). The resultant amplicons were prepared using a Nextera XT library preparation kit and indices (Illumina inc, San Diego, CA). The samples were sequenced using an Illumina MiSeq system with a MiSeq V3 2 × 300 bp sequencing kit. The demultiplexed reads were quality trimmed to Q30 and assembled using CLC genomics workbench v20.0 (Qiagen inc., Valencia, CA). The consensus sequences for two full length SSU rRNA genes were accessioned in GenBank under MT367570-MT367571.
Nextera xt library preparation kit and indices
Manufactured by Illumina
The Nextera XT library preparation kit is a product offered by Illumina for the preparation of DNA libraries for next-generation sequencing. The kit includes reagents and indices necessary for the library preparation process. The core function of the Nextera XT kit is to enable the construction of sequencing-ready libraries from DNA samples.
Automatically generated - may contain errors
2 protocols using nextera xt library preparation kit and indices
1
Amplification and Sequencing of 16S rRNA
2
Amplification and Sequencing of SSU rRNA Genes
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Polymerase chain reaction (PCR) was performed with Amplitaq Gold mastermix (Thermo sher Willington, MA) using the following parameters; 95 °C, 10 min, 35 cycles of 95 °C, 30 s; 58 °C, 30 s; 72 °C, 60 s. PCR primers targeting the SSU rRNA were: 18f, 5′-CACCAGGTTGATTCTGCC-3′ and 1537r, 5′-TTATGATCCTGC TAATGGTTC-3′ 40 . The resultant amplicons were prepared using a Nextera XT library preparation kit and indices (Illumina inc, San Diego, CA). The samples were sequenced using an Illumina MiSeq system with a MiSeq V3 2 x 300 bp sequencing kit. The demultiplexed reads were quality trimmed to Q30 and assembled using CLC genomics workbench v20.0 (Qiagen inc., Valencia, CA). The consensus sequences for two full length SSU rRNA genes were accessioned in GenBank under MT367570-MT367571.
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