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Brilliant sybr green 2 mastermix

Manufactured by Agilent Technologies

Brilliant SYBR Green II Mastermix is a ready-to-use solution for quantitative real-time PCR (qPCR) experiments. It contains SYBR Green II dye, a DNA-binding fluorescent dye, along with all the necessary components for efficient and sensitive qPCR amplification.

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2 protocols using brilliant sybr green 2 mastermix

1

Bacterial 16S rRNA Gene Sequencing

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DNA was extracted from the filters using the DNeasy PowerSoil Extraction kit (Qiagen) following the manufacturer’s protocol. Filters were cut into smaller pieces to facilitate cell disruption during the bead-beating step. Amplicon sequencing of bacterial 16S rRNA genes was carried out targeting the V3-V4 region with the primer combination Bakt_0341F/Bakt_0785R [25 (link)]. PCR amplification, library preparation, and sequencing on an Illumina MiSeq platform using v3 chemistry was performed at LGC (Berlin) as previously described [26 (link)]. Abundances of bacterial 16S rRNA genes were determined by quantitative PCR using Brilliant SYBR Green II Mastermix (Agilent Technologies) on a Mx3000P system (Agilent Technologies) and the primer combination Bakt_0341F [25 (link)] and Bakt_0799R, which is the reverse complement version of the primer 799F [27 (link)] discriminating against chloroplast-derived 16S rRNA genes. For qPCR, we used the following cycling conditions: 10 min at 95 °C, followed by 45 cycles of 30 s at 95 °C, 30 s at 53 °C, and 40 s at 72 °C, and subsequent melting curve analysis. Due to loss of plant material of some samples before determination of leaf dry weight, abundance data are only available for a subset of all samples (see Supplementary Table 2).
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2

Bacterial Community Analysis via 16S rRNA

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DNA was extracted from the filters using the DNEasy PowerSoil Extraction kit (Qiagen) following the manufacturer's protocol. Filters were cut into smaller pieces to facilitate cell disruption during the bead-beating step. Amplicon sequencing of bacterial 16S rRNA genes was carried out targeting the V3-V4 region with the primer combination Bakt_0341F/Bakt_0785R (Klindworth et al. 2013) . PCR amplification, library preparation, and sequencing on an Illumina MiSeq platform using v3 chemistry was performed at LGC (Berlin) as previously described (Rughöft et al. 2016) (link). Abundances of bacterial 16S rRNA genes were determined by quantitative PCR using Brilliant SYBR Green II Mastermix (Agilent Technologies) on a Mx3000P system (Agilent Technologies) and the primer combination Bac8Fmod (Nercessian et al. 2005) (link) and Bac338Rabc (Loy et al. 2002) (link) as previously described (Herrmann et al. 2012) (link). Due to loss of plant material of some samples before determination of leaf dry weight, abundance data are only available for a subset of all samples (see Supplementary Table 1).
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