Nanodrop 2000 nucleic acid analyzer

β-D-glucosidase was purchased from Shanghai Yuanye Biotechnology Co., Ltd., Shanghai, China; methanol (chromatographic pure), ethyl acetate (chromatographic pure), citric acid, disodium hydrogen phosphate, and anhydrous sodium carbonate were obtained from Tianjin Damao Chemical Reagent Factory, Tianjin, China; concentrator plus vacuum concentrator, ultra-low temperature refrigerator, electronic balance, ultrasonic cleaner, pipette gun were obtained from Eppendorf, Hamburg, Germany; thermostatic bath was obtained from PolyScience, Niles, IL, USA; acid meter, Trace1300 gas chromatograph, Trace ISQ mass spectrometer (GC-MS), and NanoDrop 2000 nucleic acid analyzer were obtained from Thermo, Waltham, MA, USA; high-efficiency plant total RNA extraction kit, UnionScript First-strand cDNA synthesis mix for qPCR kit, and real-time fluorescence quantitative SYBR GreenⅠMaster kit were purchased from Beijing Jinsha Biological Co., Ltd., Beijing, China; LightCycler 480 quantitative PCR instrument was obtained from Roche, Basel, Switzerland.

Rice roots from the pot were obtained for bacterial 16sRNA gene profiling. Root sampling was performed at the heading stage and washed until there were no visible soil particles. Next, 10 cm long roots from the ground were sliced into 2 mm sections and placed in a 2-ml tube. Three replicates were performed for each sample. Total DNA extraction from root bacteria was performed using the BioFast soil Genomic DNA Extraction kit (BioFlux, Hangzhou, China), as instructed by the manufacturer. Total DNA from root bacteria were detected by 1% agarose electrophoresis and their concentrations determined by the NanoDrop 2000 nucleic acid analyzer (Thermo Scientific, USA). The V5-V7 region of bacterial 16 S ribosomal RNA gene was amplified using 799 F (AACMGGATTAGATACCCKG) and 1193R (ACGTCATCCCCACCTTCC) primers. The PCR assay was performed using Trans Start Fastpfu DNA Polymerase (TransGen Biotech, China). After amplification, PCR products were identified by electrophoresis on 2% agarose gel and recovered using the AxyPrepDNA Gel Recovery Kit (Axygen Bioscience, China) after elution using Tris-HCl. The library was sequenced on a HiSeq 2500 platform (Illumina, San Diego, CA, USA).