DOI: 10.5061/dryad.9cnp5hqf0
title: Data from: Diversity and structural variability of bacterial microbial communities in rhizocompartments of desert leguminous plants

description: By assessing diversity variations of bacterial communities under different
 rhizocompartment types (i.e., roots, rhizosphere soil, root zone soil, and
 inter-shrub bulk soil), we explore the structural variability of bacterial
 communities in different root microenvironments under desert leguminous
 plant shrubs. 16S rRNA high-throughput genome sequencing was used to
 characterise the bacterial diversity in the rhizocompartments of three
 xeric leguminous plants (Caragana microphylla, Hedysarum scoparium, and
 Hedysarum mongolicum) in the Mu Us Desert.
description: 16S rRNA genome sequencing and bioinformatics
 analysis E.Z.N.A. soil DNA kits (OMEGA, the USA) were
 used with 0.5 g of fresh sample to extract DNA samples, as per the
 manufacturer’s instructions. All samples were stored at -80 °C. After
 thawing on ice, extracted DNA samples were separately centrifuged and
 fully mixed; sample quality was determined using a NanoDrop instrument,
 and 30 ng DNA was used for PCR amplification. PCR amplification was
 performed in 25 μL reaction volumes containing 10×PCR buffer, 0.5 μL
 dNTPs, 1 μL of each primer, 3 μL bovine serum albumin (2 ng/μL), 12.5 μL
 KAPA 2G Robust Hot Start Ready Mix, ultrapure H<sub>2</sub>O,
 and 30 ng template DNA. The PCR amplification program included initial
 denaturation at 95 °C for 5 min, followed by 25 cycles of denaturing at 95
 °C for 45 s, annealing at 55 °C for 50 s, and extension at 72 °C for 45 s.
 Finally, the PCR amplification program was completed at 4 °C. Forward
 primer F799 (5′-AACMGGATTAGATACCCKG-3′) and reverse primer R1193
 (5′-ACGTCATCCCCACCTTCC-3′) were used to target the V5-V7 regions of 16S
 rRNA. Both primers contained Illumina adapters, and the forward primer
 contained an 8 bp barcode sequence unique to each sample. An Agarose Gel
 DNA purification kit (Axygen Biosciences, Union City, CA, the USA) was
 used for the purification and combination of PCR amplicons. After
 purification, PCR amplicons were mixed at an equal molar concentration,
 followed by pair-end sequencing using the Illumina MiSeq sequencing system
 (Illumina, the USA) according to a standardized process.
 MiseqPE300 (Illumina, the USA) sequencing results were recorded
 in the Fastq format. Quantitative insights into microbial ecology software
 (QIIME; Version 1.8 http://qiime.org/) was used to analyze original Fastq
 files and to undertake quality control according to the following criteria
 : (i) base sequences with a quality score &lt;20 at read tails were
 removed, and the window was set at 50 bp. When the mean quality score in
 the window was &lt;20, posterior-end base sequences were discarded
 from the window and reads shorter than 50 bp were removed after quality
 control; (ii) paired reads were assembled into one sequence according to
 the overlapping relationship between reads (minimum overlapping length: 10
 bp); (iii) the maximum allowable mismatch ratio of the overlapping areas
 of assembled sequences was set to 0.1, and sequences failing to meet this
 criterion in pairs were removed; (iv) samples were distinguished according
 to barcodes and primers at the head and tail ends of sequences, and the
 sequence directions were adjusted based on the number of mismatches
 allowed by barcodes (0); and (v) different reference databases were
 selected according to the type of sequencing data. Chimeras were removed
 using the Usearch program V8.1861 (http://www.drive5.com/usearch/), and
 clean tags of high-quality sequences were acquired after smaller-length
 tags were discarded using mothur software. Sequences were clustered into
 Operational Taxonomic Units (OTUs) using UPARSE V7.1
 (http://drive5.com/uparse/) based on a 97% sequence similarity cutoff
 (excluding single sequences). Representative sequences and an OTU table
 were obtained.