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Sample GSM2719415 Query DataSets for GSM2719415
Status Public on Nov 04, 2019
Title Naturally restored secondary forests_rep7 [NRSF7]
Sample type genomic
Source name Naturally restored secondary forests
Organisms Bacteria; Archaea; Fungi; Viruses
Characteristics site: Jiangxi Province Anfu County
dominant species: Cinnamomum camphora L. and Cunninghamia lanceolata (Lamb.) Hook.
Treatment protocol The forests have been restored for about tweenty years. The restored forests were naturally restored secondary forests (NRSF), artificial reforestation with native Masson pine (Pinus massoniana Lamb.) (ARMP), and introduced slash pine (Pinus elliottii Engelm.) plantations (ARSP). For each forest restoration method, 15 plots (500 m × 500 m) were studied. Fifteen soil cores were collected from each plot, and then combined to create a composite sample.
Growth protocol Soil samples were collected from the top 10 cm in October 2008 and August, September 2009. Each sample was composition of fifteen soil cores (3.5 cm diameter). All samples were sieved through 2 mm sieves, and stored at -80 °C.
Extracted molecule genomic DNA
Extraction protocol Soil DNA was extracted by phenol/chloroform method with Al2(SO4)3 pretreatment (Peršoh et al. 2008). Firstly, 0.5 g soil was washed with an excess Al2(SO4)3 (0.2 M) which was subsequently removed with a 4 M NaOH solution. Secondly, DNA and protein were extracted from soil with Tris-HCl buffer (0.4 M LiCl, 100 mM Tris–HCl, 120 mM EDTA, pH 8) and 0.5 g glass beads by glass beading. Thirdly, DNA and protein were separated with phenol:chloroform:alcohol (25:24:1) and chloroform:isoamyl alcohol (24:1), and the supernatant was washed with chloroform overnight. Fourthly, DNA was precipitated twicely with 70% cold ethanol at 13000 rpm for 60 min and 15 min, respectively (Peršoh et al. 2008). (D. Peršoh, S. Theuerl, F. Buscot, G. Rambold, Towards a universally adaptable method for quantitative extraction of high-purity nucleic acids from soil. J. Microbiol. Meth. 75, 19-24 (2008)).
Label Cy5
Label protocol The DNA was labeled by Cy5 as described (Xue et al. 2007). Firstly, the DNA was mixed with 20 μL random primers, denatured at 99.9 °C for 5 min, and chilled on ice. Secondly, the labeling master mix (2.5 μL dNTP (5 mM dAGC-TP, 2.5 mM dTTP), 1 μL Cy-5 dUTP (Amersham, Piscataway, NJ), 80 U of the large Klenow fragment (Invitrogen, Carlsbad, CA), and 2.5 μL water) were added, incubated at 37°C for 3 h, and then heated at 95 °C for 3 min. Thirdly, the labeled DNA was purified with the QIA quick purification kit (Qiagen, Valencia, CA), measured on a NanoDrop ND-1000 spectrophotometer (NanoDrop Technologies Inc., Wilmington, DE), and then dried in a SpeedVac (ThermoSavant, Milford, MA) at 45 °C for 45 min (Xue et al. 2007). (K. Xue, M.M. Yuan, J.P. Xie, D. Li, Y. Qin, L. E. Hale, L. Wu, Y. Deng, Z. He, J. D. Van Nostrand, Y. Luo, J. M. Tiedje, J. Zhou. Annual removal of aboveground plant biomass alters soil microbial responses to warming. mBio 7, 1-12 (2016)).
Hybridization protocol The labeled DNA was suspended in 120 µl hybridization solution (50% formamide, 3 × SSC, 10 µg of unlabeled herring sperm DNA (Promega, Madison, WI), and 0.1% SDS. The mix was kept at 95°C for 5 min and kept at 50°C until deposited onto a microarray. Hybridizations were performed with a TECAN Hybridization Station HS4800 Pro (TECAN, US) (Xue et al. 2007) (K. Xue, M.M. Yuan, J.P. Xie, D. Li, Y. Qin, L. E. Hale, L. Wu, Y. Deng, Z. He, J. D. Van Nostrand, Y. Luo, J. M. Tiedje, J. Zhou. Annual removal of aboveground plant biomass alters soil microbial responses to warming. mBio 7, 1-12 (2016)).
Scan protocol The hybridized GeoChip 3.0 was scanned with a ScanArray 5000 (Perkin-Elmer, Wellesley, MA, USA) at 95% laser energy and 85% photomultiplier tube gain, and digitally analyzed using Imagene software (6.0 premium version, Biodiscovery, El Segundo, CA, USA)
Data processing The data were processed as follows: (i) Probes flagged as 1 or 3 by ImaGene and probes with signal to noise ratio < 2.0 were removed; (ii) Data were normalized firstly within a slide,then among samples,and finally devided by the mean of the signal intensities of all probes; (iii) Probes appeared less than 3 times for each reforestation approach were deleted; (iv) Samples with probe numbers outside of mean ± (2 × standard error) were considered outliers (ARMP1, NRSF2, and NRSF7) and removed. In total, 14, 14, and 11 microarray replicates were obtained for ARSP, ARMP, and NRSF, respectively.
Submission date Jul 27, 2017
Last update date Nov 04, 2019
Contact name Yun Wang
Organization name Research Center for Eco-Environmental Sciences, Chinese Academy of Sciences
Street address Shuangqing Road 18th, Haidian District
City Beijing
ZIP/Postal code 100085
Country China
Platform ID GPL23836
Series (1)
GSE100379 Artificial reforestation has less diverse soil nitrogen-cycling genes than natural restoration

Data table header descriptions
VALUE Normalized signal intensity

Data table
110551987 0.3398
154151622 0.2392
20977654 0.2945
780709 0.4135

Total number of rows: 6378

Table truncated, full table size 76 Kbytes.

Supplementary file Size Download File type/resource
GSM2719415_NRAF2.xlsx.gz 3.6 Mb (ftp)(http) XLSX
Processed data included within Sample table

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