sample site: Fengqiu soil type: Inceptisol landuse of bare fallow, maize cropping or npk fertilization: bare soil
Treatment protocol
In August-September 2011, soil samples were collected from all of the plots. Ten soil cores at a depth of 0-20 cm and diameter of 1.5 cm were randomly taken from each plot and mixed thoroughly to generate a soil sample representing the plot. Soil samples were kept on ice when transporting to laboratory, and sieved with 2 mm mesh to remove roots and stones. Soil samples were preserved at -80°C before DNA extraction.
Extracted molecule
genomic DNA
Extraction protocol
Soil genomic DNA was extracted using a FastDNA spin kit for soil (MP Biomedical, Carlsbad, CA, USA) following the manufacturer’s instructions. To purify it, DNA extract was mixed with 2.5 volume of 100% ice cold ethanol and 0.1 volume of 3 M NaOAc (pH 5.2) prior to overnight incubation at -20°C. DNA was precipitated by centrifugation for 30 minutes at 13,000 x g. Then supernatant was decanted and washed with 1 ml of 70% ethanol. DNA was air-dried and dissolved in 50 µl of nuclease-free water. DNA quality and quantity were measured using NanoDrop ND-1000 Spectrophotometer (NanoDrop Technologies Inc., Wilmington, DE, USA) and with PicoGreen (Ahn et al 1996) using a FLUOstar Optima (BMG Labtech, Jena, Germany), respectively.
Label
Cy5
Label protocol
As previously described (Yang et al 2013), DNA samples were labeled with the fluorescent dye Cy-5 using a random priming method and purified using the QIA quick purification kit (Qiagen, Valencia, CA, USA).
Hybridization protocol
Then DNA was dried in a SpeedVac (ThermoSavant, Milford, MA, USA) at 45°C for 45 minutes. The hybridization was carried out at 42°C for 16 hours on a MAUI hybridization station (BioMicro, Salt Lake City, UT, USA).
Scan protocol
After purification, GeoChip microarrays were scanned by a scanner (MS 200 Microarray Scanner, NimbieGen) using a laser power and photomultiplier tube (PMT) gain of 100% and 75%, respectively.
Description
replicate 3
Data processing
Signal intensities were quantified and processed using the data analysis pipeline as previously described (He et al 2010). Then processed GeoChip data were analyzed using the following steps: (i) remove the poor quality spots, which were flagged as 1 or 3 by ImaGene or with a signal to noise ratio (SNR) of less than 2.0; (ii) the data were then logarithmically transformed and divided by the mean value of each slide.; (iii) transform the data to the natural logarithmic form; and (iv) remove genes detected in only one out of three samples from the same elevation.
A comprehensive study to understand the effects of soil type, soil transplant and landuse changes on soil microbial community and its feedback responses
