## 2 Materials and Methods
### Soil Enzyme Assay  
Soil enzymes involving C, N, P cycling were assayed, including \(\alpha\)-glucosidase (EC 3.2.1.20), \(\beta\)-glucosidase (EC 3.2.1.21), \(\beta\)-xylosidase (EC 3.2.1.37), cellbbiosidase (EC 3.2.1.91), urase (EC 3.5.1.5), nitrate reductase (EC 1.7.99.4), chitinase (E.C. 3.2.1.30), acid phosphatase (EC 3.1.3.2), alkaline phosphatase (EC 3.1.3.1). The activities of \(\alpha\)-glucosidase, \(\beta\)-glucosidase, \(\beta\)-xylosidase, cellbbiosidase and chitinase was assayed using fluorogenic substrates in micro-well plates as described previously [26; 27; 28]. Briefly, soil suspensions containing 1 g fresh soil in 50 mL sodium acetate buffer (0.5 mol L\({}^{-1}\), pH 5.5) were ultrasonically homogenized, and then 160 uL aliquot of each soil suspension was dispensed into 96-well black microplates (Jet Bio-Filtration Co., Guangzhou, China). Substrate solution (final concentration of 500 umol L\({}^{-1}\)) were added to each well with a final volume of 200 uL. Negative control and quench control were also applied. Standard curves were produced as well. Microplates were incubated for 3 h at 30 \({}^{\circ}\)C with continuous shaking. Fluorescence was measured using a microplate fluorometer (FLx800, BioTek Instruments, Winooski, VT, USA) at 355 nm excitation and 460 nm emission. The activities of urase, nitrate reductase, acid phosphatase and alkaline phosphatase were analyzed according to [PERSON] et al. [29] and [PERSON] et al. [22].  
Polymerase Chain Reaction-Denturing Gradient Gel Electrophoresis Analysis of Soil Microbial Community  
To compare the respective effect of CaCO\({}_{3}\) or OM on bacterial community, we performed PCR-DGGE analysis. The extraction of total soil DNA was conducted using PowerSoil(r) DNA isolation kit (MO BIO Laboratories Inc., Carlsbad, CA, USA) as described previously [22]. To amplify the V3 fragments of bacterial 16S rRNA genes, the nested PCR was performed using the primer sets of 27F/1492R: 5'-AGA GTT TGA TCC TGG CTC GA-3'/5'-TAC GGC TAC CTT GTT ACG ACT T-3' (the first round [30]) and 341F-GC/518R: 5'-CGC CCG CGCG CGCG GGC GGCGGCG GGG GCG GGCG GGG GCCG GGG GCC TAC GGG ACG CGA-3'/5'-ATT ACC GCG GCT GCT GG-3' (the second round [31]).  
Arbuscular mycorrhizal fungi (AMF) are symbiotic soil fungi, which greatly improve host plant growth. It is reported that _P. natatum_ can establish symbiosis with AMF with high colonization rate up to 99% [32]; therefore, we also probed the AMF community as affected by CaCO\({}_{3}\) or OM with PCR-DGGE analysis. To amplify the 18S rDNA fragments of AMF, the nested PCR was performed according to [PERSON] et al. [23], with NS1/NS4: 5'-GTA GTC ATA TGC TTG TCT C-3'/5'-CTT CCG TCA ATT CCT TTA AG-3' (the first round [33]), AML1/AML2: 5'-ATC AAC TTT CGA TGG TAG GAT AGA-3'/5'-GAA CCC AAA CAC TTT GGT TTC C-3' (the second round [34]), NS3-GC/Glo1: 5'-CGC CCG GCCG GCCG GGG GCCA CGG GGG TTG GAG GGC AAG TCT GGT GCC-3'/5'-GCC TGC TTT AAA CAC TCTA-3' (the third round [35]) as primer sets.  
The final PCR products of bacterial community and AMF community were subjected to DGGE analysis using a D-Code Universal Mutation Detection System (Bio-Rad Laboratories, Hercules, CA, USA). For the PCR product of bacteria, DGGE were carried out on an 8% polyacrylamide gel for 10 h at a constant voltage of 80 V and 60 \({}^{\circ}\)C in a 45-65% horizontal denaturant gradient [31]. For the PCR product of AMF, DGGE were carried out on a 6% polyacrylamide gel for 15 h at a constant voltage of 70 V and 60 \({}^{\circ}\)C in a 30-60% horizontal denaturant gradient [23].