16S rRNA-Based PCR-DGGE Analysis of Actinomycete Communities in Fields with Continuous Cotton Cropping in Xinjiang, China

Abstract

The purpose of this study was to examine the variations in the microbial community structure of soil actinomycetes in fields with continuous cropping of cotton in Xinjiang Autonomous Region, China. Soil samples were collected from four depths in fields with 7-year continuous cotton cropping. The community structure of soil actinomycetes was examined using the 16S rRNA-based polymerase chain reaction-density gradient gel electrophoresis (PCR-DGGE) techniques. The microbial diversity indices of the soil samples from different depths generally decreased along with the period of continuous cotton cropping. When the period of continuous cropping of cotton reached 5 years, the diversity indices rose again and gradually stabilized at a level slightly lower than that of soils with original ecology (i.e., 0-year cotton cropping). Cluster analysis showed that at the 1-20-cm depth, the actinomycete community structure of the soil subjected to 1-year cotton cropping was similar to that of soil subjected to 0-year cotton cropping, whereas that of soils after 3-year continuous cotton cropping showed high similarity. At the 21-40-cm depth, the actinomycete community structure showed various changes but generally recovered to its original pattern after repeated fluctuations. Principal component analysis showed that at the 1-30-cm depth, the actinomycete community structure varied similarly regardless of the period of continuous cotton cropping. In contrast, there were no clear actinomycete community structure variation trends at the 31-40-cm soil depth. Homology comparison of sequences recovered from the DGGE bands showed that the obtained sequences shared similarities >88 %. Alignment with the known homologous sequences indicated a lack of microorganisms related to soil-borne cotton diseases. Continuous cotton cropping exerted significant influences on the community structure of soil actinomycetes in Xinjiang Autonomous Region, which were largely determined by the soil depth and the period of continuous cotton cropping. The microbial diversity of soil actinomycete communities gradually recovered after 5-year continuous cropping. Thereafter, a new actinomycete community structure that was beneficial for continuous cropping of cotton was formed and stabilized each year.

Effect of karst rocky desertification on soil fungal communities in Southwest China

“…Polymerase chain reaction coupled with denaturing gradient gel electrophoresis (PCR-DGGE) has been widely used to determine the species richness and structure of fungal communities in a variety of environments (Sigler and Turco, 2002;Costa et al, 2006;Yu et al, 2013;Zhang et al, 2013;Dieng et al, 2014). Primarily, this technique is carried out by amplifying specific sequences of interest, such as ribosomal RNA (rRNA) genes (Cahyani et al, 2004;Zhao et al, 2012;Yan et al, 2013;Yoshitake et al, 2013).…”

Bacterial communities of Beijing surface waters as revealed by 454 pyrosequencing of the 16S rRNA gene

“…Traditionally, culture-dependent techniques were used to study bacterial diversities (Alegría et al 2012;Ferrando et al 2012;Vandecandelaere et al 2012), but these methods could not provide accurate representations of the bacterial populations, as most bacteria in the environment are viable, but currently unculturable. Molecular approaches, such as 16S ribosomal RNA (rRNA) clone libraries (Eichler et al 2006;Gihring et al 2012;Zhang et al 2013a) and polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE) (Garrido et al 2014;González-Arenzana et al 2013;Sun et al 2013;Zhang et al 2013b), have greatly increased our comprehension of microbial communities, although these methods poorly represent the rare bacterial groups. In recent years, 454 pyrosequencing has been introduced as a next-generation sequencing method for revealing relatively detailed profiles of microbial communities (Beloshapka et al 2013;Jackson et al 2012).…”

16S rRNA-based Metagenomic Analysis of Endophytic Actinomycetes Diversity from Tinospora crispa L. Miers

“…PCR conditions used were pre-denaturation at 94 °C for 5 min, denaturation at 94 °C for 60 s, annealing at 65 °C for 45 s (a decrease at 0.5 °C per cycle was set for the first 20 cycles, the last decrease was at 55 ºC), elongation at 72 ºC for 120 s for 30 cycles and final elongation at 72 °C for 7 min (Zhang et al 2013).…”

“…Amplification was conducted using T1-thermocycler (Biometra, Goettingen, Germany) with these following optimizations: pre-denaturation at 94 °C for 5 min, denaturation at 94 °C for 60 s, annealing at 55 °C for 45 s, elongation at 72 °C for 60 s for 30 cycles and final elongation at 72 ºC for 5 min (Zhang et al 2013). …”