Identification of anaerobic ammonium-oxidizing bacteria in environmental samples by T-RFLP

Abstract:

To rapidly identify anaerobic ammonium-oxidizing (anammox) bacteria in environmental samples, a cloning-independent method based on Terminal Restriction Fragment Length Polymorphism (T-RFLP) was proposed. Based on SILVA (R108) SSU Ref database containing 400 16S rRNA gene sequences of anammox bacteria, defined lengths of terminal restriction fragments (T-RFs) were predicted by virtual PCR and restriction enzyme digestion using suitable PCR primers (Amx368f-Amx820r) and restriction enzyme (MspI ＆ RsaI). Then the genus of anammox bacteria was identified by comparative analysis of T-RFs. The repeatability and sensibility analysis results show that the anammox bacteria specific T-RFLP analysis is a reliable tool to rapidly assess the complexity of anammox bacteria in environmental samples.