Performance comparison of different microbial DNA extraction methods on bird feces

Background:As an important player during food digestion,gut microbiota has attracted much attention in diet adaptation studies in birds.Microbiota extracted from feces has been widely used as a proxy for gut microbiota.Although several methods have been developed for microbial DNA extraction,their performances in the bird feces have not been systematacially evaluated yet.Methods:In this study,we applied three DNA extraction methods(Qiagen,MoBio and Bead)to extract DNA from feces of three avian dietary guilds(granivore,omnivore and carnivore),sequenced V4 region of 16S rRNA gene for each extract and evaluated the performances of DNA yield,DNA integrity,microbial composition,cell lysis capacity and alpha diversity for the three methods on each dietary guild.Results:Bead method was the best on the performance of both DNA yield and DNA integrity regardless of dietary guild.In granivore,microbial relative abundance at both species and phylum levels,alpha diversity and cell lysis capacity were comparable among all methods.In omnivore,Qiagen had the best performance on alpha diversity,fol-lowed by Bead and MoBio.There were small variations on microbial relative abundance at both species and phylum levels among different extraction methods.MoBio exhibited the best performance on cell lysis capacity.In carnivore,considerable variations were found on microbial relative abundance at both species and phylum levels.Qiagen had the best performance on alpha diversity,followed by MoBio and Bead.MoBio had the highest cell lysis capacity.Conclusions:DNA yield and integrity have no obvious impact on microbial composition,alpha diversity or cell lysis capacity.The microbiota results(e.g.,microbial composition,cell lysis capacity,alpha diversity)obtained from differ-ent methods are comparable in granivorous avian species but not in omnivorous or carnivorous birds.Either method could be used in granivore microbiota studies.For omnivores and carnivores,we recommend Qiagen method when the research purpose is microbial diversity and MoBio when gram-positive bacteria is the research target.

A modified isooctane-based DNA extraction method from crude oil

Microbes from oil reservoirs shape petroleum composition through processes such as biodegradation or souring.Such processes are considered economically detrimental and might pose health and safety hazards.It is therefore crucial to understand the composition of a reservoir's microbial community and its metabolic capabilities.However,such analyses are hindered by difficulties in extracting DNA from such complex fluids as crude oil.Here,we present a novel DNA extraction method from oils with a wide American Petroleum Institute(API)gravity(density)range.We investigated the ability to extract cells from oils with different solvents and surfactants,the latter both nonionic and ionic.Furthermore,we evaluated three DNA extraction methods.Overall,the best DNA yields and the highest number of 16S rRNA reads were achieved with isooctane as a solvent,followed by an ionic surfactant treatment using sodium dodecyl sulfate and DNA extraction using the PowerSoil Pro Kit(Qiagen).The final method was then applied to various oils from oil reservoirs collected in aseptic conditions.Despite the expected low cell density of 10^(1)–10^(3)cells/ml,the new method yielded reliable results,with average 16S rRNA sequencing reads in the order of 41431(±8860)per sample.Thermophilic,halophilic,and anaerobic taxa,which are most likely to be indigenous to the oil reservoir,were found in all samples.API gravity and DNA yield,despite the sufficient DNA obtained,did not show a correlation.