CN109706235A - A kind of the detection and analysis method and its system of intestinal microflora - Google Patents
A kind of the detection and analysis method and its system of intestinal microflora Download PDFInfo
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Abstract
The present invention relates to field of biological detection, in particular to the detection and analysis method and its system of a kind of intestinal microflora.The detection and analysis method of the intestinal microflora includes acquisition fecal sample；Microbial flora genome DNA in fecal sample is extracted, and PCR amplification is carried out to the total DNA；Bioinformatic analysis is carried out based on sequencing data, obtains analysis result；And analysis result is compared with the Intestinal microbial bacteria group database, obtain the intestinal microflora state of subject；Intestinal microflora state and its subject's basal conditions correspondence based on subject provide adjustable index and monitor control index.The testing result that detection and analysis method based on above-mentioned intestinal microflora obtains provides corresponding enteral nutrition structural adjustment suggestion, increases health index in subject.
Description
[technical field]
The present invention relates to microbial flora detection field, in particular to a kind of detection and analysis side of intestinal microflora
Method and its system.
[background technique]
Enteric microorganism has important regulating and controlling effect to organism metabolism, energy flow, signal transduction etc., and right
The diet of animal, nutrition, immune, nerve modulation and chronic disease have important influence.Therefore, to enteric microorganism
Carrying out thoroughgoing and painstaking research is to understand various chronic diseases and the immune important side with pathogenic microorganism disease genesis mechanism
Formula method.
In recent years, with the continuous development of technique of gene detection and maturation, which is probing into and is identifying microbial flora
In be also widely used, this method breaches the limitation that microorganism is separately cultured, and can analyze nature comprehensively
Microbe groups simultaneously sufficiently excavate genetic resources.But existing microbial flora DNA is extracted and accuracy in detection is lower, no
It can meet the genesis mechanism that various chronic diseases are furtherd investigate based on intestinal microflora；High-flux sequence is as a kind of new
The technique of gene detection of type, measuring speed is fast, precision is high, can meet and further investigate various chronic diseases based on intestinal microflora
The genesis mechanism of disease, but the analysis of the data of high-flux sequence is difficult, testing cost is higher, can be only applied to scientific research at present
Stage.Therefore, how to carry out intestinal microbial DNA detection to Cheap highly effective and will test result applied to human body diseases analysis change
It must be even more important.
For this purpose, the present invention provides the detection and analysis methods and its system of a kind of intestinal microflora.
[summary of the invention]
In order to solve technical problem present in existing microbial DNA detection and analysis application, the present invention provides a kind of intestines
The detection and analysis method and its system of road microbial flora.
The present invention to solve above-mentioned technical problem the following technical schemes are provided: a kind of detection of intestinal microflora and point
Analysis method, the detection and analysis method of the intestinal microflora is the following steps are included: step S1: acquisition fecal sample；Step
Rapid S2: microbial flora genome DNA in fecal sample is extracted, and PCR is carried out to the total DNA (DNA)
(polymerase chain reaction) amplification；Step S3: the total DNA after amplification is carried out to build library and sequencing, to obtain raw sequencing data；
Step S4: raw sequencing data is subjected to data Quality Control processing, obtains sequencing data；Step S5: it is given birth to based on sequencing data
Object bioinformatics analysis obtains analysis result；And step S6: an Intestinal microbial bacteria group database is provided, result will be analyzed
It is compared with the Intestinal microbial bacteria group database, obtains the intestinal microflora state of subject；Based on by
Intestinal microflora state and its subject's basal conditions correspondence of inspection person provides adjustable index and monitor control index.
Preferably, above-mentioned steps S1 further comprises following steps: step S11: using the appropriate excrement sample of sterile swab picking
This；Step S12: by the sterile swab for speckling with excrement, stirring makes to save liquid discoloration in containing the test tube for saving liquid；And step
S13: the discoloration containing fecal sample is saved into liquid and is stored at room temperature.
Preferably, the preservation liquid includes: thio hydrohalogenic acid salt, Organic Alcohol, complexing agent, phosphate buffer, tween
(Tween), seralbumin and benzalkonium bromide；The thio hydrohalogenic acid salt concentration is 0.01M-5M, and the Organic Alcohol accounts for preservation liquid
The 30%-60% of total weight；The concentration of complexing agent is 0.001M-0.35M；The phosphate buffer accounts for the 5%- for saving liquid weight
15%；The tween accounts for the 0.05%-1% for saving liquid weight, and sero-abluminous concentration is 2.5mM- in the preservation liquid
100mM；Benzalkonium bromide accounts for the 0.01%-5% for saving liquid total weight.Preferably, above-mentioned steps S2 further comprises following steps:
Step S21: CTAB (Cetyltrimethylammonium Bromide, cetyl trimethylammonium bromide) method or SDS are utilized
(Sodium Dodecyl Sulfate, lauryl sodium sulfate) method extracts microbial flora total DNA in fecal sample；And step
S22: it is expanded to obtain using the area the V3-V4 segment of the 16S rRNA gene in primer pair microbial flora genome DNA
Total DNA after amplification.
Preferably, above-mentioned steps S3 further comprises following steps: step S31: the DNA after amplification is purified to obtain
Total DNA after amplification；Step S32: based on the total DNA building mixing library after amplification；And step S33: pass through high-flux sequence
Platform is sequenced, and raw sequencing data is obtained.
Preferably, above-mentioned steps S4 further comprises: step S41: the interference data in removal raw sequencing data；Step
Rapid S42: sequencing fragment splicing (reads splicing) is carried out to the raw sequencing data for going interference data to obtain and obtains original tag
(Raw Tags)；Step S43: high quality is obtained by label (Tags) interception and label (Tags) length filtration, label is sequenced
(Clean Tags)；And step S44: it carries out being fitted into (Chimeric Tags) gymnastics work by the way that label is sequenced to high quality, obtain
To the effective label (Effective Tags) that can carry out subsequent analysis.
Preferably, above-mentioned steps S5 includes: step S51: carrying out OTUs (Operational based on effective label
Taxonomic Units, the taxonomical unit of operation) clustering, and obtain analysis result；Step S52: based on analysis as a result,
Obtain group composition of the intestinal microflora on boundary, door, guiding principle, mesh, section, each level of category kind；And step S53: by group's group
It is associated analysis at the metabolic pathway with software prediction, obtains the influence of the pairs of metabolic pathway of group's group.
Preferably, above-mentioned steps S51 includes: step S511: each effective label is picked out, with RDP (The
Ribosomal Database Project) database is compared, it is clustered with effective tag compliance greater than 97%
And obtain sequence taxon；Step S512: the representative series of sequence taxon are picked out；And step S513: in conjunction with generation
Table sequence carries out the division of species taxonomy information using presetting database.
Preferably, above-mentioned steps S52 further comprises: step S521: right using rank sum test method (Rank-Sum Test)
Species between different grouping are for statistical analysis, to find out the species for forming on group and dividing and generating significant difference and influence；And
Step S522: being analyzed using LEfSe (LDA Effect Size), in all levels of species taxonomy, passes through linear discriminant point
It analyses (LDA:Linear Discriminant Analysis), estimates the size that each species abundance influences differential effect, look for
The species for generating significant difference and influencing are divided on sample out.
The present invention to solve above-mentioned technical problem the following technical schemes are provided: a kind of detection of intestinal microflora and point
Analysis system, it is characterised in that: include: sample collection unit: it is configured to acquisition fecal sample；PCR amplification unit: it is configured to
Microbial flora genome DNA in fecal sample is extracted, and PCR amplification is carried out to the total DNA；Data capture unit: match
It sets for carrying out building library and sequencing to the total DNA after amplification, to obtain raw sequencing data；And raw sequencing data is counted
It is handled according to Quality Control, obtains sequencing data；Bioinformatic analysis unit: it is configured to carry out bioinformatics based on sequencing data
Analysis obtains analysis result；And information comparison unit: being configured to provide for an Intestinal microbial bacteria group database, will divide
Analysis result is compared with the Intestinal microbial bacteria group database, obtains the intestinal microflora state of subject；
Intestinal microflora state and its subject's basal conditions correspondence based on subject provide adjustable index and monitor control index.
Compared with prior art, the detection and analysis method of a kind of intestinal microflora provided by the invention, for taking
The cooperation between links and each link such as sample, Jian Ku, sequencing and data analysis, makes intestinal microflora detection come into big all living creatures
It is living, it escorts for public health, this will be a promising length and breadth of land market.This is also in contemporary internet and big
Data, a kind of digitization under background and intelligentized carry out evaluating method to subject's health based on enteric microorganism information
It feels free to try, there is subversive innovation and promote meaning to traditional biological medical industry.
Firstly, a kind of detection and analysis method of intestinal microflora provided by the invention, based on the micro of subject
Fecal sample is detected, and be can effectively avoid existing embarrassment during business sampling operation, is kept detection more humanized.
Secondly, a kind of detection and analysis method of intestinal microflora provided by the invention, using PCR method in excrement
Microorganism total DNA is expanded, and gene cloning is bypassed, and the mode that hybridization or microorganism are separately cultured directly obtains a large amount of expansions
Increase sub-sequence information, sensitivity is higher, the species relative abundance of each microorganism in intestinal flora can be accurately detected, for further
Bioinformatic analysis is carried out, the diversity and abundance of subject enteric microorganism are accurately reflected.
Again, the detection and analysis method of a kind of intestinal microflora provided by the invention utilizes international advanced sequencing
Platform carries out 16S rRNA sequencing, and sequencing speed is fast, the sequencing time is short, and sequencing accuracy is high.After the sequencing is completed using life
The abundance and diversity information for believing big data analysis processing technique decryption intestinal microflora, gradually accumulate different crowd enteron aisle
The data of microbial flora, building belong to the enteric microorganism database of Chinese population, make the decline of sequencing cost, analyze result
Accuracy so that genetic test is gradually come into popular life.
Finally, by the personal information of input subject, by the enteron aisle of its enteric microorganism test data and Chinese population
Microbiological data library is matched, using the abundance and multiplicity of raw letter big data analysis processing technique decryption intestinal microflora
Property information, the composition of intestinal microflora is analyzed, a large amount of gene carries out metabolism group in binding sequence
(Metabonomics) research；It is found and body phase by metabolism with organism and related immunological diseases binding analysis
The mechanism of interaction, and then be adjusted according to human body basal conditions, increase human health index, analyzes current intestines for subject
Road health status simultaneously provides specific aim suggestion and is adjusted.
A kind of detection and analysis method of intestinal microflora provided by the invention saves liquid to excrement sample using excrement
Product are saved, it is ensured that can effectively be kept the diversity of intestinal microflora structure in 15 days at room temperature, be reduced intestines
Road microbial flora saves requirement condition and cost realizes the micro- life of enteron aisle in excrement under the specific conditions such as outgoing sampling, sample transport
Effective preservation of object flora.
A kind of detection and analysis method of intestinal microflora provided by the invention makes intestinal microflora detection more
Add it is quick and easy, efficient, can be formed high-flux sequence analysis in microbial function database.
The detection and analysis system of intestinal microflora provided by the invention, the detection of the intestinal microflora and
Analysis system has beneficial effect identical with the detection and analysis method of above-mentioned intestinal microflora, and details are not described herein.
[Detailed description of the invention]
Fig. 1 is the step process of the detection and analysis method of the intestinal microflora provided in first embodiment of the invention
Schematic diagram；
Fig. 2 is the feces collection of the detection and analysis method of the intestinal microflora provided in first embodiment of the invention
Steps flow chart schematic diagram；
Fig. 3 is the PCR of the DNA of the detection and analysis method of the intestinal microflora provided in first embodiment of the invention
The step flow diagram of amplification procedure；
After Fig. 4 is the amplification of detection and analysis method of the intestinal microflora provided in first embodiment of the invention
The step flow diagram of the sequencing procedure of DNA；
Fig. 5 is the data Quality Control of the detection and analysis method of the intestinal microflora provided in first embodiment of the invention
The step flow diagram of process；
Fig. 6 is the biological information of the detection and analysis method of the intestinal microflora provided in first embodiment of the invention
The step flow diagram of credit analysis；
Fig. 7 is the OTUs cluster of the detection and analysis method of the intestinal microflora provided in first embodiment of the invention
The step flow diagram of analysis；
Fig. 8 is micro- life of the detection and analysis method of the offer intestinal microflora provided in first embodiment of the invention
The step flow diagram of object composition analysis process；
Fig. 9 is the frame of the detection and analysis system of the offer intestinal microflora provided in second embodiment of the invention
Structural schematic diagram.
Attached drawing mark:
20, intestinal microflora detection and analysis system；21, sample collection unit；22, PCR amplification unit；23, number
According to acquiring unit；24, bioinformatic analysis unit；25, information comparison unit.
[specific embodiment]
In order to make the purpose of the present invention, technical solution and advantage are more clearly understood, below in conjunction with attached drawing and embodiment,
Invention is further elaborated.It should be appreciated that described herein, specific examples are only used to explain the present invention, not
For limiting the present invention.
The first embodiment of the present invention provides a kind of detection and analysis method of intestinal microflora, suitable for by grinding
Study carefully the microbial flora composition in subject's enteron aisle, exists between the health of subject to probe into intestinal microflora and form
Substantial connection.
Microorganism in enteron aisle reaches 100,000,000,000,000 order of magnitude, and the interaction between microorganism and host maintains enteron aisle
The stabilization of Tiny ecosystem structure.
Enteric microorganism has important regulating and controlling effect to host substance metabolism, energy flow, signal transduction etc., and to dynamic
The diet of object, nutrition, immune, nerve modulation and chronic disease have important influence.Intestinal flora in a healthy and balanced way
Structure can promote the growth and development of organism；Unbalance intestinal flora can then get muddled to the adjusting of organism, and then produce
It is raw a series of to unbalance related chronic or endocrine disturbance the complication of intestinal microecology；Such as obesity, cardiovascular disease, cancer
Disease, inflammatory bowel disease, diabetes, allergy, metabolic syndrome, irritable bowel syndrome etc..Therefore, by being carried out to enteric microorganism
It furthers investigate, further predicts human body overview situation, and make adjustment appropriate to become particularly important.
Possible bacterial species include: Lactobacillus, bifidobacterium adolescentis, animal bifid bar in human body intestinal canal flora
Bacterium, bifidobacterium bifidum, bifidobacterium breve, bifidobacterium longum, Pediococcus acidilactici, acinetobacter, actinomyces, Campylobacter spp
Belong to, bite carbon dioxide cellulose Pseudomonas, Crow promise Pseudomonas, enterococcus spp, Fusobacterium, Leptothrix, Mycobacterium, digestion
Streptococcus, Providence Pseudomonas, clostridium difficile etc..
Referring to Fig. 1, the first embodiment of the present invention provides the detection and analysis method S10 of an intestinal microflora,
It is further described, is specifically comprised the following steps: with the measurement to microbial flora in enteron aisle
Step S1: acquisition fecal sample；
Step S2: microbial flora genome DNA in fecal sample is extracted, and PCR amplification is carried out to the total DNA；
Step S3: the total DNA after amplification is carried out to build library and sequencing, to obtain raw sequencing data；
Step S4: raw sequencing data is subjected to data Quality Control processing, obtains sequencing data；
Step S5: carrying out bioinformatic analysis based on sequencing data, obtains analysis result；And
Step S6: providing an Intestinal microbial bacteria group database, by analysis result and the Intestinal microorganism
Flora database is compared, and obtains the intestinal microflora state of subject；Intestinal microflora based on subject
State and its subject's basal conditions correspondence provide adjustable index and monitor control index.
Contain a large amount of enteric microorganism in subject's excrement, therefore, the microbial bacteria in detection animal wastes can be passed through
Group's composition further finds out the microbial flora information in subject's enteron aisle, and then understands the drink of enteric microorganism and subject
The relationship of the biological characteristics such as food, nutrition, immune, nerve modulation and the generation of chronic disease.It is carried out based on micro fecal sample
Detection can effectively avoid existing embarrassment during business sampling operation.
As shown in Fig. 2, acquiring fecal sample in step S1, further comprising following steps:
Step S11: the appropriate fecal sample of sterile swab picking is used；
Step S12: by the sterile swab for speckling with excrement, stirring makes to save liquid discoloration in containing the test tube for saving liquid；And
Step S13: the discoloration containing fecal sample is saved into liquid and is stored at room temperature.
When carrying out feces collection, the quality of required excrement is 0.02g-0.2g；For the accuracy for increasing measurement, each excrement
Sample collection 3 times, use when to detect.
Specifically, the cotton swab for speckling with excrement stirs 1 minute or so in the test tube for filling clarification preservation liquid, clear preservation
Liquid becomes yellowish-brown, i.e. collected sample, can meet testing requirements.
The preservation liquid includes: thio hydrohalogenic acid salt, Organic Alcohol, complexing agent, phosphate buffer, tween, seralbumin, benzene
Prick bromine ammonium, potassium permanganate, sterile glycerol buffered saline, any one or more combinations in sterile distilled water.The thio hydrohalogenic acid salt
Concentration is 0.01M-5M, and the Organic Alcohol accounts for the 30%-60% for saving liquid total weight；The concentration of complexing agent is 0.001M-
0.35M；The phosphate buffer accounts for the 5%-15% for saving liquid weight；The tween accounts for the 0.05%-1% for saving liquid weight,
Sero-abluminous concentration is 2.5mM-100mM in the preservation liquid；Benzalkonium bromide accounts for the 0.01%-5% for saving liquid total weight；
Potassium permanganate is 0.5mM-10mM；Sterile glycerol buffered saline accounts for the 1%-15% for saving liquid total weight.
The preservation liquid of above-mentioned configuration is stored at room temperature the microbial flora in fecal sample, it is ensured that can be in room temperature condition
Effectively keep the diversity of intestinal microflora structure in lower 15 days, reduce intestinal microflora save requirement condition and at
Under the specific conditions such as the outgoing sampling of this realization, sample transport in excrement intestinal microflora effective preservation.
As shown in figure 3, step S2: flora genomic DNA in fecal sample is extracted, and PCR amplification is carried out to the DNA,
Further comprise following steps:
Step S21: microbial flora genome DNA in fecal sample is extracted using CTAB method or SDS method；And
Step S22: the area the V3-V4 segment of the 16S rRNA gene in primer pair microbial flora genome DNA is utilized
Total DNA after being expanded.
Specifically, 16S rRNA is located on the ribosomes of prokaryotes such as bacterium, shares 10 conservative regions
(Conserved Regions) and 9 hypervariable regions (Hypervariable Regions)；Wherein conserved region is between kind
Difference is little, and hypervariable region have specificity and it is variant according to affiliation, therefore can be used as bacterial population identify
Index.
16S rRNA sequencing generally selects hypervariable region progress, generally selects at this stage to Bacteria Identification in enteric microorganism
V3-V4 is carried out, and the present embodiment uses the (place of production: the U.S. Illumina；Model: Miseq 2500) to the region 16Sr DNA V3-V4
It is sequenced and carries out subsequent analysis.
Wherein, microbial flora genome DNA in fecal sample is extracted using CTAB method, further comprises grasping as follows
Make:
It configures CTAB extracting solution and extracts microbial flora genome DNA, the CTAB extracting solution includes: NaCl (chlorination
Sodium), CTAB, TrisCl (three (methylol) aminomethanes), EDTA (ethylenediamine tetra-acetic acid) and mercaptoethanol.
Wherein, it is 50mM- that the mass percent of NaCl concentration 1M-2M, CTAB, which is the concentration of 1%-5%, TrisCl,
The concentration of 200mM, EDTA are 5mM-40mM, the mass percent of mercaptoethanol is 0.1%-0.5%.
Configure phenol: chloroform: isoamyl alcohol=(10-30): (10-30): the mixed liquor of (0.5-5) has extracted albuminate
Matter impurity.
The extraction process specifically:
1) CTAB extracting solution described in heating water bath is to 60 DEG C -65 DEG C；
2) the preservation liquid containing fecal sample is added in CTAB extracting solution and forms mixed liquor, the guarantor containing fecal sample
Liquid storage: extraction liquid proportional is 1-3；
3) mixed liquor is transferred in centrifuge tube, 60 DEG C -65 DEG C of water-baths -3 hours 0.5 hour；
4) isometric phenol/chloroform/isoamyl alcohol is added, overturns mixing, the protein impurities contained in denaturated extract；
6) 10000rpm-20000rpm is centrifuged 10min-30min under room temperature (25 DEG C), removes supernatant.
7) excrement total DNA sample extraction complete after, mark and be placed on -20 DEG C it is spare.
8) the total DNA sample extracted is detected using 1% Ago-Gel, judges the concentration and matter of extracting sample
Amount.
The DNA extracted by above-mentioned CTAB method, accuracy and integrity degree are higher, meet subsequent sequencing demands.
Then, it is obtained using extraction and detects qualified total DNA as template, using HIFI (high fidelity enzyme) to the 16S of selection
The region rRNA V3-V4 is expanded.
In some specific embodiments, primer sequence used in first embodiment of the invention is 27F and 1492R.
The 27F are as follows: 5'-AGAGTTTGATCMTGGCTCAG-3'；1492R are as follows: 5'-
TACGGYTACCTTGTTACGACTT-3'。
In the present invention, by PCR amplification process, compared to gene cloning is utilized, hybridization or microorganism are separately cultured
Mode, massive amplification sub-sequence information can be directly obtained, and microbial flora is probed into according to varying environment and condition
Otherness.
In the other embodiments of first embodiment of the invention, microbial flora base in fecal sample is extracted using SDS method
Because of a group total DNA.The DNA extracted by the SDS method, accuracy and integrity degree are higher, meet subsequent sequencing demands.
As shown in figure 4, step S3: carrying out building library and sequencing to the total DNA after amplification, to obtain raw sequencing data；Into
One step includes:
Step S31: the DNA after amplification is purified to the total DNA after being expanded；
Step S32: based on the total DNA building mixing library after amplification；And
Step S33: it is sequenced by high-flux sequence platform, obtains raw sequencing data.
Specifically, include: to the depositing process of product after amplification
Step S311: the free primer after removing amplification 16S rRNA V3-V4 regional gene using magnetic bead；And
Step S312: it is cleaned again before last quantitative to building library sample, concrete operation step and step S311 class
Seemingly.
Specific cleaning step are as follows:
1) amplified sample at room temperature (25 DEG C), is centrifuged 1min-10min；Revolving speed is 10000rpm-20000rpm；
2) whirlpool shakes magnetic bead 10s-50s, it is ensured that all magnetic beads are suspended；
3) magnetic bead of 10 μ L-30 μ L is added into the PCR pipe of each amplification；
4) sample is drawn to piping and druming 5-20 times repeatedly with liquid-transfering gun；
5) 1min-10min is stood at room temperature；
6) PCR pipe is placed in magnetic sheet 1min-10min until liquid becomes to clarify；
7) sample supernatant is absorbed using liquid-transfering gun in magnetic sheet；
8) in magnetic sheet, magnetic bead is cleaned with the ethyl alcohol of 50%-80%；
A. it is added in each sample with the ethyl alcohol that liquid-transfering gun draws 200 μ L；
B. 30s is stood in magnetic sheet；
C. it carefully draws supernatant and discards.
9) in magnetic sheet, second of cleaning operation is carried out using the ethyl alcohol of 50%-80%:
A. the ethyl alcohol of 200 μ L is drawn into each sample using liquid-transfering gun；
B. 30s is stood in magnetic field；
C. it carefully draws supernatant and discards；
D. remaining ethyl alcohol is all removed as far as possible using the rifle and pipette tips that range is 20 μ L.
10) in magnetic field, centrifuge tube is uncapped and stands 1min-20min, make ethanol evaporation and drying in pipe；
11) centrifuge tube is left into magnetic field, and is added 5mM-10mM Tris (tromethamine) pH8.5's into each pipe
Buffer 30 μ L-80 μ L；
12) it using liquid-transfering gun absorption gently and blows and beats 5-20 times, all magnetic beads is resuspended；
13) 1min-10min is stood at room temperature；
14) PCR pipe is placed in magnetic field, stands 1min-10min；Supernatant is set to become to clarify completely；
15) 50 μ L of absorption supernatant gently, is placed in new centrifuge tube, marks spare.
Cleaned DNA is subjected to mixing library construction and upper machine is sequenced, but sequencing result is more accurate.
Each sample in library will be mixed and carry out quantitative work, make various kinds Ben Wenku in the same concentration level, and by each sample
It is mixed to form a mixing library.And obtained library is denaturalized, is diluted, the final Miseq using Illumina is sequenced
Platform is sequenced, and raw sequencing data (Raw PE) is obtained.
As shown in figure 5, above-mentioned steps S4: raw sequencing data being carried out data Quality Control processing, sequencing data is obtained, into one
Step includes:
Step S41: the interference data in removal raw sequencing data；
Step S42: sequencing fragment is carried out to the raw sequencing data for going interference data to obtain and splices to obtain original tag；
Step S43: high quality is obtained by filtration by label interception and tag length, label is sequenced；And
Step S44: it is operated by carrying out chimera to high quality sequencing label, obtains that having for subsequent analysis can be carried out
Criterion label.
Specifically, it can be used in the data of subsequent analysis in order to obtain, first removal interference data.The interference data packet
It includes: the impurity data generated when primer or other amplifications.
Raw sequencing data carries out sequencing fragment splicing, and removal DNA bar code (Barcode) and primer sequence obtain original
Label；The DNA bar code refer to can be represented in organism the species, standard, it is having enough variations, easily amplification and phase
To shorter DNA fragmentation.DNA bar code has become the important tool of ecological study, is applied not only to species identification, while
Biologist is helped to further appreciate that the interaction occurred in the ecosystem.Finding the one of a kind of unknown species or species
When part, researcher just describes its DNA bar code, is then compared with other bar codes in international data center.If
Match with one of them, researcher can confirm the classification of this species.
Quality Control processing is carried out to original tag according to Qiime analysis platform, specifically, carries out label interception and label respectively
High quality sequencing label is obtained after the operation such as length filtration；High quality is sequenced label and obtains after UCHIME platform removes chimera
To the effective label that can carry out subsequent analysis.
Wherein, the Qiime analysis platform is the 16S process analysis platform for gathering numerous softwares and script.
The UCHIME platform is a kind of detection instrument of chimera.
The chimera is to extend the stage during extension increasing sequence X in sequence, only produce the extension of part X sequence
Stage just finishes, and in the PCR reaction of next round, this partial sequence then extends as the primer of sequence Y, and amplification will shape
At the chimera sequence of X and Y.
As shown in fig. 6, step S5: bioinformatic analysis is carried out to sequencing data, to be analyzed as a result, further wrapping
It includes:
Step S51: carrying out OTUs clustering based on effective label, and obtains analysis result；
Step S52: the group on kind of each level is belonged in Jie Men detailed outline section as a result, obtaining intestinal microflora based on analysis
Composition；And
Step S53: group is formed, and is associated analysis with the metabolic pathway of software prediction, obtains group's group pairs of generation
Thank to the influence of access.
OTUs clustering is carried out by using Qiime analysis platform in step s 51；In step S53 by using
The metabolic pathway of PICRUSt software prediction is associated analysis.
The PICRUSt software is based on nearly edge species after the information and comparison presetting database for having surveyed bacterial 16 S rRNA
OTU information infers the gene function spectrum of their common ancestor, while to the gene function for not surveying species other in presetting database
Power spectrum is inferred, the Gene correlation spectrum of Archimycetes and the full pedigree in bacterium domain is constructed；Finally, the flora group that sequencing is obtained
At being mapped in database, then bacterial metabolism function is predicted.
As shown in fig. 7, above-mentioned steps S51 further comprises:
Step S511: picking out each effective label, be compared with RDP database, to be greater than 97% effective label
Consistency is clustered and obtains sequence taxon；
Step S512: the representative series of sequence taxon are picked out；And
Step S513: in conjunction with representative series using presetting database into.The presetting database is Greengenes number
According to combinations one or two kinds of in library or SILVA database.
Specifically, the effective label obtained after above-mentioned Quality Control is carried out with 97% sequence identity using UPARSE software
It clusters and obtains sequence taxon；The sequence that the sequence label and sequence label frequency that can not be wherein clustered are 1 will be given up
It abandons；Occurring that frequency is highest in sequence label will be by the representative sequence as sequence taxon.
Wherein, the UPARSE software is ultrafast sequence analysis software, multi-field in sequence alignment, cluster, operation etc.
It is widely applied.It is most popular in the OTUs cluster of amplicon analysis field, it has been integrated with whole amplicon analysis process at present.
Using Mothur software and SILVA database carry out OTUs clustering species annotation and successively obtain boundary,
Door, guiding principle, mesh, section, the group belonged on kind of each level form；Base can be constructed according to the case where dividing OTUs clustering in sample
In the phylogenetic tree of OTUs clustering.For the ease of carrying out subsequent Alpha diversity analysis (Alpha Diversity)
With Beta diversity analysis (Beta Diversity) need to carry out sample data uniform processing and with data volume it is least
Sample is standard.
As shown in figure 8, above-mentioned steps S52, which is based on analysis, belongs to kind in Jie Men detailed outline section as a result, obtaining intestinal microflora
Group's composition on each level further comprises:
Step S521: it is for statistical analysis between the species different grouping using rank sum test method, to find out to group's group
The species influenced at generation significant difference is divided；And
Step S522: being analyzed using LEfSe, and in all levels of species taxonomy, by linear discriminant analysis, estimation is every
The size that the abundance of a species influences differential effect finds out the species for dividing on sample and generating significant difference and influencing.Wherein,
LEfSe analysis is a kind of for finding and explaining high number of latitude according to the analysis work of biological identification (gene, access and taxon etc.)
Tool, can carry out the comparison of two or more components, emphasize statistical significance and biological relevance, can seek to have between group and group
There is the biological identification of statistical difference.
It is raising scientific experiment as a result, systematic error inspection must be carried out to experiment the data obtained.Rank sum test is one
The kind effectively simple method of inspection, uses sum of ranks to carry out hypothesis testing as statistic, and whether principle is to be based on depositing between two groups of data
In significant difference；Rank sum test can also be used to prove the reliability of new test method.
By above-mentioned statistical analysis, the significant species of its abundance difference can be found out, and analyze the species in different samples
Or the enrichment condition in different groups；The conspicuousness of group difference and group difference can be judged, finally by statistical analysis simultaneously
Judge whether group difference reaches the level of conspicuousness.
Specifically, it is carried out based on OTUs cluster annotation results, the analysis of Alpha diversity analysis, i.e. sample complex can
For the analysis to one-time detection fecal sample；Between Beta diversity analysis, i.e. multisample comparative analysis, that is, microbe colony group
Variance analysis (Statistics).
The relative abundance size of each microbial population in test sample is obtained, and then carries out species difference comparative analysis,
And it seeks influence of the microbial flora otherness to subject's physical condition and provides recommendation on improvement.
Subject's physical condition includes: human body constitution, immunity level, health risk and nutrient absorption level etc..
Beta diversity analysis: comparison and building disease risks mould for same organism different time sections detection data
Type.
By the way that microbial flora information contained in single measurement sample can be obtained after above-mentioned analysis.The microbial bacteria
Group's information includes microbial flora type, the relative abundance of each microbial flora, microbial flora growth and breeding level etc..
Intestinal microbial bacteria group database is provided in above-mentioned step S6, by analysis result and the Intestinal
Microbial bacteria group database is compared, and obtains the intestinal microflora state of subject；The micro- life of enteron aisle based on subject
Object flora state and its subject's basal conditions correspondence provide adjustable index and monitor control index.
Data in the Intestinal microbial bacteria group database are that first embodiment provides an enteron aisle through the invention
Step S1, step S2, step S3, step S4 and step S5 in the detection and analysis method S10 of microbial flora are obtained；Crowd
Data in intestinal microflora database can be updated at any time.
The Intestinal microbial bacteria group database cover the different crowd age, gender, height, weight, eating habit,
The intestinal microfloras information such as constitution, territorial scope.
Due to, be in the infants being just born it is sterile, a period of time of baby after birth, microorganism starts slowly
It is colonized in inside enteron aisle and gradually forms a stable intestinal microflora.Therefore, crowd of the age less than 3 years old carries out year
When age divides, it can further be accurate to month.
Intestinal microflora state and its subject's basal conditions based on subject, can be rapidly from database
Obtain corresponding adjustable index and monitor control index.
Specifically, in the database can using the age of subject, gender, height and location as index,
These four conditions are considered as corresponding four indexs A, B, C and D, and the corresponding weight of four different indexs is different, for example, referring to
The weight for marking A is 30%, and the weight of index B is 40%, and the weight of index C is 10%, and the weight of index D is 20%, different
Index can obtain corresponding intestinal microflora state from database.Subject inputs personal essential information (name, year
Age, height, gender and region), database is successively based on gender, age, location and the automatic search of height progress and looks into
Find the intestinal microflora information with the health of subject's information matches.
By the way that the intestinal microflora information of subject and the enteron aisle in Intestinal microbial bacteria group database is micro-
Biological flora information is compared, and obtains the intestinal microflora state of subject, the enteric microorganism bacterium based on subject
Group's state and its subject's basal conditions correspondence provide adjustable index and monitor control index.
Specifically, the adjustable index includes: prebiotic bacterial content, diversity, trophic level；The monitor control index packet
It includes: intestinal disorder coefficient, type-2 diabetes mellitus risk, enteritis risk, immunity level, anxiety level etc..
Adjustable index: the index that can be improved by probiotic supplemented or diet.
Monitor control index: the health risk situation of subject instantly is represented, suggesting effect is played.
Intestinal microflora status information is presented to subject by report form, and the report is interior with evaluation index
Form is presented: including physique assessment, health risk and trophic level.
The physique assessment further comprises: intestinal disorder coefficient, probiotics quantity and bacterial diversity etc..
The health risk further comprises: intestinal inflammatory risk, type-2 diabetes mellitus risk, immunity level and anxiety wind
Danger etc..
The trophic level include: essential amino acid synthesis capability, mineral absorption ability, vitamin A synthesis capability and
Protein, fat, carbon hydrate metabolic capability etc..
The report form includes any one or two kinds of combinations in electronic report or papery report form.
On the basis of giving result, for adjustable index, then personalized guiding opinion is provided.
A kind of detection and analysis method of intestinal microflora provided by the invention, for sample, build library, sequencing sum number
According to the cooperation between links and each link such as analysis, intestinal microflora detection is made to come into popular life, is public
Health escorts, this will be a promising length and breadth of land market.This is also one under contemporary internet and big data, background
Kind of digitization and it is intelligentized evaluating method carried out to subject's health based on enteric microorganism information feel free to try, to tradition
Biologic medical industry has subversive innovation and promote meaning.
The second embodiment of the present invention provides a kind of intestinal microflora detection and analysis system 20, the micro- life of enteron aisle
Object bacteria detection and analysis system 20 include hardware or software, can be used for implementing the intestinal microflora detection and analysis side
Method
As shown in figure 9, intestinal microflora detection and analysis system 20 includes:
Sample collection unit 21: it is configured to acquisition fecal sample.
The sampling unit 21 includes any one or two kinds of groups in artificial sample acquisition module or machine sample collection module
It closes.The artificial sample acquisition module includes that sample collection person respectively acquires equipment.
PCR amplification unit 22: it is configured to extract microbial flora genome DNA in fecal sample, and to described total
DNA carries out PCR amplification.
In some specific embodiments, PCR amplification unit 22 includes an at least PCR amplification device and its matched
DNA extract equipment.
Data capture unit 23: it is configured to that the total DNA after amplification is carried out to build library and sequencing, to obtain primitive sequencer number
According to；And raw sequencing data is subjected to data Quality Control processing, obtain sequencing data.
In some specific embodiments, data capture unit 23 includes an at least sequencing equipment and its matched all kinds of
Software.
Bioinformatic analysis unit 24: it is configured to carry out bioinformatic analysis based on sequencing data, be analyzed
As a result；And
In some specific embodiments, bioinformatic analysis unit 24 includes an at least analytical equipment and its mating
All kinds of softwares.
Information comparison unit 25: being configured to provide for an Intestinal microbial bacteria group database, by analysis result and institute
It states Intestinal microbial bacteria group database to be compared, obtains the intestinal microflora state of subject；Based on subject
Intestinal microflora state and its subject's basal conditions correspondence provide adjustable index and monitor control index.The information ratio
Include: to unit 25
Contrast module: being configured to provide for an Intestinal microbial bacteria group database, by analysis result and the crowd
Intestinal microflora database is compared, and obtains the intestinal microflora state of subject；And
Output module: intestinal microflora state and its subject's basal conditions based on subject are configured to and is corresponded to
Provide adjustable index and monitor control index.
The detection and analysis system of intestinal microflora provided by the invention, the detection with above-mentioned intestinal microflora
Beneficial effect identical with analysis method.
Compared with prior art, the detection and analysis method of a kind of intestinal microflora provided by the invention, for taking
The cooperation between links and each link such as sample, Jian Ku, sequencing and data analysis, makes intestinal microflora detection come into big all living creatures
It is living, it escorts for public health, this will be a promising length and breadth of land market.This is also in contemporary internet and big
Data, a kind of digitization under background and intelligentized carry out evaluating method to subject's health based on enteric microorganism information
It feels free to try, there is subversive innovation and promote meaning to traditional biological medical industry.
Firstly, a kind of detection and analysis method of intestinal microflora provided by the invention, based on the micro of subject
Fecal sample is detected, and be can effectively avoid existing embarrassment during business sampling operation, is kept detection more humanized.
Secondly, a kind of detection and analysis method of intestinal microflora provided by the invention, using PCR method in excrement
Microorganism total DNA is expanded, and gene cloning is bypassed, and the mode that hybridization or microorganism are separately cultured directly obtains a large amount of expansions
Increase sub-sequence information, sensitivity is higher, the species relative abundance of each microorganism in intestinal flora can be accurately detected, for further
Bioinformatic analysis is carried out, the diversity and abundance of subject enteric microorganism are accurately reflected.
Again, the detection and analysis method of a kind of intestinal microflora provided by the invention utilizes international advanced sequencing
Platform carries out 16S rRNA sequencing, and sequencing speed is fast, the sequencing time is short, and sequencing accuracy is high.After the sequencing is completed using life
The abundance and diversity information for believing big data analysis processing technique decryption intestinal microflora, gradually accumulate different crowd enteron aisle
The data of microbial flora, building belong to the enteric microorganism database of Chinese population, make the decline of sequencing cost, analyze result
Accuracy so that genetic test is gradually come into popular life.
Finally, by the personal information of input subject, by the enteron aisle of its enteric microorganism test data and Chinese population
Microbiological data library is matched, using the abundance and multiplicity of raw letter big data analysis processing technique decryption intestinal microflora
Property information, the composition of intestinal microflora is analyzed, a large amount of gene carries out metabolism group in binding sequence
(Metabonomics) research；It is found and body phase by metabolism with organism and related immunological diseases binding analysis
The mechanism of interaction, and then be adjusted according to human body basal conditions, increase human health index, analyzes current intestines for subject
Road health status simultaneously provides specific aim suggestion and is adjusted.
A kind of detection and analysis method of intestinal microflora provided by the invention saves liquid to excrement sample using excrement
Product are saved, it is ensured that can effectively be kept the diversity of intestinal microflora structure in 15 days at room temperature, be reduced intestines
Road microbial flora saves requirement condition and cost realizes the micro- life of enteron aisle in excrement under the specific conditions such as outgoing sampling, sample transport
Effective preservation of object flora.
A kind of detection and analysis method of intestinal microflora provided by the invention makes intestinal microflora detection more
Add it is quick and easy, efficient, can be formed high-flux sequence analysis in microbial function database.
The detection and analysis system of intestinal microflora provided by the invention, the detection of the intestinal microflora and
Analysis system has beneficial effect identical with the detection and analysis method of above-mentioned intestinal microflora, and details are not described herein.
The foregoing is merely illustrative of the preferred embodiments of the present invention, is not intended to limit the invention, all in original of the invention
Any modification made within then, equivalent replacement and improvement etc. should all be comprising within protection scope of the present invention.
Claims (10)
1. a kind of detection and analysis method of intestinal microflora, feature exist: the following steps are included:
Step S1: acquisition fecal sample；
Step S2: microbial flora genome DNA in fecal sample is extracted, and PCR amplification is carried out to the total DNA；
Step S3: the total DNA after amplification is carried out to build library and sequencing, to obtain raw sequencing data；
Step S4: raw sequencing data is subjected to data Quality Control processing, obtains sequencing data；
Step S5: carrying out bioinformatic analysis based on sequencing data, obtains analysis result；And
Step S6: providing an Intestinal microbial bacteria group database, by analysis result and the Intestinal microbial flora
Database is compared, and obtains the intestinal microflora state of subject；Intestinal microflora state based on subject
And its subject's basal conditions correspondence provides adjustable index and monitor control index.
2. intestinal microflora detection and analysis method according to claim 1, it is characterised in that: above-mentioned steps S1 into
One step includes the following steps:
Step S11: the appropriate fecal sample of sterile swab picking is used；
Step S12: by the sterile swab for speckling with excrement, stirring makes to save liquid discoloration in containing the test tube for saving liquid；And
Step S13: the discoloration containing fecal sample is saved into liquid and is stored at room temperature.
3. intestinal microflora detection and analysis method according to claim 2, it is characterised in that: the preservation liquid packet
It includes: thio hydrohalogenic acid salt, Organic Alcohol, complexing agent, phosphate buffer, tween, seralbumin and benzalkonium bromide；The thio hydracid
Salinity is 0.01M-5M, and the Organic Alcohol accounts for the 30%-60% for saving liquid total weight；The concentration of complexing agent is 0.001M-
0.35M；The phosphate buffer accounts for the 5%-15% for saving liquid weight；The tween accounts for the 0.05%-1% for saving liquid weight,
Sero-abluminous concentration is 2.5mM-100mM in the preservation liquid；Benzalkonium bromide accounts for the 0.01%-5% for saving liquid total weight.
4. intestinal microflora detection and analysis method according to claim 1, it is characterised in that: above-mentioned steps S2 into
One step includes the following steps:
Step S21: microbial flora genome DNA in fecal sample is extracted using CTAB method or SDS method；And
Step S22: it is carried out using the area the V3-V4 segment of the 16S rRNA gene in primer pair microbial flora genome DNA
Total DNA after being expanded.
5. intestinal microflora detection and analysis method according to claim 1, it is characterised in that: above-mentioned steps S3 into
One step includes the following steps:
Step S31: the DNA after amplification is purified to the total DNA after being expanded；
Step S32: based on the total DNA building mixing library after amplification；And
Step S33: it is sequenced by high-flux sequence platform, obtains raw sequencing data.
6. intestinal microflora detection and analysis method according to claim 1, it is characterised in that: above-mentioned steps S4 into
One step includes the following steps:
Step S41: the interference data in removal raw sequencing data；
Step S42: sequencing fragment is carried out to the raw sequencing data for going interference data to obtain and splices to obtain original tag；
Step S43: high quality is obtained by filtration by label interception and tag length, label is sequenced；And
Step S44: carrying out chimera and operate by the way that label is sequenced to high quality, and obtain can carrying out subsequent analysis has criterion
Label.
7. intestinal microflora detection and analysis method according to claim 1, it is characterised in that: above-mentioned steps S5 into
One step includes the following steps:
Step S51: carrying out OTUs clustering based on effective label, and obtains analysis result；
Step S52: group's group on kind of each level is belonged in Jie Men detailed outline section as a result, obtaining intestinal microflora based on analysis
At；And
Step S53: group is formed, and is associated analysis with the metabolic pathway of software prediction, obtain group's group be metabolized in pairs it is logical
The influence on road.
8. intestinal microflora detection and analysis method according to claim 7, it is characterised in that: above-mentioned steps S51
Further comprise following steps:
Step S511: picking out each effective label, be compared with RDP database, consistent with effective label greater than 97%
Property is clustered and obtains sequence taxon；
Step S512: the representative series of sequence taxon are picked out；And
Step S513: the division of species taxonomy information is carried out using presetting database in conjunction with representative series.
9. intestinal microflora detection and analysis method according to claim 7, it is characterised in that: above-mentioned steps S52
Further comprise following steps:
Step S521: significant difference analysis is carried out to the microbial flora different grouping using rank sum test method, to find out
Between the microbial flora for dividing generation significant difference influence group；And
Step S522: being analyzed using LEfSe, in all levels of microbial bacteria heap sort, passes through linear discriminant analysis, estimation
The size that each microbial flora abundance influences differential effect finds out the micro- life for dividing on sample and generating significant difference and influencing
Object flora.
10. a kind of intestinal microflora detection and analysis system, it is characterised in that: include:
Sample collection unit: it is configured to acquisition fecal sample；
PCR amplification unit: it is configured to extract microbial flora genome DNA in fecal sample, and the total DNA is carried out
PCR amplification；
Data capture unit: it is configured to that the total DNA after amplification is carried out to build library and sequencing, to obtain raw sequencing data；And
Raw sequencing data is subjected to data Quality Control processing, obtains sequencing data；
Bioinformatic analysis unit: being configured to carry out bioinformatic analysis based on sequencing data, obtains analysis result；And
Information comparison unit: being configured to provide for an Intestinal microbial bacteria group database, by analysis result and the crowd
Intestinal microflora database is compared, and obtains the intestinal microflora state of subject；Enteron aisle based on subject
Microbial flora state and its subject's basal conditions correspondence provide adjustable index and monitor control index.
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| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN110111897A (en) * | 2019-05-24 | 2019-08-09 | 天益健康科学研究院（镇江）有限公司 | A method of obtaining genital tract flora sequencer address |
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Citations (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2014036182A2 (en) * | 2012-08-29 | 2014-03-06 | California Institute Of Technology | Diagnosis and treatment of autism spectrum disorder |
| CN105385599A (en) * | 2016-01-21 | 2016-03-09 | 南京医科大学第一附属医院 | Coprophilous fungus preservation solution and method for preserving coprophilous fungi |
| CN107058490A (en) * | 2017-01-12 | 2017-08-18 | 杨俊杰 | The analysis method of enteron aisle and oral cavity flora diversity and otherness |
| CN108841974A (en) * | 2018-06-28 | 2018-11-20 | 北京水母科技有限公司 | Extract the method that fecal sample 16s rRNA is monitored in infant's intestinal microecology maturity |
| CN108893523A (en) * | 2018-05-31 | 2018-11-27 | 厦门安捷致善医学数据科技有限公司 | A kind of fecal specimens room-temperature extender |
| CN109122667A (en) * | 2018-09-27 | 2019-01-04 | 广州新诚生物科技有限公司 | Save liquid and its preparation method and application |
-
2019
- 2019-01-29 CN CN201910087145.2A patent/CN109706235A/en not_active Withdrawn
Patent Citations (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2014036182A2 (en) * | 2012-08-29 | 2014-03-06 | California Institute Of Technology | Diagnosis and treatment of autism spectrum disorder |
| CN105385599A (en) * | 2016-01-21 | 2016-03-09 | 南京医科大学第一附属医院 | Coprophilous fungus preservation solution and method for preserving coprophilous fungi |
| CN107058490A (en) * | 2017-01-12 | 2017-08-18 | 杨俊杰 | The analysis method of enteron aisle and oral cavity flora diversity and otherness |
| CN108893523A (en) * | 2018-05-31 | 2018-11-27 | 厦门安捷致善医学数据科技有限公司 | A kind of fecal specimens room-temperature extender |
| CN108841974A (en) * | 2018-06-28 | 2018-11-20 | 北京水母科技有限公司 | Extract the method that fecal sample 16s rRNA is monitored in infant's intestinal microecology maturity |
| CN109122667A (en) * | 2018-09-27 | 2019-01-04 | 广州新诚生物科技有限公司 | Save liquid and its preparation method and application |
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