CN109762915A - A kind of detection method and its dedicated kit of bacterial resistance gene - Google Patents
A kind of detection method and its dedicated kit of bacterial resistance gene Download PDFInfo
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Abstract
The invention discloses a kind of detection method of bacterial resistance gene and its dedicated kits.Kit provided by the invention includes 296 single stranded DNAs shown in sequence 1-296 in sequence table.Bacterial resistance gene detection method provided by the invention includes the following steps: that (1) extracts the genomic DNA of tested bacteria, is expanded using complete single stranded DNA to the genomic DNA, obtains amplified production；(2) sequencing library is constructed using the amplified production based on Ampliseq technology；(3) sequencing library is sequenced, obtains sequencing result, and the sequencing result is compared with drug resistant gene sequence, determines whether tested bacteria carries drug resistant gene.Detection method and kit of the invention high-throughput, fast and accurately can detect multiple drug resistant genes of sample to be tested simultaneously, have applications well prospect.
Description
Technical field
The present invention relates to field of biotechnology, and in particular to a kind of detection method and its special agent of bacterial resistance gene
Box.
Background technique
Bacterial resistance gene detection is of great significance in clinical diagnosis.It is resistance to using wider commercialization bacterium at this stage
Medicine gene tester mainly includes following four: the Vitek2 of automatic checkout system, flight mass spectrum MALDI-TOF, quickly
Biochemistry detection and real-time imaging monitoring.Vitek2 carries out showing preferable consistency when clinical sample detection, but it is examined
Test sample originally need to be positive culture, and general blood sample reports that the positive time needs at least 24 hours, greatly delays treatment
Opportunity.There is also same problems for real-time imaging monitoring.Flight mass spectrum and biochemistry detection can only determine produce carbapenem enzyme and
The pathogen of beta lactamase is difficult accurately to be detected for the bacterium of other drug resistances.In addition to this, with polymerase chain
Bacterial drug resistance detection method based on formula reaction also apply it is relatively wide, such as Prove it, FilmArray, Verigene etc., but
It is that these methods can only detect several frequently seen drug resistant gene, there are certain limitations.
Ampliseq technology refers to specific amplification target fragments during sequencing, finally surveys to the segment
The method that sequence determines base composition.Different from two traditional generation sequencing approaches, this method is not by genome by the way of ultrasound
It interrupts for short-movie section, but in same reaction system, is pre-placed primer, utilizes the interested mesh of the amplification of primer specificity
Standard film section, to generate the short-movie section for meeting subsequent sequencing reading length.This method can be under same reaction condition, same reactant
In system, up to 24000 pairs of primers are placed, target fragment is expanded.
Ampliseq technology has been widely applied in the screening of people's common disease at present, screening, heredity such as cancer
The screening of disease.In terms of etiological diagnosis, which is applied to the diagnosis of Ebola virus and the drug resistance of mycobacterium tuberculosis
The diagnosis of gene.Ampliseq technology shows the identification of excellent identification capacity and mutational site in various diagnosis
Ability shows stronger advantage in Multiple detection.The technology has become domestic and international disorder in screening, cause of disease screening and drug resistance sieve
The main technique methods looked into.
Summary of the invention
The first purpose of the invention is to provide complete single stranded DNAs.
Complete single stranded DNA provided by the invention 296 single stranded DNAs shown in sequence 1-296 in sequence table form.
A second object of the present invention is to provide the new applications of above-mentioned complete single stranded DNA.
The present invention provides above-mentioned complete single stranded DNAs in following a1)-a6) in it is any in application:
A1) detect or assist detection bacterium drug resistant gene；
A2) the product of preparation detection or auxiliary detection bacterium drug resistant gene；
A3 it) detects or assists detection tested bacteria whether to carry drug resistant gene and/or carry the type of drug resistant gene；
A4) whether preparation detection or auxiliary detection tested bacteria carry drug resistant gene and/or carry the type of drug resistant gene
Product；
A5) detect or assist whether detection sample to be tested contains the bacterium for carrying drug resistant gene；
A6) whether preparation detection or auxiliary detection sample to be tested contain the product for carrying the bacterium of drug resistant gene.
Third object of the present invention is to provide the kits for detection bacterium drug resistant gene.
Kit provided by the invention includes above-mentioned complete single stranded DNA.
The function of the kit be it is following 1) or 2) or 3):
1) detect or assist detection bacterium drug resistant gene；
2) it detects or assists detection tested bacteria whether to carry drug resistant gene and/or carry the type of drug resistant gene；
3) detect or assist whether detection sample to be tested contains the bacterium for carrying drug resistant gene.
Reagent needed for the kit may also include building library, reagent needed for progress PCR amplification, preparation sequencing mould
It reagent needed for plate and/or carries out that required reagent is sequenced.
Reagent needed for the building library can be 2.0 (Thermo Fisher of Ion Ampliseq Library Kit
Company, number No.4480441) in reagent.Reagent needed for the preparation sequencing template can be Ion PGM OT2 200
Reagent in Template Kit (Thermo Fisher company, number No.448094).It is described carry out that required reagent is sequenced can be
Ion PITM Hi-QTMReagent in 200 Kit of Sequencing (Thermo Fisher company, number No.26433).
Final object of the present invention be to provide it is a kind of detection or auxiliary detection tested bacteria whether carry drug resistant gene
And/or the method for carrying the type of drug resistant gene.
Whether detection or auxiliary detection tested bacteria provided by the invention carries drug resistant gene and/or carries drug resistant gene
The method of type includes the following steps:
(1) genomic DNA for extracting tested bacteria, expands the genomic DNA using above-mentioned complete single stranded DNA
Increase, obtains amplified production；
(2) sequencing library is constructed using the amplified production based on Ampliseq technology；
(3) sequencing library is sequenced, obtains sequencing result, and by the sequencing result and drug resistant gene sequence
It is compared, determines whether tested bacteria carries drug resistant gene and carry the type of drug resistant gene.
In above-mentioned application or kit or method, the drug resistant gene is following at least one: aac (2 ')-lb, aadA1,
aadA3、aadB、aadD、adeb、ant(4’-llb)、aph3-la、armA、blaZ、catB3、catB6、catB8、cmlA5、
CTX-M-14、dfrA19、dfrG、E.coli_gyrA、E_coli_parC、IMP-1、KPC-1、mecA、mecC、mcr-1、
mtrC、mtrD、mtrE、mtrR、NDM-1、oprD、oqxA、oqxB、blaOXA-10、blaOXA-23、blaOXA-24、blaOXA-1、
blaOXA-51、blaOXA-58、blaOXA-66、blaOXA-69、blaOXA-82、PAO_gyrA_parC、patA、patB、pbp1a、pbp2b、
pbp2x、phoP、phoQ、pmrA、pmrB、qacA、qnrA1、qnrB4、rpsJ、sat-1、blaSHV-60、strA、strB、sul1、
blaTEM-12、tetA、tetD、tetM、tetR、vanA、vanB、VIM-2。
The bacterium is following at least one: Acinetobacter bauamnnii Acinetobacter baumannii, enterococcus faecalis
Enterococcus faecalis, enterococcus faecium Enterococcus faecium, Klebsiella Pneumoniae Klebsiella
Pneumoniae, the Klebsiella Pneumoniae Klebsiella pneumoniae with beta- with beta- lactam resistant
Lactamase, enterococcus vancomycin-resistant Enterococci of drug resistance of vancomycin, pseudomonas aeruginosa
Pseudomonas aeruginosa, staphylococcus aureus Staphylococcus aureus, staphylococcus epidermis
Staphylococcus epidermidis, staphylococcus haemolyticus Staphylococcus haemolyticus, Human fetal cardiomyocytes
The staphylococcus aureus methicillin-resistant of Staphylococcus hominis, methicillin resistance
Staphylococcus aureus。
The sample to be tested can be blood sample.
The present invention provides a kind of detection method of bacterial resistance gene and its dedicated kits.Reagent provided by the invention
Box includes 296 single stranded DNAs shown in sequence 1-296 in sequence table.Bacterial resistance gene detection method packet provided by the invention
It includes following steps: (1) extracting the genomic DNA of tested bacteria, the genomic DNA is expanded using complete single stranded DNA,
Obtain amplified production；(2) sequencing library is constructed using the amplified production based on Ampliseq technology；(3) to the sequencing text
Library is sequenced, and is obtained sequencing result, and the sequencing result is compared with drug resistant gene sequence, is determined that tested bacteria is
The no type for carrying drug resistant gene and carrying drug resistant gene.Detection method of the invention, can be high-throughput, fast in primary first-order equation
68 drug resistant genes of sample to be tested are detected while fast, accurate, thus it is speculated that its drug-resistant phenotype provides guidance for clinic accurate medication
Property effect, reduce abuse of antibiotics and the superbacteria that therefore generates, there is applications well prospect.
Specific embodiment
Embodiment below facilitates a better understanding of the present invention, but does not limit the present invention.Experiment in following embodiments
Method is unless otherwise specified conventional method.Test material as used in the following examples is unless otherwise specified certainly
What routine biochemistry reagent shop was commercially available.Quantitative test in following embodiment is respectively provided with three repeated experiments, as a result makes even
Mean value.
The Klebsiella Pneumoniae with beta- lactam resistant in following embodiments is recorded in document: Feng J, Yin Z,
Zhao Q,et al.Genomic characterization of novel IncFII-type multidrug
resistant plasmids p0716-KPC and p12181-KPC from Klebsiella pneumoniae[J].Sci
Rep, 2017,7 (1): in 5830., the public can obtain from applicant.
The staphylococcus aureus of methicillin resistance in following embodiments is recorded in document: Li L, Pian Y, Chen
S,et al.Phenol-soluble modulin alpha4 mediates Staphylococcus aureus-
associated vascular leakage by stimulating heparin-binding protein release
From neutrophils [J] .Sci Rep, in 2016,6:29373., the public can obtain from applicant.
Embodiment 1, the acquisition of bacterial resistance gene and its design of detection primer
One, the acquisition of drug resistant gene
According to the type of clinical common antibiotics and the type of common clinical substance, in The Comprehensive
Antibiotic Resistance Database(CARD；Http:// apcard.mcmaster.ca) database and ARDB number
According to the associated clip of downloading drug resistant gene in library (http://ardb.cbcb.umd.edu), and according to pathogen, antibiotic, resistance to
The type of medicine gene is arranged, and method for sorting is as follows: by the drug resistant gene downloaded in ARDB database and CARD database
Associated clip is clustered by cd-hit, is threshold value, de-redundancy sequence with 95%；BLAST is reused to plant the segment of acquisition
Between compare, be threshold value with 97%, the specific fragment in drug resistance site screened, remove redundant sequence.Finally it is obtained 68
It is a to be often used that antibiotic generates drug resistant drug resistant gene and its sequence, 68 drug resistant genes are as follows to clinic: aac (2 ')-lb,
aadA1、aadA3、aadB、aadD、adeb、ant(4’-llb)、aph3-la、armA、blaZ、catB3、catB6、catB8、
cmlA5、CTX-M-14、dfrA19、dfrG、E.coli_gyrA、E_coli_parC、IMP-1、KPC-1、mecA、mecC、mcr-
1、mtrC、mtrD、mtrE、mtrR、NDM-1、oprD、oqxA、oqxB、blaOXA-10、blaOXA-23、blaOXA-24、blaOXA-1、
blaOXA-51、blaOXA-58、blaOXA-66、blaOXA-69、blaOXA-82、PAO_gyrA_parC、patA、patB、pbp1a、pbp2b、
pbp2x、phoP、phoQ、pmrA、pmrB、qacA、qnrA1、qnrB4、rpsJ、sat-1、blaSHV-60、strA、strB、sul1、
blaTEM-12, tetA, tetD, tetM, tetR, vanA, vanB, VIM-2, GenBank and the submission date difference it is as shown in table 1.
Table 1, the GenBank of drug resistant gene and submission date
Note: | AF078527 |+| 3770-4303 | the sequence for indicating aadB drug resistant gene is GenBank for AF078527 institute
3770-4303 of the gene order shown, other are similarly.
The antibiotic and drug resistant gene that different pathogens generate are specifically as shown in table 2.
The antibiotic and drug resistant gene that table 2, different pathogens generate
Two, the design of primer
68 drug resistant gene sequences that step 1 obtains are committed to Ampliseq design of primers website Ion Ampliseq
Design of primers is carried out on Designer (http://www.ampliseq.com/browse.action), to the primer sequence of return
It is listed on NCBI and is compared, evaluate its specificity, and carry out adjustment and optimization appropriate, final acquisition is resistance to for detecting 68
The primer of medicine gene is totally 148 to (table 3).
Table 3, the primer sequence for detecting 68 drug resistant genes
Embodiment 2, for detection bacterium drug resistant gene primer application
(1) the drug resistant gene detection of pathogen to be measured
One, the extraction of pathogen nucleic acid
Using the MagAttract HMW DNA Kit kit of Qiagen, extracted using paramagnetic particle method to be detected in table 4
Pathogen nucleic acid.Specific extraction step is carried out according to the specification of kit.
Table 4, pathogen to be detected
Two, the building of gene library
2.0 (Thermo Fisher of Ion Ampliseq Library Kit is utilized based on Ampliseq technology
Scientific, Cat.no.4480441) carry out library construction.Specific step is as follows:
1, the DNA target amplification reaction in 2 primer ponds is established
Ampliseq primer is divided into primer pond 1 (primer pool 1) and primer pond 2 (primer pool 2), primer pond
For primer in 1 by marking the primer for being Pool=1 " to form in table 2, the primer in primer pond 2 is Pool=by marking in table 2
2 " primer composition, concentration of the every primer in primer pond 1 or primer pond 2 is 100nM.
In 96 orifice plates, totally two reacting holes, primer are respectively to draw in primer and primer pond 2 in primer pond 1 to each sample
Object.The system of each reaction tube is 10 microlitres: 2 × primer pool:5 microlitre, 5 × Ion AmpliseqTMHiFi Mix:2
Microlitre, Nuclease-free Water:0.5 microlitre, DNA:2.5 microlitres.After above-mentioned system is prepared as requested, according to table 5
Shown in reaction condition reacted (recurring number 19).
Table 5, reaction condition
| Step | Temperature | Time |
| Initial denaturation | 99℃ | 2 minutes |
| Denaturation | 99℃ | 15 seconds |
| Annealing and extension | 60℃ | 4 minutes |
| - | 10℃ | Hold |
2, target amplification is reacted and is merged
After completing step 1, to each sample, the reaction product that 10 microlitres of primer ponds 1 are obtained and 10 microlitres of primer ponds 2 are obtained
To reaction product merge, the total volume of each sample is 20 microlitres.
3, partial digested product
(1) 2 microlitres of FuPa reagents (brown lid) are added into each sample cell, total volume is 22 microlitres at this time.
(2) 96 orifice plates are put on PCR reaction instrument, program shown in table 6.
Table 6, response procedures
| Temperature | Time |
| 50℃ | 10 minutes |
| 55℃ | 10 minutes |
| 60℃ | 20 minutes |
| 10℃ | (most 1 hours) |
4, connector connects
Connector (Adapter) is connected on amplicon and is purified.Specific step is as follows:
(1) connection reaction
96 orifice plate sealed membranes are carefully removed, the reagent being sequentially added into each reacting hole in table 7 later.
Table 7, reagent
| Order of addition | Ingredient | Volume |
| 1 | Switch Solution (yellow lid) | 4 microlitres |
| 2 | The Ion Xpress barcode adapter mix diluted | 2 microlitres |
| 3 | DNA ligase (blue cap) | 2 microlitres |
| - | Total volume | 30 microlitres |
96 orifice plates are put on PCR reaction instrument, program shown in table 8.
Table 8, response procedures
| Temperature | Time |
| 22℃ | 30 minutes |
| 68℃ | 5 minutes |
| 72℃ | 5 minutes |
| 10℃ | (most 24 hours) |
(2) purified library
The Agencourt of 45 microlitres (1.5 × sample volume) is addedTMAMPure Reagent (BeckmanCoulter,
Cat.A63880) into library.5 mixings of piping and druming up and down, are incubated at room temperature 5 minutes, 96 orifice plates are placed on magnetic frame,
It is incubated for 2 minutes later until solution clarification, under the premise of without impinging on sediment, carefully removes and discard supernatant.It is added 150
Microlitre 70% ethyl alcohol newly configured, washes pearl, carefully removes later and discard supernatant；Reuse 70% ethyl alcohol cleaning second
It is secondary；Ensure that the ethyl alcohol in each reacting hole is removed, 96 orifice plates are placed on magnetic frame, are placed at room temperature for 5 minutes.
5, the library not expanded is quantified using qPCR
The library Ion AmpliseqTM not expanded is diluted, passes through Ion using the method for qPCR later
LibraryQuantitation Kit (Thermo Fisher Scientific, Cat.no.4468802) is to library
Concentration quantified.The concentration in the library not expanded is generally 100-500pM.After quantitative, library is diluted to 100pM,
Template preparation is carried out using Ion template Kit so as to subsequent.
Three, template prepares and is sequenced
The library that step 2 is obtained using 200 Template Kit of Ion PGM OT2 (Thermo Fisher company,
Number No.448094) kit preparation sequencing template, obtained sequencing template is subjected to upper machine sequencing, template in sequencing procedure
Emulsification and be enriched in operate in Ion One Touch System (Thermo Fisher company) and complete, the sequencing of upper machine uses
Ion P1 chip operates completion in Ion Proton sequenator (Thermo Fisher company).All steps are according to explanation
Book is operated.
Four, sequencing result is analyzed
The raw data of sequencing gained bam format switch to sam file by Samtools (version 1.6) software, extract later
The sequencing result of effective fasta format, filters out low-quality data and short-movie is had no progeny, use bowtie (version
1.2.0) software is compared with the drug resistant gene sequence of the pathogen of NCBI downloading respectively, comparison result is extracted, according to survey
Gene reads number obtained by sequence and ratio, determine the drug resistant gene type that pathogen to be measured contains.
Samtools order line are as follows: samtools view-h original series bam file > output sam file；Bowtie
Order line are as follows: 1-al of bowtie-f-a-m 20-v compares the outgoing route-that outgoing route-un of upper file is not compared
The path norc reference needs the sequential file comparison result outgoing route compared.
Five, drug resistant gene testing result
The drug resistant gene testing result of each pathogen is as shown in table 9.
The drug resistant gene testing result of table 9, each pathogen
(2) the drug resistance evaluation of pathogen to be measured
One, pathogen drug-resistant phenotype detects
The drug-resistant phenotype of each pathogen shown in table 4 is sieved using 13 kinds of variety classes antibiotic shown in table 10
It looks into.Specific step is as follows: after 37 DEG C of shaking tables of each bacterium shown in table 4 are incubated overnight 8 hours, being diluted to 0.5 wheat using PBS
Family name's turbidity.Cotton swab dips the bacterium solution after dilution, is uniformly coated on MHA culture medium.Drug sensitive test paper is clamped using tweezers, according to
The requirement of CLSI posts corresponding drug sensitive test paper on MHA culture medium.Different bacterium are carried out 18-24 hours according to the requirement of CLSI
Culture, measures the diameter of inhibition zone using vernier caliper after culture, and is compared with the file of 2018 editions CLSI, determines
Sensitivity, intermediary and the drug resistance of the pathogen.Specific steps reference literature " Urbaniak C, the Sielaff A of drug resistance evaluation
C,Frey K G,et al.Detection of antimicrobial resistance genes associated with
the International Space Station environmental surfaces[J].Sci Rep,2018,8(1):
Method in 814. ".
Table 10, Antibiotics
| Medicine name | Dosage | Unit | Antibiotics |
| Gentamicin | 10 | μg | Aminoglycosides |
| Cefoxitin | 30 | μg | Cephalosporins |
| Oxacillin | 1 | μg | Penicillins |
| Chloramphenicol | 30 | μg | Chloramphenicol |
| Tetracycline | 30 | μg | Tetracyclines |
| Penicillin | 10 | units | Penicillins |
| Lavo-ofloxacin | 5 | μg | Quinolones(fluoroquinolones) |
| Cefotaxime | 30 | μg | Cephalosporins |
| Imipenem | 10 | μg | Carbapenems |
| Piperacillin | 100 | μg | Penicillins |
| Minocycline | 30 | μg | Tetracyclines |
| Vancomycin | 30 | μg | Glycopeptides |
| Ampicillin | 10 | μg | Penicillins |
Two, testing result
Each pathogen is as shown in table 11 to the screening results of the drug-resistant phenotype of 13 kinds of variety classes antibiotic.
Table 11, each pathogen are to the screening results of the drug-resistant phenotype of 13 kinds of variety classes antibiotic
The drug-resistant phenotype testing result and drug resistant gene testing result of each pathogen are almost the same, illustrate method of the invention
Whether detection tested bacteria that can be accurate and effective carries drug resistant gene and entrained drug resistant gene type.
Sequence table
<110>PLA Academy of Military Sciences's military medical research institute
<120>detection method and its dedicated kit of a kind of bacterial resistance gene
<160> 296
<210> 1
<211> 20
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 1
atgcacgcct tcatctgtca 20
<210> 2
<211> 21
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 2
tgtggtgtcg agttccattc c 21
<210> 3
<211> 20
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 3
gacacagcca gaggcatgta 20
<210> 4
<211> 18
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 4
gccagtcgca ggtgatct 18
<210> 5
<211> 22
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 5
cgcctttcac gtagtggaca aa 22
<210> 6
<211> 26
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 6
ggttcctgaa caggatctat ttgagg 26
<210> 7
<211> 22
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 7
ggctcgaaga tacctgcaag aa 22
<210> 8
<211> 29
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 8
gaagtatcga ctcaactatc agaggtagt 29
<210> 9
<211> 22
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 9
ggtagcggtg accatcgaaa tt 22
<210> 10
<211> 24
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 10
tccataaggt cattgagcaa tgct 24
<210> 11
<211> 23
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 11
cccggttcct gaacaggatc tat 23
<210> 12
<211> 27
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 12
tctcgccttt cacgtagtga ataaatt 27
<210> 13
<211> 22
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 13
atggacacaa cgcaggtcac at 22
<210> 14
<211> 24
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 14
atccatagtc caactcctcc atga 24
<210> 15
<211> 23
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 15
ctcgaggcaa tagttgaaat gct 23
<210> 16
<211> 21
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 16
cacgcaagac ctcaaccttt t 21
<210> 17
<211> 27
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 17
atgaatatga atggacctgc atcaatg 27
<210> 18
<211> 26
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 18
cggttgcagc gtcttttaac agtata 26
<210> 19
<211> 22
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 19
gtggaaagcg gaagttaacg tt 22
<210> 20
<211> 27
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 20
gcttgccaac atattcatac atttccc 27
<210> 21
<211> 29
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 21
ggcaccacaa caatataatc gttataacg 29
<210> 22
<211> 30
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 22
ccaataatgg taattagccc gattttgaag 30
<210> 23
<211> 29
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 23
tcattgccat tatgtctaga gggttaatg 29
<210> 24
<211> 26
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 24
ggaaaatagc ttttctactg caccca 26
<210> 25
<211> 19
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 25
ccgttgggtt gatcgcctt 19
<210> 26
<211> 22
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 26
tctatccagg tgcggtaatc ct 22
<210> 27
<211> 29
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 27
gatatcgatt tcgatgtatt ggtaagcac 29
<210> 28
<211> 23
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 28
aacaatgtcg gcactaactt tgc 23
<210> 29
<211> 24
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 29
gtgagaaatc accatgagtg acga 24
<210> 30
<211> 30
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 30
cagcattcca ggtattagaa gaatatcctg 30
<210> 31
<211> 22
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 31
ctgccagcgc atcaacaata tt 22
<210> 32
<211> 24
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 32
ccatattcaa cgggaaacgt cttg 24
<210> 33
<211> 33
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 33
atggataaga atgatgttgt taagaagata ctt 33
<210> 34
<211> 33
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 34
accaaaatct aaaatagatg agacatgttt gat 33
<210> 35
<211> 30
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 35
gagtcgcaac attaaatgac ttttacactt 30
<210> 36
<211> 26
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 36
cgtcgtcttt aacttcccaa taatgc 26
<210> 37
<211> 30
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 37
tttctagaag catatgttat tgcttgacca 30
<210> 38
<211> 30
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 38
gaggcttcaa tgaagtacag tgataataca 30
<210> 39
<211> 30
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 39
tttttgattc caccgatttc gtttataatt 30
<210> 40
<211> 30
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 40
cagttcacat gccaaagagt taaatgattt 30
<210> 41
<211> 22
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 41
ggtacgactg ggcatcatct tt 22
<210> 42
<211> 23
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 42
tgcaggccaa caatattaga cga 23
<210> 43
<211> 22
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 43
ccttcaaagg caagctgctt tc 22
<210> 44
<211> 25
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 44
gagaatgcag gttcttcctg catat 25
<210> 45
<211> 28
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 45
atggaaaatt actttgacag tcccttca 28
<210> 46
<211> 22
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 46
aagctggctc ttcttgcatg ta 22
<210> 47
<211> 22
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 47
cggcatgact gggtaacatc tt 22
<210> 48
<211> 25
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 48
ctgtgcagac caaaaatgtc tgaag 25
<210> 49
<211> 29
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 49
atgaaaaact actttaacag ccctttcaa 29
<210> 50
<211> 22
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 50
gaaagcaggc tcctcttgca ta 22
<210> 51
<211> 22
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 51
gcatgactgg gcatcatcct tc 22
<210> 52
<211> 28
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 52
acttatgcag accaaaaata tttgacga 28
<210> 53
<211> 22
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 53
ctttagttgg cggtactccc tt 22
<210> 54
<211> 24
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 54
actgttgcaa atgtggaaac aagg 24
<210> 55
<211> 22
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 55
gcttcggatt cttcaggctt gt 22
<210> 56
<211> 22
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 56
gacgaagaag ctacccattc ca 22
<210> 57
<211> 20
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 57
cgatcggcga tgagacgttt 20
<210> 58
<211> 24
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 58
gggtaaaata ggtcaccaga acca 24
<210> 59
<211> 23
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 59
gtgacaaaga gagtgcaacg gat 23
<210> 60
<211> 22
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 60
gtactgcaca ttggaaagcg tt 22
<210> 61
<211> 22
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 61
gagtcaccca caacttgagc ta 22
<210> 62
<211> 22
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 62
tcgaggagat cacgaacgat tc 22
<210> 63
<211> 25
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 63
gagcgaatca aggagaaagg aattc 25
<210> 64
<211> 22
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 64
aagtgaatgc gctcttgagt ct 22
<210> 65
<211> 26
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 65
atgaaagttt ctttgattgc tgcgat 26
<210> 66
<211> 30
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 66
cgattgattc aaggttcttc ctacctaata 30
<210> 67
<211> 32
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 67
gactgggaat atgttaaaaa tactacaaag gg 32
<210> 68
<211> 32
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 68
acggattttt atcattcttt atcccttttt gg 32
<210> 69
<211> 33
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 69
tgtattcttt atatttttgt tttgcagcat tgc 33
<210> 70
<211> 28
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 70
gccgtaaatg gagtgtcaat taggtaag 28
<210> 71
<211> 25
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 71
ctaaacatgg tttggtggtt cttgt 25
<210> 72
<211> 28
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 72
caccgaataa tattttcctt tcaggcaa 28
<210> 73
<211> 22
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 73
tgtcactgta tcgccgtcta gt 22
<210> 74
<211> 27
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 74
cctgttgtca gatatttttc cgagatg 27
<210> 75
<211> 22
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 75
ccatccgtta cggcaaaaat gc 22
<210> 76
<211> 24
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 76
cctttagcca atcaacaaac tgct 24
<210> 77
<211> 22
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 77
gctactgatc accacgctgt ta 22
<210> 78
<211> 26
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 78
cccaaaccaa tgatacgcat gataaa 26
<210> 79
<211> 22
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 79
caagccgaga ccaaggatct at 22
<210> 80
<211> 24
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 80
catatcaggc ttggttgctt gtac 24
<210> 81
<211> 28
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 81
gaagtagaaa tgactgaacg tccgataa 28
<210> 82
<211> 25
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 82
atatgctgtt cctgtattgg ccaat 25
<210> 83
<211> 27
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 83
tcagaacgtg gtaaaatttt agaccga 27
<210> 84
<211> 29
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 84
cagagttaat gggaccaaca taacctaat 29
<210> 85
<211> 33
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 85
atcatcataa acttttccag atatagtagc att 33
<210> 86
<211> 32
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 86
gaacgtgtag ttaataaaac acatagggat ga 32
<210> 87
<211> 28
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 87
ccagatttac caataattcg ggcataat 28
<210> 88
<211> 23
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 88
agcaaggtat gcaagatttg gga 23
<210> 89
<211> 22
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 89
agaagcaggt ttggcttcag at 22
<210> 90
<211> 23
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 90
gccgtaccga acgatcagaa tat 23
<210> 91
<211> 22
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 91
atcagcacgc gcacatacag ac 22
<210> 92
<211> 22
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 92
cgatttccgg ctttatcggt ca 22
<210> 93
<211> 22
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 93
gcgatgccaa aaccgtcttt aa 22
<210> 94
<211> 22
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 94
ttacccaact ggcgaaaagc at 22
<210> 95
<211> 24
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 95
ggcggaagtg gacatataat ccaa 24
<210> 96
<211> 25
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 96
ccatttttgc gtgggttatt tcgat 25
<210> 97
<211> 28
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 97
ctacattgaa aactaccttg acctctgt 28
<210> 98
<211> 22
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 98
ggctgtacgc aaactggtat tg 22
<210> 99
<211> 22
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 19
cctgcaaaag ctgatcgaca tc 22
<210> 100
<211> 25
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 100
cataacgctc gttgaaatag gcttt 25
<210> 101
<211> 26
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 101
atgagaaaaa ccaaaaccga agcctt 26
<210> 102
<211> 28
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 102
gtgttcgcat tttaaaaaca ggatgttg 28
<210> 103
<211> 22
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 103
gctgcaaagc aacgacatct ac 22
<210> 104
<211> 27
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 104
ggttttccaa gttgtccatc attatcc 27
<210> 105
<211> 23
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 105
aacggtgata ttgtcactgg tgt 23
<210> 106
<211> 22
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 106
gcttccaacg gtttgatcgt ca 22
<210> 107
<211> 22
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 107
gcaggttgat ctcctgcttg at 22
<210> 108
<211> 25
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 108
atggaattgc ccaatattat gcacc 25
<210> 109
<211> 22
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 109
tacaggatcg acagcggata gt 22
<210> 110
<211> 23
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 110
ctgtccagta ctccgacttc aac 23
<210> 111
<211> 22
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 111
ccataggagg ctaggttcag gt 22
<210> 112
<211> 23
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 112
gccgaactcg aagacatcta tcg 23
<210> 113
<211> 25
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 113
aaagacggtg tacgtctact ttgac 25
<210> 114
<211> 29
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 114
gctgtgcttt accatcttta tcaacgata 29
<210> 115
<211> 22
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 115
ggtactgacc gatcaggatc gt 22
<210> 116
<211> 20
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 116
gcaacggttt tggcgttaac 20
<210> 117
<211> 22
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 117
cgatcaacgg cgttgaaaac at 22
<210> 118
<211> 23
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 118
gccagctcat ccttcacttt cag 23
<210> 119
<211> 23
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 119
aagtacgact cgctgtatat gcg 23
<210> 120
<211> 24
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 120
ccaaactctt cttcggtatg caga 24
<210> 121
<211> 27
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 121
ttgttagccg ttaaaattaa gcccttt 27
<210> 122
<211> 27
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 122
cgttgcacac tttgctttat tgaattg 27
<210> 123
<211> 27
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 123
ttttacttta cgatgcatcc acaaacg 27
<210> 124
<211> 26
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 124
tactttcgag ccatgcttct gttaat 26
<210> 125
<211> 22
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 125
gtcaatggtg tcttcgtgct tt 22
<210> 126
<211> 22
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 126
ggccttccaa ccagaatttg tc 22
<210> 127
<211> 30
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 127
aaaaattttc ctatggcaac cagaatatca 30
<210> 128
<211> 25
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 128
tccatgattt tggtgggaat ggatt 25
<210> 129
<211> 24
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 129
cctcaggtgt gctggttatt caaa 24
<210> 130
<211> 24
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 130
cctgctgtcc aatttcagca ttac 24
<210> 131
<211> 30
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 131
tcttgatctc atgcaaaaag aagtaaaacg 30
<210> 132
<211> 29
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 132
atcaataatt cattacgtat agatgccgg 29
<210> 133
<211> 29
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 133
tattttactt gctatgtggt tgcttctct 29
<210> 134
<211> 30
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 134
gcggcttaga gcattaccat atagattaat 30
<210> 135
<211> 33
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 135
acttcctata ttcagcattt ctattctagt ttc 33
<210> 136
<211> 27
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 136
gcagggacat attctttatt tgctcgt 27
<210> 137
<211> 27
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 137
gagggtaaaa atcttagcac ctatggt 27
<210> 138
<211> 30
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 138
ggcaaaatta acttcttgta ctggtgtaat 30
<210> 139
<211> 30
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 139
gaacacaggt cgataatttt tggttagttg 30
<210> 140
<211> 30
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 140
ttaaatgatt ccaagatttt ctagcgactt 30
<210> 141
<211> 33
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 141
atgaacatta aaacactctt acttataaca agc 33
<210> 142
<211> 26
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 142
atgtcctttt cccattctgg gaatag 26
<210> 143
<211> 24
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 143
agtatttaag tgggacgggc aaaa 24
<210> 144
<211> 30
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 144
tcttctatga ataacatgga ttgcacttca 30
<210> 145
<211> 26
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 145
ctaataaaac gcttccattt agccca 26
<210> 146
<211> 33
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 146
atttctaagt taagggagaa cgctacaata ttt 33
<210> 147
<211> 26
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 147
gatgctgtgt ttgtcacata tgatgg 26
<210> 148
<211> 28
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 148
ccctttcaac caaaattgat caacttcc 28
<210> 149
<211> 27
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 149
ttgcaacgta ttggttatgg caatatg 27
<210> 150
<211> 30
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 150
cccaacttat ctagcacatc taaagacaat 30
<210> 151
<211> 31
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 151
gagtttagtt tgcttaagca taagtattgg g 31
<210> 152
<211> 25
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 152
cggtctaaat gcgtgccata ttttt 25
<210> 153
<211> 32
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 153
cttatatagt gactgctaat ccaaatcaca gt 32
<210> 154
<211> 24
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 154
tcttttccca ttctgggaat agcc 24
<210> 155
<211> 27
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 155
aaccaccaca gaagtattta aatggga 27
<210> 156
<211> 27
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 156
tgcctcttgc tgaggagtaa tttttag 27
<210> 157
<211> 20
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 157
cagccgttct cctatcgcta 20
<210> 158
<211> 20
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 158
gcacgtgttc gcacaattcg 20
<210> 159
<211> 20
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 159
gtggactggg taccgaactt 20
<210> 160
<211> 20
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 160
ggtctcgtag accttctgca 20
<210> 161
<211> 20
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 161
accgaagccg agatcatcac 20
<210> 162
<211> 22
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 162
gtcatcagct cttcgacatc ga 22
<210> 163
<211> 20
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 163
cgacccgcat cgtcatcatc 20
<210> 164
<211> 20
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 164
gcacctcgct gagttcgaag 20
<210> 165
<211> 18
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 165
tcgagcgcat cggcttta 18
<210> 166
<211> 22
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 166
ctcccaggca gtcgataaac tc 22
<210> 167
<211> 25
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 167
ggaagtttaa atacgcttgt cgaca 25
<210> 168
<211> 22
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 168
cgccaagtga tttaccgtgg aa 22
<210> 169
<211> 23
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 169
caagtaggta ccttgcggtt tca 23
<210> 170
<211> 25
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 170
aagatttgac cctctttggt cacaa 25
<210> 171
<211> 26
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 171
tttgaaggca ggattgatgg tattga 26
<210> 172
<211> 30
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 172
aacagctatt caaaccttta caaaagaagg 30
<210> 173
<211> 24
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 173
gtgggtagac aggcgtgtta atta 24
<210> 174
<211> 21
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 174
ccaaccgttt agggcaccat a 21
<210> 175
<211> 23
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 175
atgaacaaac caacgattct gca 23
<210> 176
<211> 24
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 176
ccaggatacg gatggtatca atcc 24
<210> 177
<211> 26
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 177
gtttctatcg aagaccatcg cttctt 26
<210> 178
<211> 30
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 178
agcaaggcta actgaggtaa acttaaatta 30
<210> 179
<211> 25
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 179
gacagcagct caaaactact atggt 25
<210> 180
<211> 27
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 180
cttggtctac atttgtgtag acatcca 27
<210> 181
<211> 27
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 181
ggaagtttcc aaaacctttc tatccca 27
<210> 182
<211> 27
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 182
agcatcagtt ctccaaattt gacagaa 27
<210> 183
<211> 29
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 183
agatttcagg atcagccaaa tagtaatcc 29
<210> 184
<211> 28
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 184
aaatttaaca gccattcgat tccgattc 28
<210> 185
<211> 31
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 185
ggtatttttg atacagattg ataatgtcac g 31
<210> 186
<211> 22
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 186
ggtacagccg aaagctatgt gg 22
<210> 187
<211> 28
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 187
tttagtcaat tttgcggtca ttattggg 28
<210> 188
<211> 25
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 188
gctccaaagg aaacttgctt gagat 25
<210> 189
<211> 28
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 189
tggaagaatc ctatgtaaga gagcaact 28
<210> 190
<211> 25
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 190
taccgtttgt tgggaaactt gttct 25
<210> 191
<211> 23
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 191
atgaaactgc tggtagtgga aga 23
<210> 192
<211> 20
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 192
gaacggcttg accacgtagt 20
<210> 193
<211> 21
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 193
caagtccttc ccgatcctga t 21
<210> 194
<211> 21
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 194
gtgatgccgc atgaggtatt c 21
<210> 195
<211> 18
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 195
cgctgaccgc ctacgaat 18
<210> 196
<211> 19
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 196
gcgctcggtg aacaggtag 19
<210> 197
<211> 21
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 197
tacagcatcg tcaccatgca a 21
<210> 198
<211> 20
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 198
ctgcgcaaca ggcggttaag 20
<210> 199
<211> 20
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 199
cgagctggac cagatcgaat 20
<210> 200
<211> 23
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 200
tgatagccta tctgctggct cat 23
<210> 201
<211> 201
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 201
cctgctggag aacgcctatc 20
<210> 202
<211> 21
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 202
tgtagcgaaa cgtatgcgga a 21
<210> 203
<211> 20
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 203
atgagaatac tgctggccga 20
<210> 204
<211> 20
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 204
ctgcagttcg tcgagatcga 20
<210> 205
<211> 18
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 205
gtgccgacga ctacctga 18
<210> 206
<211> 18
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 206
tcgatgccgt agccgatg 18
<210> 207
<211> 19
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 207
gggtgctcca gctgaacat 19
<210> 208
<211> 18
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 208
ggcgtgcgga tttcatgg 18
<210> 209
<211> 20
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 209
ggcctcatcg acgaactcaa 20
<210> 210
<211> 21
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 210
agctcgatgt ccttgtcgat g 21
<210> 211
<211> 21
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 211
ggctccatca ccgagatcat c 21
<210> 212
<211> 23
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 212
caggttttcg ctgatccaga tct 23
<210> 213
<211> 30
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 213
aaaaagatgg actgcactag tagtattagc 30
<210> 214
<211> 29
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 214
ctgcaatacc aagtaaaaat cgaatagct 29
<210> 215
<211> 29
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 215
cattagctat atttttcgca gaaagtgca 29
<210> 216
<211> 28
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 216
gtccaatcat gcctgcaatt gataaaat 28
<210> 217
<211> 27
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 217
caaaagaaaa gtctcactcg tgggata 27
<210> 218
<211> 29
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 218
acaacctgta accattgtga agctaataa 29
<210> 219
<211> 30
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 219
gataaagttt ttcagcaaga ggatttctca 30
<210> 220
<211> 24
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 220
gttgcaacct gagatataag ccga 24
<210> 221
<211> 27
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 221
gcttctacaa tcaagtcagc cataaga 27
<210> 222
<211> 23
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 222
cagcactatt actcccaagg gtt 23
<210> 223
<211> 22
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 223
atgatgactc tggcgttagt tg 22
<210> 224
<211> 27
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 224
gtattggtga tataggcgct acaaaac 27
<210> 225
<211> 27
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 225
ggcgcaagtt ttatgaatat gatcacc 27
<210> 226
<211> 23
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 226
ataccaagac gttccaggag caa 23
<210> 227
<211> 18
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 227
ccggcaaatc cagcttca 18
<210> 228
<211> 23
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 228
cgcctgaaag cttatgatta cgc 23
<210> 229
<211> 21
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 229
ctgagcaggt ggcggaaaca t 21
<210> 230
<211> 25
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 230
ccatagcaag cagagtcttc atcag 25
<210> 231
<211> 23
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 231
ccctaccgga aggattacat ctc 23
<210> 232
<211> 24
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 232
ccagtaccag tacatcgctg tttc 24
<210> 233
<211> 27
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 233
gcttgagcaa attaaacaaa gcgaaag 27
<210> 234
<211> 22
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 234
cgcaaaaagg cagtcaatcc tg 22
<210> 235
<211> 23
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 235
gccattacca tgagcgataa cag 23
<210> 236
<211> 23
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 236
cggtcttatc ggcgataaac cag 23
<210> 237
<211> 19
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 237
gcagtggatg gtggacgat 19
<210> 238
<211> 19
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 238
cgttgccagt gctcgatca 19
<210> 239
<211> 22
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 239
ttcttaccgg acgaggacaa ga 22
<210> 240
<211> 23
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 240
tccaatcgca gatagaaggc aag 23
<210> 241
<211> 25
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 241
tttttggtga atcgcattct gactg 25
<210> 242
<211> 22
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 242
aaatccgctc cagacagatc ag 22
<210> 243
<211> 22
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 243
atgcttggtg ataacggcaa ttc 23
<210> 244
<211> 24
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 244
ttcgacacga gccaaaagat cgag 24
<210> 245
<211> 27
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 245
tgagaacaat ctgatgttgc tcgaata 27
<210> 246
<211> 23
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 246
tcatgatgca gatcgccatg tag 23
<210> 247
<211> 29
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 247
gattatagcc gatcaaatga tgagcaatg 29
<210> 248
<211> 20
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 248
tctgtcgcac ctgcttgatc 20
<210> 249
<211> 22
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 249
cgcctgtttt tcctgctcat tg 22
<210> 250
<211> 22
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 250
gcaacgatgt gagagagcat tc 22
<210> 251
<211> 22
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 251
cggtgttcgg cattctgaat ct 22
<210> 252
<211> 23
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 252
gcctcagcaa tatcgggata gag 23
<210> 253
<211> 27
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 253
gctacctgaa cgatatccaa ggatttc 27
<210> 254
<211> 22
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 254
ttccgcgaca ccgagaccaa ta 22
<210> 255
<211> 24
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 255
gtcgaacctt caaaagctga agtc 24
<210> 256
<211> 20
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 256
cgtcgcgact gcgaaatttc 20
<210> 257
<211> 22
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 257
acatttccgt gtcgccctta tt 22
<210> 258
<211> 25
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 258
gatgcttttc tgtgactggt gagta 25
<210> 259
<211> 24
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 259
gccgcataca ctattctcag aatg 24
<210> 260
<211> 27
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 260
cctccatcca gtctattaat tgttgcc 27
<210> 261
<211> 28
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 261
gcaaactatt aactggcgaa ctacttac 28
<210> 262
<211> 27
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 262
cacctatctc agcgatctgt ctatttc 27
<210> 263
<211> 23
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 263
cccagtgctg ttgttgtcat taa 23
<210> 264
<211> 28
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 264
ccctacttcg gtatctgtat tatcacgt 28
<210> 265
<211> 24
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 265
gtggttatgt tttggttccg tgaa 24
<210> 266
<211> 29
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 266
ccttcagata taaacgctaa aaaggcaaa 29
<210> 267
<211> 22
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 267
tgcattgcct ccaattccca ta 22
<210> 268
<211> 23
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 268
agctatattc gtgctcgtcg ttt 23
<210> 269
<211> 25
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 269
tttttcgtca acctcaactc aacag 25
<210> 270
<211> 30
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 270
cagcattctc gctatcaaaa taaagctaat 30
<210> 271
<211> 22
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 271
tagctcatgt tgacgcagga aa 22
<210> 272
<211> 27
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 272
atctctcgca gaaattagta ggattgc 27
<210> 273
<211> 30
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 273
ttttagcaga agtatatcgc tcgttatcag 30
<210> 274
<211> 29
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 274
tcatttcctt ctattaccac atcccattg 29
<210> 275
<211> 26
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 275
ccctaatatg tgtgtgatga gctgta 26
<210> 276
<211> 30
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 276
aaagttcaga cttctttctg tatgttgatg 30
<210> 277
<211> 26
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 277
agtgattaac agcgcattag agct 24
<210> 278
<211> 23
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 278
ctgtaggccg tgtacctaaa tgt 23
<210> 279
<211> 25
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 279
atgtgcttta ctaagtcatc gcgat 25
<210> 280
<211> 30
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 280
gacccacttt cacatttaag ttgtttttct 30
<210> 281
<211> 32
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 281
atgaatagaa taaaagttgc aatactgttt gg 32
<210> 282
<211> 25
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 282
ctgaatagca attgtcgttt tccca 25
<210> 283
<211> 26
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 283
tggtgtatgg aaaatgtgcg aaaaac 26
<210> 284
<211> 30
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 284
ttaacaaaaa caggataggt aaacgtagct 30
<210> 285
<211> 22
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 285
catgggctgc ttgtcatgaa ag 22
<210> 286
<211> 22
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 286
ttacgccaaa ggacgaacct ga 22
<210> 287
<211> 23
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 287
gcttacctac cctgtctttg tga 23
<210> 288
<211> 28
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 288
gtaatcatcg cattttctga gccttttt 28
<210> 289
<211> 22
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 289
gtatcttccg catccatcag ga 22
<210> 290
<211> 22
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 290
aatcaggctg tcaatcagtg ca 22
<210> 291
<211> 22
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 291
tctacccgtc caatggtctc at 22
<210> 292
<211> 24
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 292
cacaactaag ttgtcggtcg aatg 24
<210> 293
<211> 26
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 293
cttcggtcca gtagaactct tctatc 26
<210> 294
<211> 24
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 294
gcgcttttac aacattcgtt gtgt 24
<210> 295
<211> 27
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 295
gttattggtc tatttgaccg cgtctat 27
<210> 296
<211> 28
<212> DNA
<213>artificial sequence
<220>
<223>
<400> 296
gctgtatcaa tcaaaagcaa ctcatcac 28
Claims (10)
1. complete single stranded DNA, 296 single stranded DNAs shown in sequence 1-296 in sequence table are formed.
2. complete single stranded DNA described in claim 1 is detecting or is assisting the application in detection bacterium drug resistant gene；
Or, complete single stranded DNA described in claim 1 answering in the product of preparation detection or auxiliary detection bacterium drug resistant gene
With.
3. complete single stranded DNA described in claim 1 detect or assist detection tested bacteria whether carry drug resistant gene and/or
Carry the application in drug resistant gene type；
Or, whether complete single stranded DNA described in claim 1 carries drug resistant gene in preparation detection or auxiliary detection tested bacteria
And/or the application in the product of carrying drug resistant gene type.
4. complete single stranded DNA described in claim 1 is detecting or is assisting whether detection sample to be tested contains carrying drug resistant gene
Bacterium in application；
Or, whether complete single stranded DNA described in claim 1 contains carrying drug resistance in preparation detection or auxiliary detection sample to be tested
Application in the product of the bacterium of gene.
5. according to any application of claim 2-4, it is characterised in that: the drug resistant gene is following at least one: aac
(2’)-lb、aadA1、aadA3、aadB、aadD、adeb、ant(4’-llb)、aph3-la、armA、blaZ、catB3、catB6、
catB8、cmlA5、CTX-M-14、dfrA19、dfrG、E.coli_gyrA、E_coli_parC、IMP-1、KPC-1、mecA、
mecC、mcr-1、mtrC、mtrD、mtrE、mtrR、NDM-1、oprD、oqxA、oqxB、blaOXA-10、blaOXA-23、blaOXA-24、
blaOXA-1、blaOXA-51、blaOXA-58、blaOXA-66、blaOXA-69、blaOXA-82、PAO_gyrA_parC、patA、patB、pbp1a、
pbp2b、pbp2x、phoP、phoQ、pmrA、pmrB、qacA、qnrA1、qnrB4、rpsJ、sat-1、blaSHV-60、strA、
strB、sul1、blaTEM-12、tetA、tetD、tetM、tetR、vanA、vanB、VIM-2。
6. according to any application of claim 2-5, it is characterised in that: the bacterium is following at least one: Bao Man is not
Lever bacterium Acinetobacter baumannii, enterococcus faecalis Enterococcus faecalis, enterococcus faecium
Enterococcus faecium, Klebsiella Pneumoniae Klebsiella pneumoniae, the lung with beta- lactam resistant
The enterococcus of scorching klebsiella Klebsiella pneumoniae with beta-lactamase, drug resistance of vancomycin
Vancomycin-resistant Enterococci, pseudomonas aeruginosa Pseudomonas aeruginosa, golden yellow Portugal
Grape coccus Staphylococcus aureus, staphylococcus epidermis Staphylococcus epidermidis, haemolysis grape ball
Bacterium Staphylococcus haemolyticus, Human fetal cardiomyocytes Staphylococcus hominis, methicillin resistance
Staphylococcus aureus methicillin-resistant Staphylococcus aureus.
7. a kind of kit comprising complete single stranded DNA described in claim 1；
The function of the kit be it is following 1) or 2) or 3):
1) detect or assist detection bacterium drug resistant gene；
2) it detects or assists detection tested bacteria whether to carry drug resistant gene and/or carry the type of drug resistant gene；
3) detect or assist whether detection sample to be tested contains the bacterium for carrying drug resistant gene.
8. whether a kind of detection or auxiliary detection tested bacteria carry drug resistant gene and/or carry the side of the type of drug resistant gene
Method includes the following steps:
(1) genomic DNA for extracting tested bacteria, using complete single stranded DNA described in claim 1 to the genomic DNA
It is expanded, obtains amplified production；
(2) sequencing library is constructed using the amplified production based on Ampliseq technology；
(3) sequencing library is sequenced, obtains sequencing result；And the sequencing result and drug resistant gene sequence are carried out
It compares, determines whether tested bacteria carries drug resistant gene and carry the type of drug resistant gene.
9. kit according to claim 7 or method according to any one of claims 8, it is characterised in that: the drug resistant gene
Be following at least one: aac (2 ')-lb, aadA1, aadA3, aadB, aadD, adeb, ant (4 '-llb), aph3-la,
armA、blaZ、catB3、catB6、catB8、cmlA5、CTX-M-14、dfrA19、dfrG、E.coli_gyrA、E_coli_
parC、IMP-1、KPC-1、mecA、mecC、mcr-1、mtrC、mtrD、mtrE、mtrR、NDM-1、oprD、oqxA、oqxB、
blaOXA-10、blaOXA-23、blaOXA-24、blaOXA-1、blaOXA-51、blaOXA-58、blaOXA-66、blaOXA-69、blaOXA-82、PAO_
gyrA_parC、patA、patB、pbp1a、pbp2b、pbp2x、phoP、phoQ、pmrA、pmrB、qacA、qnrA1、qnrB4、
rpsJ、sat-1、blaSHV-60、strA、strB、sul1、blaTEM-12、tetA、tetD、tetM、tetR、vanA、vanB、VIM-
2。
10. kit according to claim 7 or method according to any one of claims 8, it is characterised in that: the bacterium is such as
Lower at least one: Acinetobacter bauamnnii Acinetobacter baumannii, enterococcus faecalis Enterococcus faecalis,
Enterococcus faecium Enterococcus faecium, Klebsiella Pneumoniae Klebsiella pneumoniae, band beta- lactams
The intestines of drug resistant Klebsiella Pneumoniae Klebsiella pneumoniae with beta-lactamase, drug resistance of vancomycin
It is coccus vancomycin-resistant Enterococci, pseudomonas aeruginosa Pseudomonas aeruginosa, golden yellow
Color staphylococcus Staphylococcus aureus, staphylococcus epidermis Staphylococcus epidermidis, haemolysis Portugal
Grape coccus Staphylococcus haemolyticus, Human fetal cardiomyocytes Staphylococcus hominis, methicillin are resistance to
The staphylococcus aureus methicillin-resistant Staphylococcus aureus of medicine.
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Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN110438247A (en) * | 2019-08-20 | 2019-11-12 | 中国医学科学院北京协和医院 | For detecting the nucleic acid reagent, kit, system and method for Escherichia coli and Klebsiella pneumoniae and its toxicity and drug resistance |
| CN111593131A (en) * | 2020-04-26 | 2020-08-28 | 领航基因科技（杭州）有限公司 | Primer probe system and kit capable of simultaneously detecting multiple drug-resistant genes |
| CN112481398A (en) * | 2020-12-21 | 2021-03-12 | 江苏汇先医药技术有限公司 | Real-time fluorescent quantitative PCR detection method and kit for multiple respiratory tract pathogenic bacteria |
| CN112941214A (en) * | 2021-03-29 | 2021-06-11 | 中国农业大学 | Primer group for gram-negative bacterium drug-resistant gene high-throughput amplicon sequencing and application |
| CN116875717A (en) * | 2023-09-07 | 2023-10-13 | 四川大学华西医院 | Primer, probe, method and kit for detecting klebsiella pneumoniae and carbapenem drug resistance genes |
Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US20150344977A1 (en) * | 2011-11-01 | 2015-12-03 | Pathogenica, Inc. | Method And System For Detection Of An Organism |
| CN105420371A (en) * | 2015-12-21 | 2016-03-23 | 张明 | Multi-pathogen and drug resistance gene detection method |
| CN108384782A (en) * | 2018-05-04 | 2018-08-10 | 中国人民解放军军事科学院军事医学研究院 | Detection causes the reagent set and kit of bloodstream infection pathogen |
-
2019
- 2019-02-18 CN CN201910119759.4A patent/CN109762915B/en active Active
Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US20150344977A1 (en) * | 2011-11-01 | 2015-12-03 | Pathogenica, Inc. | Method And System For Detection Of An Organism |
| CN105420371A (en) * | 2015-12-21 | 2016-03-23 | 张明 | Multi-pathogen and drug resistance gene detection method |
| CN108384782A (en) * | 2018-05-04 | 2018-08-10 | 中国人民解放军军事科学院军事医学研究院 | Detection causes the reagent set and kit of bloodstream infection pathogen |
Non-Patent Citations (1)
| Title |
|---|
| 李 倩等: "检测4 种血流感染病原体的靶向序列富集与测序相结合技术方法", 《军事医学》 * |
Cited By (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN110438247A (en) * | 2019-08-20 | 2019-11-12 | 中国医学科学院北京协和医院 | For detecting the nucleic acid reagent, kit, system and method for Escherichia coli and Klebsiella pneumoniae and its toxicity and drug resistance |
| CN111593131A (en) * | 2020-04-26 | 2020-08-28 | 领航基因科技（杭州）有限公司 | Primer probe system and kit capable of simultaneously detecting multiple drug-resistant genes |
| CN112481398A (en) * | 2020-12-21 | 2021-03-12 | 江苏汇先医药技术有限公司 | Real-time fluorescent quantitative PCR detection method and kit for multiple respiratory tract pathogenic bacteria |
| CN112481398B (en) * | 2020-12-21 | 2024-03-08 | 江苏汇先医药技术有限公司 | Real-time fluorescent quantitative PCR detection method and kit for various respiratory tract pathogens |
| CN112941214A (en) * | 2021-03-29 | 2021-06-11 | 中国农业大学 | Primer group for gram-negative bacterium drug-resistant gene high-throughput amplicon sequencing and application |
| CN116875717A (en) * | 2023-09-07 | 2023-10-13 | 四川大学华西医院 | Primer, probe, method and kit for detecting klebsiella pneumoniae and carbapenem drug resistance genes |
| CN116875717B (en) * | 2023-09-07 | 2023-11-21 | 四川大学华西医院 | Primer, probe, method and kit for detecting klebsiella pneumoniae and carbapenem drug resistance genes |
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