CN108949745A - A kind of unicellular sequencing library construction method of high throughput - Google Patents
A kind of unicellular sequencing library construction method of high throughput Download PDFInfo
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- CN108949745A CN108949745A CN201810796145.5A CN201810796145A CN108949745A CN 108949745 A CN108949745 A CN 108949745A CN 201810796145 A CN201810796145 A CN 201810796145A CN 108949745 A CN108949745 A CN 108949745A
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N15/00—Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
- C12N15/09—Recombinant DNA-technology
- C12N15/10—Processes for the isolation, preparation or purification of DNA or RNA
- C12N15/1034—Isolating an individual clone by screening libraries
- C12N15/1093—General methods of preparing gene libraries, not provided for in other subgroups
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
- C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
- C12Q1/68—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving nucleic acids
- C12Q1/6806—Preparing nucleic acids for analysis, e.g. for polymerase chain reaction [PCR] assay
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
- C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
- C12Q1/68—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving nucleic acids
- C12Q1/6869—Methods for sequencing
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- C—CHEMISTRY; METALLURGY
- C40—COMBINATORIAL TECHNOLOGY
- C40B—COMBINATORIAL CHEMISTRY; LIBRARIES, e.g. CHEMICAL LIBRARIES
- C40B50/00—Methods of creating libraries, e.g. combinatorial synthesis
- C40B50/06—Biochemical methods, e.g. using enzymes or whole viable microorganisms
Abstract
The invention discloses a kind of unicellular sequencing library construction methods of high throughput, include the following steps, S1, the first substance will be contacted from the single celled nucleic acid and include the polymerase of strand-displacement activity, contact betides under conditions of 4 DEG C to about 25 DEG C temperature ranges, primer and the Nucleic acids anneal, and the primer is extended through digestion, the mixture is placed under conditions of so that two ends of full amplicon is annealed with one another, so that full amplicon cannot anneal with the first substance or the second substance, wherein the second substance is rich in the G or C of 30%-40%；And the first substance or the second substance is made to anneal and extend with half amplicon linearly generated remaining in mixture, wherein the annealing and extending in nucleic acid and will not further occur in the case where unwinding, thus generate the full amplicon A linearly generated.The present invention is ingenious in design, and method is reasonable, high-throughput sequencing library, and effect is good, and accuracy is high, is suitble to promote.
Description
Technical field
The present invention relates to unicellular sequencing library constructing technology field more particularly to a kind of unicellular sequencing libraries of high throughput
Construction method.
Background technique
The unicellular genomics technologies for being not based on culture are successfully realized, and are applied to various microbial ecologies and are ground
Study carefully.And the difficult point analyzed microbial cell is, most of microbe, such as bacterium and archeabacterial cell, is difficult to
The epicyte structure effectively cracked；RNA half-life short, the stability of prokaryotic micro-organisms are low；In addition, cell size is much smaller than the food in one's mouth
Newborn zooblast (10~30 pik), therefore the total concentration of intracellular RNA molecule is very low, about each cell 4~10 is luxuriant and rich with fragrance
Gram, this RNA concentration mentions for RNA extracting and analysis far below the RNA concentration in eukaryotic cells and has given birth to many challenges.Though
So the single celled full transcriptome analysis technology of eucaryote is directed to it has been established that but being directed to the single celled full transcription of prokaryotic micro-organisms
The technology of group analysis could not enough be completely set up always.
Biology can be divided into unicellular organism and multicellular organism according to the cell number of composition.Unicellular organism is only by list
A cell composition, and often aggregation becomes cell colony.Earliest biology is about before 3,500,000,000 years away from the present to 41 on the earth
It was formed before 100000000 years, prokaryotes are the biologies of most original, such as bacterium and blue-green alge and occurred in warm water.Unicellular life
Object includes all archeobacterias and eubacteria and many protists.There are many animals according to old classification, plant and fungi are more
It is multicellular organism.Amoeba can be regarded as unicellular animal, its some types but can be regarded as Acarasiales, such as eye worm of the flagellate with flagellum
Sometimes it is classified as unicellular alga either unicellular animal.
Through retrieving, Patent No. CN201310481921.X proposes a kind of microbial single-cell based on two generation sequencing technologies
Transcriptome analysis method, the separation of microbial single-cell: fresh microbial cell culture sample is collected by centrifugation at normal temperature,
It is suspended in RNA protective agent RNALater solution immediately；It is carried out under inverted microscope using micro-injection system single celled
Separation is chosen, and isolated microbial single-cell is resuspended in RNALater solution, if total serum IgE extracts, the line of total serum IgE
Property amplification, and the building for RNA sequencing library a variety of commercial reagents boxes application effect comparative evaluation, obtain a set of
Complete techniqueflow finally realizes and carries out the full transcriptome analysis based on two generation sequencing technologies to microbial single-cell, only
The analysis between the gene expression difference different microorganisms cell individual may be implemented, can also realize can not to extreme environment
The microorganism of culture carries out the physiological function analysis of individual cell level；This application file is when constructing sequencing library, false positive rate
It cannot be reduced and simply increasing sequencing depth, amplification mistake independently generates, and can be dilute in linear amplified production
It releases, for this purpose, the present invention proposes a kind of unicellular sequencing library construction method of high throughput.
Summary of the invention
The purpose of the present invention is to solve disadvantages existing in the prior art, and a kind of unicellular survey of high throughput proposed
Preface base construction method.
To achieve the goals above, present invention employs following technical solutions:
A kind of unicellular sequencing library construction method of high throughput, includes the following steps,
S1 will contact the first substance from the single celled nucleic acid and comprising the polymerase of strand-displacement activity, contact hair
It is born under conditions of 4 DEG C to about 25 DEG C temperature ranges, primer and the Nucleic acids anneal, and the primer is prolonged by digestion
It stretches, wherein first substance includes the G rich in the 30%-40% or C rich in 30%-40%；With include restriction enzyme core
Sour enzyme site, to generate the nucleic acid-templated mixture comprising primer annealing；To the nucleic acid-templated carry out two-wheeled of primer annealing
Or more wheel extend, unwinding and annealing steps, thus generate nucleic acid-templated, half amplicon that is linearly generating and non-linear generation
The mixture of full amplicon；The mixture is placed under conditions of so that two ends of full amplicon is annealed with one another, from
And generate the full amplicon of cyclization；The full amplicon of the cyclization is contacted into restriction endonuclease, so that full amplification
Son cannot anneal with the first substance or the second substance, wherein the second substance is rich in the G or C of 30%-40%；And make the first substance
Or second substance anneal and extend with half amplicon linearly generated remaining in mixture, wherein the annealing and extending in core
Acid will not occur in the case where further unwinding, thus generate the full amplicon A linearly generated；
Full amplicon A in S2, the S1 is used as sequencing library.
Preferably, second substance is rich in 35% G or C.
Preferably, the enzyme that the digestion uses is including but not limited to any of them or several: AluI, Taq α I,
SphI、MspI、BamHI、ApeKI、BanII、HpaII、HpyCH4V、BstNI、HaeIII、HpyCH4III、BglII、BssSI
And KpnI.
Preferably, first substance and the second substance include the enzyme of strand-displacement activity.
Preferably, the enzyme of the strand-displacement activity is selected from but not limited to any of them or several: klenow segment,
Klenow segment, bst archaeal dna polymerase, vent archaeal dna polymerase, vent archaeal dna polymerase, Phi 29DNA polymerase, deep
Vent archaeal dna polymerase, deepvent archaeal dna polymerase.
Preferably, first structure of matter are as follows: the sequence of the 5 '-specific recognition connectors-island random sequence-CpG identification
The structure of sequence -3 ', the second substance are as follows: the sequence of 5 '-specific recognition connectors-island random sequence-CpG identifies sequence -3 '；Or
The structure of the first substance of person are as follows: sequence-molecular label of 5 '-specific recognition connectors-island random sequence-CpG identifies sequence-
3 ', the structure of the second substance are as follows: sequence-molecular label of 5 '-specific recognition connectors-island random sequence-CpG identifies sequence-
3’。
Preferably, described unicellular by microflow control technique, microflow control technique can utilize gravity centrifugation, hydrodynamics, electricity
Field force captures cell, can also be sorted by hydrodynamics using cell size, cell surface marker, while can be with
Integrator cell culture, cell capture sorting, cell cracking and subsequent test and analyze are based on one, can intuitively distinguish that purpose is thin
Born of the same parents, realize high-throughput unicellular separation, automate it is integrated.
Preferably, in the S1, the first substance will be contacted from the single celled nucleic acid and include strand-displacement activity
Polymerase, under conditions of contact betides 20 DEG C of temperature ranges, primer and the Nucleic acids anneal, and the primer is obtained by digestion
To extension, wherein first substance includes the G rich in the 35% or C rich in 35%.
Preferably, the length of the random sequence in first substance is 4~20nt, the stochastic ordering in second substance
The length of column is 4~20nt.
Preferably, sequencing library is illustrated using the NuGen Encore kit of NuGen company of the U.S. according to kit
The building of book progress RNA sequencing library.
Compared with prior art, the beneficial effects of the present invention are: betiding 4 DEG C to about 25 DEG C temperature ranges by contact
Under the conditions of, primer and the Nucleic acids anneal, and the primer is extended through digestion, wherein first substance includes being rich in
The G of 30%-40% or C rich in 30%-40% is made using the activated centre region of these small-molecule substances stimulation ligase
It obtains it and connects increased activity, improve connector joint efficiency, so that more DNA plerosis segments are successfully connected joint sequence, into
And it is expanded by the primer complementary with joint sequence, and then obtained the more high-flux sequences of purpose amplified fragments content
Library；To improve micro initial amount sample；The G or C of 30%-40% are rich in by the second substance；And make the first substance
Or second substance anneal and extend with half amplicon linearly generated remaining in mixture, wherein the annealing and extending in core
Acid will not occur in the case where further unwinding, thus generate the full amplicon A linearly generated, measure the full amplification linearly generated
Son one or more of them specific nucleotide compared with standard items whether there is, and false positive rate is sequenced depth by this method and subtracts
Few, the present invention is ingenious in design, and method is reasonable, high-throughput sequencing library, and effect is good, and accuracy is high, is suitble to promote.
Specific embodiment
The technical scheme in the embodiments of the invention will be clearly and completely described below, it is clear that described implementation
Example is only a part of the embodiment of the present invention, instead of all the embodiments.
Embodiment one
A kind of unicellular sequencing library construction method of high throughput, includes the following steps,
S1 will contact the first substance from the single celled nucleic acid and comprising the polymerase of strand-displacement activity, contact hair
It is born under conditions of 4 DEG C of temperature, primer and the Nucleic acids anneal, and the primer is extended through digestion, wherein described the
One substance includes the G rich in the 30% or C rich in 30%；With comprising restriction endonuclease site, to generate comprising drawing
The nucleic acid-templated mixture of object annealing；The extension of nucleic acid-templated carry out two-wheeled to primer annealing or more wheel, unwinding and annealing
Thus step generates the mixture of the full amplicon of nucleic acid-templated, half amplicon that is linearly generating and non-linear generation；It will be described
Mixture is placed under conditions of so that two ends of full amplicon is annealed with one another, to generate the full amplicon of cyclization；It will
The full amplicon of the cyclization contacts restriction endonuclease, so that full amplicon cannot be with the first substance or the second object
Matter annealing, wherein the second substance is rich in 30% G or C；And make remaining line in the first substance or the second substance and mixture
Property half amplicon that generates anneal and extends, wherein described anneal and extend in nucleic acid and will not issue the case where further unwinding
It is raw, thus generate the full amplicon A linearly generated；
Full amplicon A in S2, the S1 is used as sequencing library；
The enzyme that the digestion uses is including but not limited to any of them or several: AluI, Taq α I, SphI, MspI,
BamHI, ApeKI, BanII, HpaII, HpyCH4V, BstNI, HaeIII, HpyCH4III, BglII, BssSI and KpnI.
Embodiment two
A kind of unicellular sequencing library construction method of high throughput, includes the following steps,
S1 will contact the first substance from the single celled nucleic acid and comprising the polymerase of strand-displacement activity, contact hair
It is born under conditions of 20 DEG C of temperature, primer and the Nucleic acids anneal, and the primer is extended through digestion, wherein described the
One substance includes the G rich in the 35% or C rich in 35%；With comprising restriction endonuclease site, to generate comprising drawing
The nucleic acid-templated mixture of object annealing；The extension of nucleic acid-templated carry out two-wheeled to primer annealing or more wheel, unwinding and annealing
Thus step generates the mixture of the full amplicon of nucleic acid-templated, half amplicon that is linearly generating and non-linear generation；It will be described
Mixture is placed under conditions of so that two ends of full amplicon is annealed with one another, to generate the full amplicon of cyclization；It will
The full amplicon of the cyclization contacts restriction endonuclease, so that full amplicon cannot be with the first substance or the second object
Matter annealing, wherein the second substance is rich in 35% G or C；And make remaining line in the first substance or the second substance and mixture
Property half amplicon that generates anneal and extends, wherein described anneal and extend in nucleic acid and will not issue the case where further unwinding
It is raw, thus generate the full amplicon A linearly generated；
Full amplicon A in S2, the S1 is used as sequencing library；
The enzyme of the strand-displacement activity is selected from but not limited to any of them or several: klenow segment, klenow piece
Section, bst archaeal dna polymerase, vent archaeal dna polymerase, vent archaeal dna polymerase, Phi 29DNA polymerase, deep vent DNA
Polymerase, deep vent archaeal dna polymerase.
Embodiment three
A kind of unicellular sequencing library construction method of high throughput, includes the following steps,
S1 will contact the first substance from the single celled nucleic acid and comprising the polymerase of strand-displacement activity, contact hair
It is born under conditions of 25 DEG C of temperature, primer and the Nucleic acids anneal, and the primer is extended through digestion, wherein described the
One substance includes the G rich in the 40% or C rich in 40%；With comprising restriction endonuclease site, to generate comprising drawing
The nucleic acid-templated mixture of object annealing；The extension of nucleic acid-templated carry out two-wheeled to primer annealing or more wheel, unwinding and annealing
Thus step generates the mixture of the full amplicon of nucleic acid-templated, half amplicon that is linearly generating and non-linear generation；It will be described
Mixture is placed under conditions of so that two ends of full amplicon is annealed with one another, to generate the full amplicon of cyclization；It will
The full amplicon of the cyclization contacts restriction endonuclease, so that full amplicon cannot be with the first substance or the second object
Matter annealing, wherein the second substance is rich in 40% G or C；And make remaining line in the first substance or the second substance and mixture
Property half amplicon that generates anneal and extends, wherein described anneal and extend in nucleic acid and will not issue the case where further unwinding
It is raw, thus generate the full amplicon A linearly generated；
Full amplicon A in S2, the S1 is used as sequencing library；
First structure of matter are as follows: the sequence of 5 '-specific recognition connectors-island random sequence-CpG identifies sequence -3 ',
The structure of second substance are as follows: the sequence of 5 '-specific recognition connectors-island random sequence-CpG identifies sequence -3 '；Or first object
The structure of matter are as follows: sequence-molecular label of 5 '-specific recognition connectors-island random sequence-CpG identifies sequence -3 ', the second object
The structure of matter are as follows: sequence-molecular label of 5 '-specific recognition connectors-island random sequence-CpG identifies sequence -3 '；
Described unicellular by microflow control technique, microflow control technique can be caught using gravity centrifugation, hydrodynamics, electric field force
Cell is obtained, can also be sorted by hydrodynamics using cell size, cell surface marker, while can be with integrator cell
Culture, cell capture sort, cell cracking and subsequent test and analyze are based on one, can intuitively distinguish aim cell, realize height
Flux it is unicellular separation, automate it is integrated.
In the use of the present invention, under conditions of betiding 4 DEG C to about 25 DEG C temperature ranges by contact, primer and the core
Acid annealing, and the primer is extended through digestion, wherein first substance includes being rich in the G of 30%-40% or being rich in
The C of 30%-40% is mentioned using the activated centre region of these small-molecule substances stimulation ligase so that it connects increased activity
High connector joint efficiency, so that more DNA plerosis segments are successfully connected joint sequence, and then by complementary with joint sequence
Primer expanded, and then obtained the more high-throughput sequencing libraries of purpose amplified fragments content；It is micro to improve
Initial amount sample；The G or C of 30%-40% are rich in by the second substance；And make in the first substance or the second substance and mixture
Remaining half amplicon linearly generated is annealed and is extended, wherein it is described annealing and extend in nucleic acid will not further unwinding feelings
Occur under condition, thus generate the full amplicon A linearly generated, measures the full amplicon linearly generated wherein one compared with standard items
Kind or a variety of specific nucleotides whether there is, and false positive rate is sequenced depth by this method and reduces, and the present invention is ingenious in design, side
Method is reasonable, high-throughput sequencing library, and effect is good, and accuracy is high, is suitble to promote.
The foregoing is only a preferred embodiment of the present invention, but scope of protection of the present invention is not limited thereto,
Anyone skilled in the art in the technical scope disclosed by the present invention, according to the technique and scheme of the present invention and its
Inventive concept is subject to equivalent substitution or change, should be covered by the protection scope of the present invention.
Claims (10)
1. a kind of unicellular sequencing library construction method of high throughput, which is characterized in that include the following steps,
S1 will contact the first substance from the single celled nucleic acid and comprising the polymerase of strand-displacement activity, contact betide 4
DEG C under conditions of about 25 DEG C of temperature ranges, primer and the Nucleic acids anneal, and the primer is extended through digestion, wherein
First substance includes the G rich in the 30%-40% or C rich in 30%-40%；With include restriction endonuclease position
Point, to generate the nucleic acid-templated mixture comprising primer annealing；To the nucleic acid-templated carry out two-wheeled or more of primer annealing
Wheel extension, unwinding and annealing steps, thus generate the full amplification of nucleic acid-templated half amplicon that is linearly generating and non-linear generation
The mixture of son；The mixture is placed under conditions of so that two ends of full amplicon is annealed with one another, to generate
The full amplicon of cyclization；The full amplicon of the cyclization is contacted into restriction endonuclease, so that full amplicon cannot
It anneals with the first substance or the second substance, wherein the second substance is rich in the G or C of 30%-40%；And make the first substance or second
Substance is annealed and is extended with half amplicon linearly generated remaining in mixture, wherein it is described annealing and extend in nucleic acid will not
Occur in the case where further unwinding, thus generates the full amplicon A linearly generated；
Full amplicon A in S2, the S1 is used as sequencing library.
2. the unicellular sequencing library construction method of a kind of high throughput according to claim 1, which is characterized in that described second
Substance is rich in 35% G or C.
3. the unicellular sequencing library construction method of a kind of high throughput according to claim 1, which is characterized in that the digestion
The enzyme of use is including but not limited to any of them or several: AluI, Taq α I, SphI, MspI, BamHI, ApeKI,
BanII, HpaII, HpyCH4V, BstNI, HaeIII, HpyCH4III, BglII, BssSI and KpnI.
4. the unicellular sequencing library construction method of a kind of high throughput according to claim 1, which is characterized in that described first
Substance and the second substance include the enzyme of strand-displacement activity.
5. the unicellular sequencing library construction method of a kind of high throughput according to claim 4, which is characterized in that the chain is set
Active enzyme is changed selected from but not limited to any of them or several: klenow segment, klenow segment, bst archaeal dna polymerase,
Vent archaeal dna polymerase, vent archaeal dna polymerase, Phi 29DNA polymerase, deep vent archaeal dna polymerase, deepvent
Archaeal dna polymerase.
6. the unicellular sequencing library construction method of a kind of high throughput according to claim 1, which is characterized in that described first
The structure of matter are as follows: the sequence of the 5 '-specific recognition connectors-island random sequence-CpG identification sequence -3 ', the second substance structure
Are as follows: the sequence of 5 '-specific recognition connectors-island random sequence-CpG identifies sequence -3 '；Or first substance structure are as follows: 5 '-
Sequence-molecular label of the specific recognition connector-island random sequence-CpG identification sequence -3 ', the second substance structure are as follows: 5 ' -
Sequence-molecular label of specific recognition connector-island random sequence-CpG identifies sequence -3 '.
7. the unicellular sequencing library construction method of a kind of high throughput according to claim 1, which is characterized in that described slender
Born of the same parents can be captured cell using gravity centrifugation, hydrodynamics, electric field force, can also be led to by microflow control technique, microflow control technique
It crosses hydrodynamics to be sorted using cell size, cell surface marker, while can be with integrator cell culture, cell capture point
Choosing, cell cracking and it is subsequent test and analyze based on one, can intuitively distinguish aim cell, realize high-throughput unicellular separation,
It automates integrated.
8. the unicellular sequencing library construction method of a kind of high throughput according to claim 1, which is characterized in that the S1
In, the first substance will be contacted from the single celled nucleic acid and comprising the polymerase of strand-displacement activity, contact betides 20 DEG C
Under conditions of temperature range, primer and the Nucleic acids anneal, and the primer is extended through digestion, wherein first object
Matter includes the G rich in the 35% or C rich in 35%.
9. the unicellular sequencing library construction method of a kind of high throughput according to claim 1, which is characterized in that described first
The length of random sequence in substance is 4~20nt, and the length of the random sequence in second substance is 4~20nt.
10. the unicellular sequencing library construction method of a kind of high throughput according to claim 1, which is characterized in that sequencing text
Library is carried out according to kit specification the structure of RNA sequencing library using the NuGen Encore kit of NuGen company of the U.S.
It builds.
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN110453000A (en) * | 2019-08-15 | 2019-11-15 | 深圳谱元科技有限公司 | A kind of two generations sequencing banking process for identifying the typing of bacteria and resistant type of helicobacter pylori in sample |
| CN111363795A (en) * | 2018-12-26 | 2020-07-03 | 清华大学 | Single cell whole genome sequencing method |
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| CN106661620A (en) * | 2014-05-06 | 2017-05-10 | 贝勒医学院 | Methods of linearly amplifying whole genome of a single cell |
| CN107475779A (en) * | 2017-09-22 | 2017-12-15 | 上海美吉医学检验有限公司 | Library method for building up and its application suitable for unicellular RRBS sequencings |
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2018
- 2018-07-19 CN CN201810796145.5A patent/CN108949745A/en active Pending
Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN106661620A (en) * | 2014-05-06 | 2017-05-10 | 贝勒医学院 | Methods of linearly amplifying whole genome of a single cell |
| CN107475779A (en) * | 2017-09-22 | 2017-12-15 | 上海美吉医学检验有限公司 | Library method for building up and its application suitable for unicellular RRBS sequencings |
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN111363795A (en) * | 2018-12-26 | 2020-07-03 | 清华大学 | Single cell whole genome sequencing method |
| CN110453000A (en) * | 2019-08-15 | 2019-11-15 | 深圳谱元科技有限公司 | A kind of two generations sequencing banking process for identifying the typing of bacteria and resistant type of helicobacter pylori in sample |
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