CN103484458A - Oligonucleotide sequence containing universal base and application thereof to DNA hybridization analysis - Google Patents
Oligonucleotide sequence containing universal base and application thereof to DNA hybridization analysis Download PDFInfo
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- CN103484458A CN103484458A CN201310469720.8A CN201310469720A CN103484458A CN 103484458 A CN103484458 A CN 103484458A CN 201310469720 A CN201310469720 A CN 201310469720A CN 103484458 A CN103484458 A CN 103484458A
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Abstract
The invention discloses an oligonucleotide sequence containing universal bases and application thereof to DNA hybridization analysis. The oligonucleotide sequence comprises basic bases and universal bases. The basic bases are completely complementary to a target sequence, and comprise basic bases corresponding to SNV (single nucleotide variation) loci of the target sequence; the universal bases can not be corresponding to SNV loci of the target sequence and are located at one or two sides of the SNV loci of the target sequence; the basic bases comprise A, G, C and T; and the universal bases are nucleosides capable of forming hydrogen bond pairing with A, G, C and T. The most outstanding advantage of the invention is that effective analysis of single base mismatching in the DNA sequence is realized.
Description
Technical field
The present invention relates to the oligonucleotide field, be specifically related to a kind of containing universal base oligonucleotide sequence and the application in the DNA hybridization analysis thereof.
Background technology
Along with completing that human genome is evolved; to the research of human genome from measuring DNA sequence dna, the research aspect of explaining all genetic information transfers to the aspect of biological function being studied from the molecule integral level and comes; thereby more the good utilisation gene information is carried out medical diagnosis on disease, treatment and prevention, reach the purpose of understanding life, protection life.The research mutant dna sequence relevant to genetic phenotype is one of theme of genetics research, the variation great majority of human genomic sequence are change (the single nucleotide variation of mononucleotide, SNV) (comprise single nucleotide polymorphism (single nucleotide polymorphism, SNP) and point mutation (Mutation)), and in different crowds, the frequency distribution of SNV is variant, and these differences can represent the hereditary difference between a certain race or crowd.Therefore, research SNV contributes to explain individual phenotypic difference, different groups and individual to disease, particularly to the susceptibility of complex disease, and to the tolerance of various medicines with to the reaction of environmental factor.The characteristic of SNV self has determined that it is more suitable in the research of the genetic dissection to complex character and disease relationship and the aspects such as gene recognition based on colony than other polymorphism marks.SNV can be used as genetic marker and carry out linkage analysis, the location diseases predisposing gene, and also the ratio of precision microsatellite marker of its location is much meticulous, can be directly used in and instruct the tumor susceptibility gene clone.Erect image many investigators think: the understanding for these molecule markers will make the discovery of the disease related genes such as asthma, diabetes, schizophrenia and cancer become simply, and SNV also can be used as the association analysis of disease simultaneously.Be applied at present the classifying method in SNV site mainly based on four kinds of ultimate principles: i.e. endonuclease digestion technology, allele-specific hybridization, primer extension and oligonucleotide interconnection technique.Prerequisite based on the enzyme incision technology detection method is that restriction endonuclease recognition sequence need be contained in the both sides in SNV site to be detected, so the SNV site does not cause the sequence of the change of restriction enzyme site can't analyze.Its excess-three kind technology is without any restrictions to analytical sequence on principle, and can carry out high throughput analysis in conjunction with biochip technology.
Gene chip (gene chip), also being DNA chip, DNA microarray (DNA microarray), oligonucleotide arrays (Oligonucleotide array), is to fix in an orderly manner, to high-density different target sequences or oligonucleotide fragment on the carriers such as slide.At present, gene chip has been widely used in and has found disease related gene, set up medical diagnosis on disease index and fundamental biological knowledge and medical research field.Most important application of gene chip is to analyze and detect the sequence difference between different organism genomes, and these differences mainly comprise sudden change and single nucleotide polymorphism.Although gene chip goes out many different SNV analytical technologies in conjunction with other derives from technology: as the hybridization typing method based on the DNA chip; Single-basic extension based on the DNA chip-label typing method; Oligonucleotide based on the DNA chip connects typing method; Classifying method based on DNA chip and multiplex PCR amplification etc.But in fact the core essence of these technology take the solid-phase hybridization technology of large-scale integrated is prerequisite, each analytical sequence needs, with probe sequence, specific hybridization is arranged, so the hybridization of specific probe sequence is a very important index of these analytical procedure accuracys.
Being comprised of base A, G, C, T of specific probe sequence in existing gene chip, according to the sequence of target object, design according to specific Tm value.Probe sequence design can modulation mode comprise length and the kind of sequence, by one section of selected target sequence or several sections sequences, design the one or more of probe sequences with the target sequence coupling, finally, by take hybridization as the signal detection of essential analytical technology enforcement to sample, draw the genetic information (information of gene order and genetic expression) of target sample.Clearly, this regional as hybridization by selected certain specific fragment, can only rely on this condition of change base number to determine that the probe sequence design of Tm value is limited by very large undoubtedly.On the one hand, for number, for more DNA chip, all sequences is designed to as far as possible to close Tm value itself is exactly a difficulty; On the other hand, even on the DNA chip, the Tm value of all sequences design is close, but can effectively distinguish single base mismatch also has problems, because the differentiation of single base mismatch often needs more exacting terms to realize, as reduced under the prerequisite of non-specific sequence hybridization at the optimization hybridization temperature, also need the intensity (being washed) of cleaning by strengthening or adopt the means such as electric field electrophoresis under at specified temp, remove the probe sequence of non-specific hybridization.
Summary of the invention
The purpose of this invention is to provide a kind of containing the universal base oligonucleotide sequence, by introduce universal base in oligonucleotide sequence, utilize it can form with four basic bases the characteristics of weak hydrogen bond pairing, effectively distinguish the single base mismatch sequence, solved and can not effectively distinguish the problem of single base mismatch in the prior art.The present invention also provides this containing the application of universal base oligonucleotide sequence in the DNA hybridization analysis.
For addressing the above problem, the present invention by the following technical solutions:
A kind of containing the universal base oligonucleotide sequence, the described universal base oligonucleotide sequence that contains comprises basic base and universal base, and described basic base and target sequence complete complementary, comprising the basic base corresponding with target sequence SNV site; Described universal base can not corresponding target sequence SNV site, is positioned at the one or both sides of the basic base corresponding with target sequence SNV site; Described basic base is A, G, C, T, and described universal base is can form the nucleosides of hydrogen bond pairing with A, G, C, T.
Described basic base is 11-50; Described universal base is 1-8.
Described basic base is 13-18; Described universal base is 2-4.
Described universal base is hypoxanthine deoxyriboside, 2 ' ribodesose-3 ' nitro-pyrrole or 2 ' ribodesose-5 ' nitroindoline.
The described universal base oligonucleotide sequence that contains comprises marker or does not comprise marker.
Above-mentioned containing the application of universal base oligonucleotide sequence in the DNA hybridization analysis.
The above-mentioned application in the DNA hybridization hybrid chip is analyzed containing the universal base oligonucleotide sequence.
Principle of the present invention: have the general nucleosides of a class to realize pairing by forming hydrogen bond with A, G, C, tetra-basic bases of T in nucleic acid, this class nucleosides is called as universal base, its kind is numerous, as hypoxanthine deoxyriboside (Hypoxanthine deoxyriboside), 2 ' ribodesose-3 ' nitro-pyrrole etc.The present invention, exactly by introduce universal base in sequence, utilizes it to form the characteristics of weak hydrogen bond pairing with four basic bases, effectively distinguishes the single base mismatch sequence.Introduce one or several universal base in sequence, the concentration kinetic factor of this sequence can be strengthened, and Thermodynamics can be weakened.On the one hand, for the sequence of mating fully, although it is low with respect to forming double-stranded stability by normal alkali basic sequence and matching sequence hybridization fully in the past to form double-stranded stability containing universal base sequence and complete matching sequence hybridization, but still can ensure and effective enforcement of hybridization maintain its normal biochemical function; And for the single base mismatch sequence, except base mismatch, increased again and formed the lower universal base of double-spiral structure stability, so this stability is just low more than original normal alkali basic sequence, therefore, under a maneuverable condition, containing universal base sequence and this double-spiral structure of single base mismatch sequence, just may not form, perhaps destroyed, so just can effectively be distinguished the single base mismatch sequence.
Beneficial effect of the present invention:
1. great advantage of the present invention is to have realized effective analysis of single base mismatch in the DNA sequence dna.Ensureing that sequence and complete matching sequence maintain under effective hybridization and normal biochemical function condition, the hybridization stability of decrease single base mismatch sequence, improve the separating capacity of single base mismatch, increase the accuracy that in nucleotide sequence, SNV analyzes, for SNV in the analysis of nucleic acids sequence provides a kind of accurate, reproducible, quick and cheap typing method.
2. the present invention can adopt traditional DNA chemical synthesis to prepare containing the universal base oligonucleotide sequence, and its using method and operation steps are all carried out according to popular molecular biology method, easily in existing technical enforcement.
The accompanying drawing explanation
Fig. 1 be conventional probe and containing the universal base probe respectively with the crossing pattern of fixing DNA sequence dna
The hybridization of a, DNA sequence dna and complete complementary conventional probe sequence; The hybridization of b, DNA sequence dna and single base mismatch conventional probe sequence; C, DNA sequence dna and complete complementary are containing the hybridization of universal base hypoxanthine deoxyriboside probe sequence; D, DNA sequence dna and single base mismatch are containing the hybridization of universal base hypoxanthine deoxyriboside probe sequence.
Fig. 2 be fixing conventional probe and containing the universal base probe respectively with the crossing pattern of marker DNA sequence
The complete complementary hybridization of a, conventional probe sequence and marker DNA sequence; B, conventional probe sequence and marker DNA sequence have the hybridization of a single base mismatch; C, containing the complete complementary of universal base hypoxanthine deoxyriboside probe sequence and marker DNA sequence, hybridize; D, the hybridization of a single base mismatch is arranged containing universal base hypoxanthine deoxyriboside probe sequence and marker DNA sequence.
Fig. 3 is the DNA chip analysis Cy3 scintigram as a result of 80 sample line plastochondria mutational site C3206T.
Fig. 4 is the DNA chip analysis Cy5 scintigram as a result of 80 sample line plastochondria mutational site C3206T.
Fig. 5 is the DNA chip analysis Cy3 scintigram as a result in mankind rs988748 site
A, wild-type homozygote Cy3 scintigram; B, heterozygote Cy3 scintigram; C, saltant type homozygote Cy3 scintigram.
Embodiment
Below in conjunction with embodiment and accompanying drawing, the present invention is done further and explains.The following example is only for the present invention is described, but is not used for limiting practical range of the present invention.
A kind of containing the universal base oligonucleotide sequence, the described universal base oligonucleotide sequence that contains comprises basic base and universal base, and described basic base and target sequence complete complementary, comprising the basic base corresponding with target sequence SNV site; Described universal base can not corresponding target sequence SNV site, is positioned at the one or both sides of the basic base corresponding with target sequence SNV site, can guarantee like this specificity of sequence hybridization; Described basic base is A, G, C, T; Described universal base is can form the nucleosides of hydrogen bond pairing with A, G, C, T, including, but not limited to hypoxanthine deoxyriboside, 2 ' ribodesose-3 ' nitro-pyrrole, 2 ' ribodesose-5 ' nitroindoline etc.
Described is to design according to concrete evaluating objects sequence containing the basic base in the universal base oligonucleotide sequence and sequence and the number of universal base, and finally be determined by experiment.Tm value containing the universal base oligonucleotide sequence is relevant to the hybridization temperature of experiment, generally higher than 1～5 ℃ of hybridization temperature.The number of basic base and universal base should be as far as possible little, is generally 11-50, and wherein the number of universal base is 1-8; Be preferably 13-18, wherein the number of universal base is 2-4.Should can complete normal hybridization with the target complementary sequence containing the universal base oligonucleotide sequence, and maintain the extension (synthesizing) of hybridization sequences, the biochemical functions such as connection, the analysis for the DNA chip to nucleotide sequence; But can not form the duplex hybrid structure to the single base mismatch sequence, although or can form unsettled double-spiral structure, this double-spiral structure can be destroyed under mild conditions.
The described universal base oligonucleotide that contains can comprise marker, also can not comprise marker.What comprise marker is directly used in the DNA hybridization check, especially DNA chip hybridization and detection containing the universal base oligonucleotide sequence; Do not contain first hybridizing with target sequence containing the universal base oligonucleotide of marker, the oligonucleotide sequence of then hybridizing is realized the detection to target sequence by biochemical reactions such as extension, connections.
As shown in Figure 1, detecting sequence 1 in figure is that DNA(is as the PCR product) part of sequence; Probe 2 and 3 is the conventional hybridization probes that consist of base A, G, C, T, its middle probe 2 and detection sequence 1 complete complementary, and probe 3 has a single base mismatch with detection sequence 1; Probe 4 and 5 is hybridization probes of the present invention of introducing universal base hypoxanthine deoxyriboside (I), its middle probe 4 and detection sequence 1 complete complementary, and probe 5 has a single base mismatch with detection sequence 1; Pigment 6 and 7 be mark different fluorescence molecule.In the conventional hybridization probe, owing between probe 3 and detection sequence 1, only having a base mispairing, and all the other bases are all just being joined, and therefore need strict hybridization and hybridization post-treatment condition, could ensure probe 3 and detect between sequence 1 not hybridize; And at the hybridization probe of the present invention 5 containing the universal base hypoxanthine deoxyriboside, for and detect the hybridization between sequence 1, except single base mismatch, increased again and formed the lower universal base of double-spiral structure stability, therefore under a maneuverable condition, this double-spiral structure just may not form, and only has probe 4 to form hybridization with detecting sequence, and probe 5 can not form hybridization with the detection sequence.
As shown in Figure 2, detect the part that sequence 8 is marker DNA (as the PCR product) sequence in figure; Probe 9 and 10 is the conventional hybridization probes that consist of base A, G, C, T, its middle probe 9 and detection sequence 8 complete complementaries, and probe 10 has a single base mismatch with detection sequence 8; Probe 11 and 12 is hybridization probes of the present invention of introducing universal base hypoxanthine deoxyriboside (I), its middle probe 11 and detection sequence 8 complete complementaries, and probe 12 has a single base mismatch with detection sequence 8; The fluorescence molecule that pigment 6 is mark.In the conventional hybridization probe, owing between probe 10 and detection sequence 8, only having a base mispairing, and all the other bases are all just being joined, and therefore need strict hybridization and hybridization post-treatment condition, could ensure probe 10 and detect between sequence 8 not hybridize; And at the hybridization probe of the present invention 11 and 12 containing the universal base hypoxanthine deoxyriboside, and only having probe 11 to form hybridization with detecting sequence, probe 12 can not form hybridization with the detection sequence.
(1), DNA extraction
80 peripheral blood sample adopt traditional protein kinase K and phenol/chloroform extraction process to extract the genomic dna in peripheral blood.
(2), PCR reaction
(5 ' amido modified (NH2)) shown in forward primer SEQ ID NO:1:
5′-NH
2-TTTTTTTTTTGCAGCCGCTATTAAAGGTTCG-3′
Shown in reverse primer SEQ ID NO:2:
5′-GGGCTCTGCCATCTTAACAAA-3′
Adopt above-mentioned forward primer and reverse primer to carry out pcr amplification to the fragment containing mitochondrial mutations site C3206T in different DNA samples.The pcr amplification system of 25 μ l comprises: 200ng genomic dna, 0.2mM dNTP, the forward primer of 0.8 μ M and 0.8 μ M reverse primer, 1U Taq archaeal dna polymerase, 1 * amplification buffer, 1.5mMMgCl
2.Amplification condition: 94 ℃ of denaturations 5 minutes, 35 thermal cyclings: 45 seconds～72 ℃ of 30 seconds～55 ℃ annealing of 94 ℃ of sex change are extended 45 seconds, and last 72 ℃ are extended 7 minutes, obtain the PCR product.
(3), the processing of PCR product
Add 95% ethanol of-20 ℃ of 4 times of PCR product amounts in the PCR product, place under-20 ℃ of freezing states 3 hours, under 10000 rotating speeds centrifugal 4 minutes, supernatant liquid is shifted out, and, 50 ℃ of dryings 30 minutes, obtain PCR ethanol purification throw out.
(4), the preparation of DNA chip
Dissolve PCR ethanol purification throw out with the aqueous solution of the 1-(3-dimethylamino-propyl) containing 1～2%-3-ethyl-carbodiimide hydrochloride, and solute is transferred in 384 orifice plates, with DNA chip point sample instrument point sample, each sample repeats 4 times, preparation PCR product D NA chip.After point sample completes, the DNA chip is at 37 ℃, and the Water Under 3h of humidity 80%, then use respectively 2 * SSC/0.5%SDS, 0.1 * SSC/0.5%SDS, distilled water washing DNA chip, and wherein SSC means sodium citrate buffer solution, and SDS means sodium laurylsulfonate.
(5), the hybridization of DNA chip
With the probe hybridization PCR product D NA chip that contains universal base hypoxanthine deoxyriboside (I), (0.5 μ M probe one detects wild-type to the probe bulk crossing liquid of 1 μ M, 0.5 μ M probe two detects saltant type) be evenly distributed on PCR product D NA chip, then cover conversion zone with cover glass, and reduce the evaporation of reaction solution, at 37 ℃ of reaction 0.5h.
Shown in probe one SEQ ID NO:3 (5 ' Cy3 fluorochrome label (Cy3)):
5′-Cy3–GGIIGTGGGIITA-3′
Shown in probe two SEQ ID NO:4 (5 ' Cy5 fluorochrome label (Cy5)):
5′-Cy5-GGIIGTAGGIITA-3′
(6), hybridization signal obtaining and analyzing
After hybridization completes, use respectively 2 * SSC/0.5%SDS, 0.1 * SSC/0.5%SDS washings washing DNA chip, finally clean the washings on slide with distilled water, then the fluorescent signal with scanner scanning Cy3 and Cy5 by the DNA chip.As shown in Figure 3 and Figure 4, the analytical results of 80 samples shows, the order according to sample in 384 orifice plates, and except the 4th, 34,35,36,38,39,79 being saltant type, all the other samples are wild-type.
The SNP in embodiment 2 mankind rs988748 sites detects
(1), the preparation of DNA chip
By 2 probe three and probes four that detect mankind rs988748 site, be mixed with the concentration and probe concentration of 4 μ M with the 1-(3-dimethylamino-propyl) containing 1～2%-aqueous solution of 3-ethyl-carbodiimide hydrochloride respectively, and transfer in 384 orifice plates, with DNA chip point sample instrument point sample, each sample repeats 2 times, prepares the DNA chip of 2 * 2 lattice points.After point sample completes, the DNA chip is at 37 ℃, and the Water Under 3h of humidity 80%, then use respectively 2 * SSC/0.5%SDS, 0.1 * SSC/0.5%SDS, distilled water washing DNA chip.
(5 ' amido modified (NH2)) shown in probe Three S's EQ ID NO:5:
5′-NH
2-TTTTTTTTTTAGGIICTCTGIIGTA-3′
(5 ' amido modified (NH2)) shown in probe four SEQ ID NO:6:
5′-NH
2-TTTTTTTTTTAGGIICTGTGIIGTA-3′
(2), PCR reaction
Shown in forward primer SEQ ID NO:7:
5′-TAGGGTTCCTCCAGTCCTTT-3′
Shown in reverse primer SEQ ID NO:8:
5′-CAGCACAGATGGCAGAGTTTA-3′
Adopt above-mentioned forward primer and reverse primer to carry out asymmetric PCR to DNA sample.The pcr amplification system of 25 μ l comprises: the genomic dna that 200ng extracts from the human blood sample, 0.2mM dNTP(is Cy3-dCTP/dCTP=4/6 wherein), the forward primer of 0.2 μ M and 2.4 μ M reverse primers, 1U Taq archaeal dna polymerase, 1 * amplification buffer, 1.5mM MgCl
2.Amplification condition: 94 ℃ of denaturations 5 minutes, 40 thermal cyclings: 30 seconds～72 ℃ of 20 seconds～57 ℃ annealing of 95 ℃ of sex change are extended 30 seconds, and last 72 ℃ are extended 5 minutes.
(3), the hybridization of DNA chip
The PCR reaction solution is heated to 95 ℃, then cooling fast on ice, and cooled PCR reaction solution is evenly distributed on the DNA chip, then with cover glass, cover conversion zone, and reduce the evaporation of reaction solution, at 37 ℃ of reaction 0.5h.
(4), hybridization signal obtaining and analyzing
After hybridization completes, use respectively 2 * SSC/0.5%SDS, 0.1 * SSC/0.5%SDS washings washing DNA chip, then clean the washings on the DNA chip with distilled water, finally the fluorescent signal with scanner scanning Cy3 by the DNA chip.According to the position distribution of 2 probes on the DNA chip, if the DNA chip only has probe three to have the fluorescent signal of obvious Cy3, rs988748 site that can judgement sample be the wild-type homozygote as shown in Figure 5 a; If the DNA chip only has probe four to have the fluorescent signal of obvious Cy3, rs988748 site that can judgement sample is the saltant type homozygote, as shown in Figure 5 c; If DNA chip probe three and probe four all have the fluorescent signal of obvious Cy3, rs988748 site that can judgement sample is shown in heterozygote Fig. 5 b.
Claims (7)
1. one kind contains the universal base oligonucleotide sequence, it is characterized in that, the described universal base oligonucleotide sequence that contains comprises basic base and universal base, and described basic base and target sequence complete complementary, comprising the basic base corresponding with target sequence SNV site; Described universal base can not corresponding target sequence SNV site, is positioned at the one or both sides of the basic base corresponding with target sequence SNV site; Described basic base is A, G, C, T, and described universal base is can form the nucleosides of hydrogen bond pairing with A, G, C, T.
2. the universal base oligonucleotide sequence that contains according to claim 1, is characterized in that, described basic base is 11-50; Described universal base is 1-8.
3. the universal base oligonucleotide sequence that contains according to claim 2, is characterized in that, described basic base is 13-18; Described universal base is 2-4.
4. according to claim 1 or the 2 or 3 described universal base oligonucleotide sequences that contain, it is characterized in that, described universal base is hypoxanthine deoxyriboside, 2 ' ribodesose-3 ' nitro-pyrrole or 2 ' ribodesose-5 ' nitroindoline.
5. according to claim 1 or the 2 or 3 described universal base oligonucleotide sequences that contain, it is characterized in that, the described universal base oligonucleotide sequence that contains comprises marker or does not comprise marker.
6. claim 1 or 2 or 3 described containing the application of universal base oligonucleotide sequence in the DNA hybridization analysis.
7. claim 1 or the 2 or 3 described application in the DNA hybridization hybrid chip is analyzed containing the universal base oligonucleotide sequence.
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| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN109868309A (en) * | 2019-03-05 | 2019-06-11 | 厦门市安美捷生物工程有限公司 | Single stranded DNA amplification method based on universal base replacement insertion |
Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1833034A (en) * | 2003-06-20 | 2006-09-13 | 埃克斯魁恩公司 | Probes, libraries and kits for analysis of mixtures of nucleic acids and methods for constructing the same |
| CN1840695A (en) * | 2006-02-08 | 2006-10-04 | 北京博奥生物芯片有限责任公司 | Sequence specific oligonucleotide probe and its use |
| CN101072882A (en) * | 2004-09-10 | 2007-11-14 | 塞昆纳姆股份有限公司 | Methods for long-range sequence analysis of nucleic acids |
| CN101090979A (en) * | 2004-12-22 | 2007-12-19 | 埃克斯魁恩公司 | Probes, libraries and kits for analysis of mixtures of nucleic acids and methods for constructing the same |
| CN101240341A (en) * | 2008-03-14 | 2008-08-13 | 东南大学 | DNA sequencing method by using thiooligonucleotide probe |
-
2013
- 2013-10-10 CN CN201310469720.8A patent/CN103484458A/en active Pending
Patent Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1833034A (en) * | 2003-06-20 | 2006-09-13 | 埃克斯魁恩公司 | Probes, libraries and kits for analysis of mixtures of nucleic acids and methods for constructing the same |
| CN101072882A (en) * | 2004-09-10 | 2007-11-14 | 塞昆纳姆股份有限公司 | Methods for long-range sequence analysis of nucleic acids |
| CN101090979A (en) * | 2004-12-22 | 2007-12-19 | 埃克斯魁恩公司 | Probes, libraries and kits for analysis of mixtures of nucleic acids and methods for constructing the same |
| CN1840695A (en) * | 2006-02-08 | 2006-10-04 | 北京博奥生物芯片有限责任公司 | Sequence specific oligonucleotide probe and its use |
| CN101240341A (en) * | 2008-03-14 | 2008-08-13 | 东南大学 | DNA sequencing method by using thiooligonucleotide probe |
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN109868309A (en) * | 2019-03-05 | 2019-06-11 | 厦门市安美捷生物工程有限公司 | Single stranded DNA amplification method based on universal base replacement insertion |
| CN109868309B (en) * | 2019-03-05 | 2022-11-11 | 苏州恩可医药科技有限公司 | Single-stranded DNA amplification method based on universal base substitution insertion |
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