CN102226177A - Fluorescent quantitative detection kit for Notch pathway key molecule - Google Patents
Fluorescent quantitative detection kit for Notch pathway key molecule Download PDFInfo
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- CN102226177A CN102226177A CN2011101132998A CN201110113299A CN102226177A CN 102226177 A CN102226177 A CN 102226177A CN 2011101132998 A CN2011101132998 A CN 2011101132998A CN 201110113299 A CN201110113299 A CN 201110113299A CN 102226177 A CN102226177 A CN 102226177A
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Abstract
The invention discloses a kit for performing a relative quantitative detection to a key molecule in Notch pathway by using a reverse transcription primer and multi gene specific primer with matching amplification. The kit comprises a reagent A: 50 mu L of 5 mu M reverse transcription primer; a reagent B: 50 mu L of 20 U/mu L M-Mu LV reverse transcriptase; a reagent C: 200 mu L of 5*reverse transcription buffer containing the with dNTP concentration of 5m M, Tris-HCl concentration of 250 mM, KCl concentration is 250 mM, MgCl2 concentration of 20 mM, and 50nM of DTT; a reagent D: which is composed of 2*PCR buffer, 1mL liquid comprises 100 U of tap enzyme, 600 nM of ROX, 1*Green I dye, 2mM of dNTPs, 80mM of Tris-HCl, 80mM of KCl, 40mM of (NH4)2SO4, 6mM of MgSO4; 9 reaction tubes provided in 8 reactions of PCR containing primer. The full set of kit provided in the invention performs a relative quantification to the multi key molecules such as mRNA expression of NOTCH1, NUMB, DLL3, GCN5L2, HDAC2, EP300, Hes6, Hes1Mrna in NOTCH pathway at a time, thereby the variation of the expression can be reflected.
Description
Technical field
The present invention relates to biotechnology, is a kind of molecular biology method based on polymerase chain reaction (PCR), particularly relates to the polymerase chain reaction (Q-PCR) of fluorescent quantitation.The expression that reflects each molecule in this path by the relative content that detects a plurality of key molecules in the Notch path.In the primer of synthetic cDNA its 3 ' end introduce 11 specific nucleotide sequences can reverse transcription mRNA, and be with a high-quality nucleotide sequence to carry out Q-PCR at 5 ' end various mRNA increased and relative quantification as label (also being primer location) and offside gene-specific primer.After the content with β-Actin compares, draw the relative content of various mRNA.The relative content of NOTCH1, NUMB, DLL3, GCN5L2, HDAC2, EP300, Hes6, Hes1mRNA can reflect the expression of individuality NOTCH path under different pathological status.This method can disposable genetic expression to a plurality of key molecules in the NOTCH path be carried out relative quantification, thus key molecule expression difference in the reflection path.After the optimization experiment condition, make up simply, relative quantification detection kit accurately.
Background technology
MRNAs is prevalent in the intravital Nucleotide of people, and its expression is subjected to the adjusting of various factors.The content of mRNA is to the proteinic synthetic important effect that has.Can reflect proteinic synthesizing by quantification of mrna, indirectly the biological function of reflection performance range protein.The differential expression of NOTCH key molecule has been brought into play important effect in important physical, pathologic processes such as the generation of cell development, tumour, progression of disease.In the cell in static and the activated NOTCH path key molecule mRNAs express spectra also different with abundance, show significant differential expression.And under the different pathological states in the NOTCH path key molecule mRNAs express and also to have significant difference, and then in different diseases, brought into play important effect.The expression that detects key molecule in the NOTCH path fast and effectively has very important value to pathologic process and the molecular mechanism of studying various malignant tumours.
The Notch signal path is a very conservative signal pass through mechanism on evolving, this path occurs tending to cause some congenital disorders unusually, has now confirmed the unusual relevant with heredopathias such as CADASIL, Aligile syndromes and spinal cord agenesiss of rib of key molecule in the NOTCH signal path.In addition, the NOTCH signal path plays important effect in the differentiation of cell, propagation and individual growth, therefore also must be relevant with it in the developing of tumour.The differential expression that detects key molecule in the NOTCH path in the generating process of disease fast and effectively is very important to the effect that we clearly are familiar with these key molecules, but at present detection method since technological deficiency, operation loaded down with trivial details, experimental period long, can't stdn, need specific apparatus, situation such as cost an arm and a leg allows the experimenter face very big difficulty.We are by setting up open fluorescent quantitative PCR technique platform, and the mRNA detection scheme of combined innovation is developed brand-new detection method.This method relies on the high sensitivity of round pcr and the effect of fluorescent quantitation, can disposablely carry out relative quantification to a plurality of genetic expressions in the NOTCH path, detect under the different pathological states expression difference of key molecule in the NOTCH signal path, and the method by fluorescent quantitation with visual result show.
The NOTCH signal path plays keying action in the developing of various tumours, be the potential target spot of treatment kinds of tumors.By in body tissue and cultured tumor cells in vitro, finish the extraction of total RNA or short-movie section RNA, and on the fluorescent PCR instrument, detect, obtain the result fast.Sum up above process and can find that this method can effectively shorten experimental period, reduce economic cost, reagent and instrument can be opened use.Be quick, accurate, the simple triage techniques and the quantitative measurement technology of key molecule differential expression in a kind of NOTCH of detection path.Can make the standardized testing of multiple mRNA by this invention, make the original one group of Yeast Nucleic Acid that is difficult to adopt unified routine techniques to detect to carry out differential expression analysis and detection by quantitative by standardized nucleic acid nucleic acid amplification technologies.Can distinguish the big mRNAs differential expression of copy number by chip technology, and the Q-PCR technology can detection by quantitative be hanged down the nucleic acid fragment that copies, at first being with the mRNA of poly VITAMIN B4 to carry out reverse transcription by the primer of tape label to all becomes cDNA, carries out relative quantification by Q-PCR again.
At present, also untapped go out a kind of accurately, fast, the technology of key molecule relative content in simple, the measuring N otch path that expense is cheap.The present invention is based on the Q-PCR technology, introduce the Nucleotide of 11 thymus pyrimidines by the primer 3 ' end of tape label, can the nearly all mRNA of reverse transcription, and carry out Q-PCR by a cover Auele Specific Primer and analyze, carry out the detection of relative quantification, solve present technical bottleneck key molecule rapid detection in the NOTCH signal path.
Summary of the invention
The objective of the invention is to overcome existing various technology to detect the deficiency that various path key molecule differential expressions exist in the tumor research, comprise analyze length consuming time, can not high-throughput, defective such as complex operation.The technology of key molecule relative content in a kind of rapid detection NOTCH path is provided, can have reflected the different expression of body NOTCH path under different pathological states.5 ' the end of reverse transcriptase primer SEQ ID NO.1 has the label of nucleotide sequence, primer is introduced the specificity Nucleotide of 11 successive thymus pyrimidines at 3 ' end simultaneously, can the nearly all sophisticated mRNA of reverse transcription, and introduce the universal primer site by label and carry out the Q-PCR analysis, the differential expression of mRNA is carried out the detection of relative quantification.The primer SEQ ID NO.1 of tape label comprises and derives from the corn sequence, is SEQ ID NO.2, and its 3 ' end contains 11 specificity nucleotide sequences at mRNA poly VITAMIN B4; And be used for β-Actin, NOTCH1, NUMB, DLL3, GCN5L2, HDAC2, EP300, Hes6, Hes1 mRNA are carried out 1 primer of relative quantification, be respectively Auele Specific Primer SEQ ID NO.3, SEQ ID NO.4, SEQ ID NO.5, SEQ ID NO.6, SEQ ID NO.7, SEQ ID NO.8, SEQ ID NO.9, SEQ ID NO.10, SEQ ID NO.11 carries out quantitative analysis after carrying out Q-PCR with universal primer SEQ ID NO.2.
Second purpose of the present invention provides the amplification program that the supporting mRNA of a cover detects, make NOTCH1, NUMB, DLL3, GCN5L2, HDAC2, EP300, Hes6, Hes1 mRNA can be simultaneously detected, reach the purpose of real-time analysis, program is as follows: behind 95 ℃ of pre-sex change 3min; Carry out 40 round-robin amplifications, 95 ℃ of sex change 5sec, 62 ℃ of annealing are extended 35sec and are obtained fluorescence intensity level.
The 3rd purpose of the present invention provides a kind of test kit of relative quantification detection of the NOTCH of being used for signal path key molecule detection.Reagent A: 5 μ M reverse transcriptase primers, 50 μ L; Reagent B:20U/ μ L M-MuLV reversed transcriptive enzyme 50 μ L; Reagent C: it is 5mM (comprising dATP, dCTP, dGTP, dTTP) that 5 * reverse transcription damping fluid, 200 μ L contain dNTP concentration, and Tris-HCl concentration is 250mM, and KCl concentration is 250mM, MgCl
2Concentration is 20mM, 50nM DTT; Reagent D: 2 * PCR damping fluid is formed, and comprises 100U taq enzyme, 600nM ROX in the 1mL liquid, 1 *
Green I dye, 2mM dNTPs (comprising dATP, dCTP, dGTP, dTTP), 80mM Tris-HCl, 80mM KCl, 40mM (NH
4)
2SO
4, 6mM MgSO
4The PCR8 that contains primer joins 9 of reaction tubess: a reaction tubes: the mix primer that contains amplification people NOTCH1 cDNA.Comprise 1 * 10
-3Nmol SEQ ID NO.2 and 1 * 10
-3Nmol SEQ ID NO.4.No. two reaction tubess: the mix primer that contains amplification people NUMB cDNA.Comprise 1 * 10
-3Nmol SEQ ID NO.2 and 1 * 10
-3Nmol SEQ IDNO.5.No. three reaction tubess: the mix primer that contains amplification people DLL3 cDNA.Comprise 1 * 10
-3Nmol SEQID NO.2 and 1 * 10
-3Nmol SEQ ID NO.6.No. four reaction tubess: the mix primer that contains amplification people GCN5L2 cDNA.Comprise 1 * 10
-3Nmol SEQ ID NO.2 and 1 * 10
-3Nmol SEQ ID NO.7.No. five reaction tubess: the mix primer that contains amplification people HDAC2 cDNA.Comprise 1 * 10
-3Nmol SEQ ID NO.2 and 1 * 10
-3Nmol SEQ ID NO.8.No. six reaction tubess: the mix primer that contains amplification people EP300 cDNA.Comprise 1 * 10
-3Nmol SEQ ID NO.2 and 1 * 10
-3Nmol SEQ ID NO.9.No. seven reaction tubess: the mix primer that contains amplification people Hes6 cDNA.Comprise 1 * 10
-3Nmol SEQ ID NO.2 and 1 * 10
-3NmolSEQ ID NO.10.No. eight reaction tubess: the mix primer that contains amplification people Hes6 cDNA.Comprise 1 * 10
-3NmolSEQ ID NO.2 and 1 * 10
-3Nmol SEQ ID NO.11.No. nine reaction tubess: the mix primer that contains amplification people β-ActincDNA.Comprise 1 * 10
-3Nmol SEQ ID NO.2 and 1 * 10
-3Nmol SEQ ID NO.3.
Technical scheme of the present invention is summarized as follows:
One cover is used for NOTCH signal path key molecule mRNA is carried out the primer that relative quantification detects, and a cover of supporting this detection detects the primer of β-Actin internal reference.The reverse transcriptase primer SEQ ID NO.1 that comprises a tape label, its 3 ' end is introduced 11 thymus pyrimidines at the poly-A tail of mRNA; Article one, a section on its sequence of consensus primer and the SEQ ID NO.1 is all SEQ ID NO.2 mutually; Carry out 9 primers of relative quantification at β-Actin, NOTCH1, NUMB, DLL3, GCN5L2, HDAC2, EP300, Hes6, Hes1mRNA, be respectively Auele Specific Primer SEQ ID NO.3, SEQ ID NO.4, SEQ ID NO.5, SEQ ID NO.6, SEQ ID NO.7, SEQ ID NO.8, SEQ ID NO.9, SEQ ID NO.10, SEQID NO.11.
A trace routine that detects at mRNA makes β-Actin, NOTCH1, NUMB, DLL3, GCN5L2, HDAC2, EP300, Hes6, Hes1mRNA can be simultaneously detected, reaches the purpose of real-time analysis, and program is as follows: behind 95 ℃ of pre-sex change 3min; Carry out 40 round-robin amplifications, 95 ℃ of sex change 5sec, 62 ℃ of annealing are extended 35sec and are obtained fluorescence intensity level.
A kind of to β-Actin, NOTCH1, NUMB, DLL3, GCN5L2, HDAC2, EP300, Hes6, Hes1mRNA carry out relative quantification one the cover test kit, its feature comprises 1, is made up of following reagent:
Reagent A: 5 μ M reverse transcriptase primers, 50 μ L;
Reagent B:20U/ μ L M-MuLV reversed transcriptive enzyme, 50 μ L;
Reagent C: it is 5mM (comprising dATP, dCTP, dGTP, dTTP) that 5 * reverse transcription damping fluid, 200 μ L contain dNTP concentration, and Tris-HCl concentration is 250mM, and KCl concentration is 250mM, MgCl
2Concentration is 20mM, 50nM DTT;
Reagent D: 2 * PCR damping fluid is formed, and comprises 100U taq enzyme, 600nM ROX in the 1mL liquid, 1 *
Green I dye, 2mM dNTPs (comprising dATP, dCTP, dGTP, dTTP), 80mM Tris-HCl, 80mM KCl, 40mM (NH
4)
2SO
4, 6mM MgSO
4
9 of PCR 8 reaction tubess that contain primer:
Reaction tubes: the mix primer that contains amplification people NOTCH1cDNA.Comprise 1 * 10
-3Nmol SEQ IDNO.2 and 1 * 10
-3Nmol SEQ ID NO.4.
No. two reaction tubess: the mix primer that contains amplification people NUMB cDNA.Comprise 1 * 10
-3Nmol SEQ IDNO.2 and 1 * 10
-3Nmol SEQ ID NO.5.
No. three reaction tubess: the mix primer that contains amplification people DLL3 cDNA.Comprise 1 * 10
-3Nmol SEQ IDNO.2 and 1 * 10
-3Nmol SEQ ID NO.6.
No. four reaction tubess: the mix primer that contains amplification people GCN5L2cDNA.Comprise 1 * 10
-3Nmol SEQ IDNO.2 and 1 * 10
-3Nmol SEQ ID NO.7.
No. five reaction tubess: the mix primer that contains amplification people HDAC2 cDNA.Comprise 1 * 10
-3Nmol SEQ IDNO.2 and 1 * 10
-3Nmol SEQ ID NO.8.
No. six reaction tubess: the mix primer that contains amplification people EP300 cDNA.Comprise 1 * 10
-3Nmol SEQ IDNO.2 and 1 * 10
-3Nmol SEQ ID NO.9.
No. seven reaction tubess: the mix primer that contains amplification people Hes6 cDNA.Comprise 1 * 10
-3Nmol SEQ IDNO.2 and 1 * 10
-3Nmol SEQ ID NO.10.
No. eight reaction tubess: the mix primer that contains amplification people Hes1 cDNA.Comprise 1 * 10
-3Nmol SEQ IDNO.2 and 1 * 10
-3Nmol SEQ ID NO.11.
No. nine reaction tubess: the mix primer that contains amplification people β-Actin cDNA.Comprise 1 * 10
-3Nmol SEQID NO.2 and 1 * 10
-3Nmol SEQ ID NO.3.
Advantage of the present invention:
The NOTCH signal path plays keying action in the developing of various tumours, be the potential target spot of treatment kinds of tumors.Shown that in some congenital disorders and kinds of tumors generation evolution the change of mRNA express spectra appears in key molecule in the NOTCH path, by design of primers of the present invention, when reverse transcription, bring a good primer location into as second primer location of step during nucleic acid amplification.The temperature of reaction of each primer has all been carried out standard, and the fragment reaction length has also been carried out stdn, and all mRNA quantitatively all can carry out under identical condition, makes to have comparability between different experiments.The present invention can be based upon on the Q-PCR technology platform of mRNA detection, not only can shorten experimental period, can improve the rate of utilization of open instrument simultaneously, reduces economic cost, a brand-new examination criteria and evaluation system are provided.By finishing the extraction of total RNA or short-movie section RNA after the cultured tumor cells in vitro fast, and on the fluorescent PCR instrument, detect.Sum up above process and can find that this method can effectively shorten experimental period, reduce economic cost, reagent and instrument can be opened use.By optimization experiment reaction system and reaction conditions, form to detect sensitive, test kit easily, key molecule provides the reliable detection technology in the NOTCH signal path for individuality detects.
Description of drawings
Fig. 1 is the detection effect of carcinoma of the pancreas sample NOTCH1, NUMB, DLL3, GCN5L2, HDAC2, EP300, Hes6, Hes1mRNA.
Fig. 2 is the solubility curve of each sample NOTCH1, NUMB, DLL3, GCN5L2, HDAC2, EP300, Hes6, Hes1 detection.
Embodiment
Method therefor is ordinary method if no special instructions among the following embodiment.
Embodiment 1
Add 1ml Trizol in culturing cell or the cancerous tissue, vibrator vibration or pipettor are inhaled and are beaten mixing for several times, place 5min.Add chloroform 200, violent mixing left standstill 5 minutes.Behind centrifugal 10 minutes of the 12000rpm, get supernatant liquid 500 μ L, add 500 μ L Virahols-20 degree and placed 2 hours.Behind centrifugal 15 minutes of the 12000rpm, abandon supernatant liquor.Use 75% washing with alcohol, 12000rpm abandons supernatant liquor after centrifugal 15 minutes.After repeating to wash 2 times, placed 10 minutes under the room temperature, add the water 30 μ L that DEPC handles.Fully dissolving ,-80 degree are preserved.
Embodiment 2
The reverse transcription schedule of operation of a kind of supporting this method testing goal mRNA.Contain 1 μ LmRNA (0.5pg-1 μ g) in the 10 μ L systems, primer mixed solution reagent A 1 μ L, (final concentration dNTP is 1mM for reversed transcriptive enzyme reagent B 1 μ L and reverse transcription damping fluid reagent C 2 μ L, primer is 500nM, M-MuLV reversed transcriptive enzyme concentration is 40U, Tris-HCl concentration is 50mM, and KCl concentration is 50mM, MgCl
2Concentration is 4mM), add water 5 μ L.The reverse transcription program is for placing 5min on ice, 30 ℃ of 30min, 95 ℃ of 5min.After reverse transcription is finished, in vitro add 90 μ L water, carry out 10 times dilution.
Embodiment 3
A kind of Q-PCR method that detects NOTCH path key molecule mRNA, and operating process and response procedures.Each project need be carried out 2 detections that repeat pipe, and every reaction tubes reaction is to can be 10 μ L-50 μ L.In 10 μ L Q-PCR systems, the cDNA that at first gets after 18 μ L dilute adds in the barren pipe, adds the reagent D of 90 μ L, after adding water 72 μ L mixing, each 10 μ L that take out add respectively in the 1-9 pipe that final concentration is respectively in (totally 2 pipes) each pipe: primer 2 00nM, and 300nM ROX, 0.5 *
Green I dye, 1mm dNTPs, 40mM Tris-HCl, 40mM KCl, 20mM (NH
4)
2SO
4, 3mM MgSO
4, Taq enzyme 0.5U.Stick pad pasting, on ABI 7500, detect.As follows to cDNA amplification and quant program: behind 95 ℃ of pre-sex change 3min; Carry out 40 circulations, 95 ℃ of sex change 5sec, 35sec (this step is gathered fluorescent value) is extended in 62 ℃ of annealing.By ordinary method the Ct value of each PCR pipe is analyzed, after comparing, obtained a result with internal reference.
Claims (12)
1. universal primer that is used for reverse transcription mRNA is characterized in that the nucleotide sequence of a tape label, and 3 ' end is with the poly VITAMIN B4 specificity complementation of 11 successive thymus pyrimidines and mRNA, and nucleotide sequence is SEQ ID NO.1, can be used for reverse transcription mRNA.
2. the offside primer that is used for mRNA is carried out quantitative PCR according to claim 1 is SEQ ID NO.2 by comprising a high-quality primer location in the SEQ IDNO.1.
3. according to claim 1ly be used for that (β-Actin) carries out the Auele Specific Primer SEQ ID NO.3 of quantitative PCR, with people β-one section complementation of Actin mRNA to the beta Actin muscle.
4. the Auele Specific Primer SEQ ID NO.4 that is used for NOTCH1 mRNA is carried out quantitative PCR according to claim 1 is with one section complementation of people NOTCH1 mRNA.
5. the Auele Specific Primer SEQID NO.5 that is used for Numb mRNA is carried out quantitative PCR according to claim 1 is with one section complementation of people NUNB mRNA.
6. the Auele Specific Primer SEQID NO.6 that is used for DLL3 mRNA is carried out quantitative PCR according to claim 1 is with one section complementation of people DLL3 mRNA.
7. the Auele Specific Primer SEQ ID NO.7 that is used for GCN5L2 mRNA is carried out quantitative PCR according to claim 1 is with one section complementation of people GCN5L2 mRNA.
8. the Auele Specific Primer SEQID NO.8 that is used for HDAC2 mRNA is carried out quantitative PCR according to claim 1 is with one section complementation of people HDAC2 mRNA.
9. the Auele Specific Primer SEQID NO.9 that is used for EP300 mRNA is carried out quantitative PCR according to claim 1 is with one section complementation of people EP300mRNA.
10. the Auele Specific Primer SEQ ID NO.10 that is used for Hes6 mRNA is carried out quantitative PCR according to claim 1 is with one section complementation of people Hes6mRNA.
11. the Auele Specific Primer SEQ ID NO.11 that is used for Hes1 mRNA is carried out quantitative PCR according to claim 1 is with one section complementation of people Hes1mRNA.
12. according to claim 1 to mRNA carry out relative quantification one the cover test kit, its feature comprises 1, is made up of following reagent:
Reagent A: 5 μ M reverse transcriptase primers, 50 μ L;
Reagent B:20U/ μ L M-MuLV reversed transcriptive enzyme, 50 μ L;
Reagent C: it is 5mM (comprising dATP, dCTP, dGTP, dTTP) that 5 * reverse transcription damping fluid, 200 μ L contain dNTP concentration, and Tris-HCl concentration is 250mM, and KCl concentration is 250mM, MgCl
2Concentration is 20mM, 50nM DTT;
Reagent D: 2 * PCR damping fluid is formed, and comprises 100U taq enzyme, 600nM ROX in the 1mL liquid, 1 *
Green I dye, 2mM dNTPs (comprising dATP, dCTP, dGTP, dTTP), 80mM Tris-HCl, 80mM KCl, 40mM (NH
4)
2SO
4, 6mM MgSO
4
9 of PCR 8 reaction tubess that contain primer:
Reaction tubes: the mix primer that contains amplification people NOTCH1cDNA.Comprise 1 * 10
-3Nmol SEQ IDNO.2 and 1 * 10
-3Nmol SEQ ID NO.4.
No. two reaction tubess: the mix primer that contains amplification people NUMB cDNA.Comprise 1 * 10
-3Nmol SEQ IDNO.2 and 1 * 10
-3Nmol SEQ ID NO.5.
No. three reaction tubess: the mix primer that contains amplification people DLL3 cDNA.Comprise 1 * 10
-3Nmol SEQ IDNO.2 and 1 * 10
-3Nmol SEQ ID NO.6.
No. four reaction tubess: the mix primer that contains amplification people GCN5L2cDNA.Comprise 1 * 10
-3Nmol SEQ IDNO.2 and 1 * 10
-3Nmol SEQ ID NO.7.
No. five reaction tubess: the mix primer that contains amplification people HDAC2 cDNA.Comprise 1 * 10
-3Nmol SEQ IDNO.2 and 1 * 10
-3Nmol SEQ ID NO.8.
No. six reaction tubess: the mix primer that contains amplification people EP300 cDNA.Comprise 1 * 10
-3Nmol SEQ IDNO.2 and 1 * 10
-3Nmol SEQ ID NO.9.
No. seven reaction tubess: the mix primer that contains amplification people Hes6 cDNA.Comprise 1 * 10
-3Nmol SEQ IDNO.2 and 1 * 10
-3Nmol SEQ ID NO.10.
No. eight reaction tubess: the mix primer that contains amplification people Hes1 cDNA.Comprise 1 * 10
-3Nmol SEQ IDNO.2 and 1 * 10
-3Nmol SEQ ID NO.11.
No. nine reaction tubess: the mix primer that contains amplification people β-Actin cDNA.Comprise 1 * 10
-3Nmol SEQID NO.2 and 1 * 10
-3Nmol SEQ ID NO.3.
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| CN2011101132998A CN102226177A (en) | 2011-04-25 | 2011-04-25 | Fluorescent quantitative detection kit for Notch pathway key molecule |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN103627814A (en) * | 2013-12-13 | 2014-03-12 | 青岛大学医学院附属医院 | Reagent for detecting Notch signal path as well as PCR (Polymerase Chain Reaction) detecting method and application thereof |
| CN104263722A (en) * | 2014-08-06 | 2015-01-07 | 温州医科大学附属第一医院 | Amplimer used for human dermokine gene subtype mRNA expression quantity detection and kit thereof |
| CN107460237A (en) * | 2017-06-23 | 2017-12-12 | 中南大学 | HES6 is treating the purposes of chronic myelocytic leukemia as molecular target |
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| CN102154275A (en) * | 2011-01-26 | 2011-08-17 | 余震 | Relative quantification kit for detecting ribonucleic acid of Thl/Th2 cell factors |
| CN103627814A (en) * | 2013-12-13 | 2014-03-12 | 青岛大学医学院附属医院 | Reagent for detecting Notch signal path as well as PCR (Polymerase Chain Reaction) detecting method and application thereof |
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| CN102154275A (en) * | 2011-01-26 | 2011-08-17 | 余震 | Relative quantification kit for detecting ribonucleic acid of Thl/Th2 cell factors |
| CN103627814A (en) * | 2013-12-13 | 2014-03-12 | 青岛大学医学院附属医院 | Reagent for detecting Notch signal path as well as PCR (Polymerase Chain Reaction) detecting method and application thereof |
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Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103627814A (en) * | 2013-12-13 | 2014-03-12 | 青岛大学医学院附属医院 | Reagent for detecting Notch signal path as well as PCR (Polymerase Chain Reaction) detecting method and application thereof |
| CN103627814B (en) * | 2013-12-13 | 2015-05-27 | 青岛大学医学院附属医院 | Reagent for detecting Notch signal path as well as PCR (Polymerase Chain Reaction) detecting method and application thereof |
| CN104263722A (en) * | 2014-08-06 | 2015-01-07 | 温州医科大学附属第一医院 | Amplimer used for human dermokine gene subtype mRNA expression quantity detection and kit thereof |
| CN107460237A (en) * | 2017-06-23 | 2017-12-12 | 中南大学 | HES6 is treating the purposes of chronic myelocytic leukemia as molecular target |
| CN107460237B (en) * | 2017-06-23 | 2020-03-27 | 中南大学 | Application of HES6 as molecular target in treating chronic granulocytic leukemia |
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Application publication date: 20111026 |