CN112662758B - miRNA marker related to auxiliary diagnosis of endometrial receptivity and application thereof - Google Patents
miRNA marker related to auxiliary diagnosis of endometrial receptivity and application thereof Download PDFInfo
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Abstract
The invention discloses a miRNA marker for assisting in diagnosing endometrial receptivity, which comprises miRNA of hsa-miR-16-2-3p, hsa-miR-92b-5p, hsa-miR-130a-3p and has-miR-9024-3p and primers thereof; the invention also provides an application of the miRNA marker for auxiliary diagnosis of endometrial receptivity, which is characterized in that: the method comprises the following steps: s1 collecting maternal blood, and preparing supernatant; s2 centrifugal column filtering tube, and grading filtering to prepare miRNA crude product; s3 preparing miRNA samples; s4 synthesizing the miRNA into cDNA by using the kit, and performing a PCR amplification system and a PCR amplification program to obtain miRNA content data; s5, constructing a CT content curve, and determining different miRNA contents in different periods; the invention has the beneficial effects that: by limiting miRNA and primers thereof and taking four primers after specific screening as a detection basis, the endometrial receptivity period can be quickly and accurately identified.
Description
Technical Field
The invention relates to the field of genetic engineering and reproductive medicine, in particular to a miRNA marker for auxiliary diagnosis of endometrial receptivity and application thereof.
Background
Currently, the incidence of Chinese infertility is around 15-20%, and the number of infertility patients increases at a rate of hundreds of thousands every year. The delay of marriage and childbearing age, the accumulation of working pressure, and the change of life style, etc. directly or indirectly reduce the fertility of people. With the birth of the first test-tube infant in the world, in vitro fertilization-embryo transfer (IVF-ET) has become an effective way for an infertile female to obtain pregnancy as soon as possible. Although the development and technological innovation of assisted reproduction technology has enabled most infertility patients to obtain good quality embryos by embryo in vitro culture technology and then to become pregnant, a part of good quality embryos are lost in the early or even very early stages of pregnancy. Research shows that embryo factors and endometrial receptivity factors have certain correlation with the successful implantation of embryos.
Endometrial receptivity refers to the ability of the endometrium to accept the embryo, a capability that allows the zygote to locate, adhere, invade, and alter the intimal-interstitium for implantation into the endometrium. Endometrial receptivity is limited in time, which is referred to as the "Implantation window" period (WOI). The "planting window" period is also called a receptive period, generally appears 6-8 days after ovulation or 5-7 days after fertilization, needs the support of female and progestational hormones secreted by corpus luteum, and is also influenced by various genes, proteins, cytokines and adhesion molecules, in-vitro fertilization-embryo transfer (IVF-ET) in assisted reproduction, an artificial simulated menstrual cycle is usually established, when the endometrium reaches 7 mm, the endometrium is converted from a proliferative state to a differentiated state by administering progesterone in different dosage forms, the timing of administering progesterone is called a luteal transformation period, and the embryo can be placed into the uterus on the 3 rd-5 th day after the luteal transformation period. Under normal conditions, only after the 5 th day after luteal phase transformation, the endometrium can receive the embryo, and the embryo cannot be implanted unless it is received at other times.
The synchronization of embryo implantation and implantation window opening is a necessary condition for successful implantation, and the failure of 2/3 repeated implantation is caused by insufficient endometrial receptivity, so that good endometrial receptivity is a necessary condition for successful embryo implantation. However, the conventional detection means at present is to perform biopsy on the endometrium, because the biopsy is an invasive operation, embryo transplantation needs to be performed in the next IVF-ET cycle, tolerance errors of different cycles exist between the two cycles, and damage to a mother body is not reversible while a large amount of time is consumed.
However, numerous studies have shown that endometrial receptivity is associated with a number of genes (e.g., homeobox gene A10, homeobox gene C8, galectin-3, exon-rich gene forkhead box L2, human and Drosophila ems homeodomain gene Emx2, etc.), proteins (e.g., collagen type IV, pregnancy-associated proteins, estrogen receptors, progesterone receptors, osteopontin, integrins, etc.), cytokines (e.g., LIF, IGF1, interleukin I, colony stimulating factor I, vascular endothelial growth factor, etc.), ovarian steroid hormones, etc.
miRNA is a non-coding RNA, highly conserved, about 18-22 nucleotides in length, and can regulate gene expression at a post-transcriptional level by targeting mRNA for cleavage or transcriptional repression. mirnas are widely found in cells and tissues and are involved in a variety of biological processes. Studies have demonstrated the presence of hundreds of miRNAs in serum/plasma, these small molecular RNAs are stable in nature, abundant, easily detected quantitatively, and present significant disease specificity. The existing mature technologies, including qualitative and quantitative miRNA molecular technologies, show that the method using serum miRNAs as molecular biomarkers is more effective than the traditional specific protein molecular marking method, and opens up a new border for the biomarkers. And studies have shown that regulation of mirnas is associated with reproductive disorders such as polycystic ovary syndrome, endometriosis, endometrial polyps, hydrosalpinx, and the like.
Therefore, the miRNA is combined to detect the uterine receptivity, the miRNA content of the parents at different periods is harmlessly and accurately detected and specified, and data support is provided for the subsequent determination of the receptivity period.
Disclosure of Invention
The invention aims to overcome the defects of the prior art and provide a miRNA marker for assisting in diagnosing endometrial receptivity and application thereof, so as to at least achieve the aim of quickly, harmlessly and accurately determining endometrial receptivity.
The purpose of the invention is realized by the following technical scheme:
an miRNA marker for assisting in diagnosing endometrial receptivity comprises miRNA of hsa-miR-16-2-3p, hsa-miR-92b-5p, hsa-miR-130a-3p and has-miR-9024-3p and primers thereof.
Preferably, in order to further realize the purpose of rapidness and accuracy, the primers of the miRNA include a primer of hsa-miR-16-2-3p shown as SEQ ID No.1, a primer of hsa-miR-92b-5p shown as SEQ ID No.2, a primer of hsa-miR-130a-3p shown as SEQ ID No.3 and a primer of has-miR-9024-3p shown as SEQ ID No. 4; by limiting the miRNA marker primers and taking the four specifically screened primers as a detection basis, the composite detection primers are constructed, so that the miRNA sequences to be detected can be amplified in a large amount, the detected data sample is large enough, and the purpose of rapidness and accuracy is achieved.
Preferably, in order to further achieve the accurate purpose, the miRNA comprises a marker sequence of hsa-miR-16-2-3p shown in SEQ ID No.5, a primer of hsa-miR-92b-5p shown in SEQ ID No.6, a primer of hsa-miR-130a-3p shown in SEQ ID No.7 and a primer of has-miR-9024-3p shown in SEQ ID No. 8; and the DNA sequence of the marker is defined again so as to be matched with the primer sequence, a complete detection tool is constructed, and then the primers are matched to amplify a large amount of miRNA to be detected, so that the aim of rapidness and accuracy is fulfilled.
Preferably, in order to further realize the purpose of rapidness and harmlessness, the primer is applied to the preparation of an auxiliary diagnostic kit for endometrial receptivity; the diagnostic kit is used for detecting serum/plasma miRNA markers related to auxiliary diagnosis of endometrial receptivity; the kit also comprises enzymes and reagents commonly used in PCR reaction; through with the primer of design, construct detect reagent box to the peripheral plasma/serum of parent is direct detection sample object to avoid carrying out the biopsy to the parent, avoid producing the harm to the parent, the holistic detection cycle of PCR is shorter simultaneously, thereby realizes quick harmless purpose.
The invention also provides an application of the miRNA marker for auxiliary diagnosis of endometrial receptivity, which comprises the following steps:
s1, collecting sample blood of different parent corpus luteum transformation periods by using a blood collection tube containing EDTA anticoagulant factors, standing, centrifuging, and collecting supernate;
s2, adding a QIAzol lysine Reagent into the obtained supernatant at room temperature, standing, adding a chloroform solution, oscillating, standing, centrifuging to obtain a mixed supernatant, adding absolute ethanol into the obtained mixed supernatant, mixing uniformly, adding into a RNeasy MinElute spin column filter tube, carrying out graded filtration, and discarding filtrate to obtain a miRNA crude product in the filter tube;
s3, centrifuging the obtained miRNA crude product again, transferring the centrifuged centrifugal column filter tube into an miRNA collecting tube, adding RNase-free water until the whole filter membrane of the centrifugal column filter tube is immersed in the solution, and centrifuging and collecting to obtain an miRNA sample;
s4, synthesizing cDNA (complementary deoxyribonucleic acid) from the obtained miRNA by using a Takara Mir-XTM miRNA First-Strand Synthesis (Cat. number 638313) kit, selecting primers of miRNA markers of hsa-miR-16-2-3p, hsa-miR-92b-5p, hsa-miR-130a-3p and has-miR-9024-3p, mixing the primers with the obtained cDNA, carrying out a PCR (polymerase chain reaction) amplification system and program, detecting the content of the amplified miRNA, and obtaining the content data of the miRNA;
s5 construction of C by miRNA content dataTAnd (4) content curve, determining the content of different miRNA in different periods.
Preferably, for further achieving the purpose of rapidness and accuracy, the graded filtering is as follows:
a. adding 700 mu L Buffer BWT into a centrifugal column filter tube, then adopting 8000 Xg centrifugal force, centrifuging for 15s at room temperature, and abandoning the filtrate;
b. adding 500 μ L Buffer RPE into a centrifugal column filter tube, centrifuging at room temperature for 15s with 8000 Xg centrifugal force, and discarding the filtrate;
c. adding 500 μ L of 80% ethanol into a centrifugal column filter tube, centrifuging at room temperature for 2min by adopting a centrifugal force of 8000 Xg, and discarding the filtrate to finish the fractional filtration;
through carrying out multistage filtration, messenger's miRNA in the sample can fully purify to prevent the interference of other DNA, thereby realize quick accurate purpose.
Preferably, in order to further achieve the purpose of rapid and accurate synthesis, the specific process of synthesizing cDNA by miRNA is that in the kit, 3.75 μ L of miRNA sample, 5 μ L of mRQ Buffer (2 x), 1.25 μ L of mRQ Enzyme are added into 200 μ L of RNase-free EP tube, after mixing uniformly, the EP tube is heated to 37 ℃, insulated for 1h, heated to 85 ℃, insulated for 5min, and then 90 μ L of ddH is added2O is subjected to constant volume to 100 mu L to obtain the cDNA solution; by passingmiRNA is reversely transcribed into complementary DNA, so that PCR amplification is directly carried out by utilizing a complementary strand to obtain a large number of samples, and the aim of rapidness and accuracy is fulfilled.
Preferably, for further achieving the purpose of rapidness and accuracy, the PCR amplification system is: 2 μ L of cDNA solution, 0.5 μ L of miRNA marker primers, 0.5 μ L of 10 μmol/L mRQ 3' Primer, 12.5 μ L of TB Green Advantage Premix (2X), 0.5 μ L of ROX Dye (50X) and 9 μ L of ddH2O；
The amplification procedure is pre-denaturation at 95 ℃ for 10min, the target sequence is amplified at 95 ℃ for 15s and 60 ℃ for 30s in 40 cycles, and the dissolution reaction is at 95 ℃ for 15s, 60 ℃ for 60s and 95 ℃ for 15 s; the quantity of the cDNA is amplified in large quantity by limiting a PCR amplification system and a PCR amplification program, so that the whole quantity of the amplified cDNA is conveniently measured, the content of miRNA is indirectly deduced, and the aim of rapidness and accuracy is fulfilled.
The invention has the beneficial effects that:
1. by limiting miRNA and primers thereof and taking four primers after specific screening as detection bases, a composite detection primer is constructed, so that the miRNA sequences to be detected can be amplified in a large amount, the detected data sample is large enough, and the purpose of rapidness and accuracy is achieved.
2. Through with the primer of design, construct detect reagent box to the peripheral plasma/serum of parent is direct detection sample object to avoid carrying out the biopsy to the parent, avoid producing the harm to the parent, the holistic detection cycle of PCR is shorter simultaneously, thereby realizes quick harmless purpose.
3. And the DNA sequence of the marker is defined again so as to be matched with the primer sequence, a complete detection tool is constructed, and then the primers are matched to amplify a large amount of miRNA to be detected, so that the aim of rapidness and accuracy is fulfilled.
4. Through carrying out multistage filtration, messenger's miRNA in the sample can fully purify to prevent the interference of other DNA, thereby realize quick accurate purpose.
5. By reverse transcribing miRNA into complementary DNA, PCR amplification is directly carried out by using complementary strands to obtain a large number of samples, thereby achieving the purpose of rapidness and accuracy.
6. The quantity of the cDNA is amplified in large quantity by limiting a PCR amplification system and a PCR amplification program, so that the whole quantity of the amplified cDNA is conveniently measured, the content of miRNA is indirectly deduced, and the aim of rapidness and accuracy is fulfilled.
Drawings
FIG. 1 is a graph of the C of hsa-miR-16-2-3p with different miRNA contents at different stagesTA graph is shown schematically;
FIG. 2 is a schematic representation of CT curves for hsa-miR-92b-5p with different miRNA content at different time periods;
FIG. 3 is a graph of C of hsa-miR-130a-3p with different miRNA contents at different stagesTA graph is shown schematically;
FIG. 4 is a schematic representation of CT curves of has-miR-9024-3p at different miRNA contents at different stages;
wherein, denotes a p value < 0.05;
FIG. 5 is a schematic view of a ROC curve.
Detailed Description
The technical solutions of the present invention are further described in detail below with reference to the accompanying drawings, but the scope of the present invention is not limited to the following.
Example 1
An miRNA marker for assisting in diagnosing endometrial receptivity comprises miRNA of hsa-miR-16-2-3p, hsa-miR-92b-5p, hsa-miR-130a-3p and has-miR-9024-3p and primers thereof.
In order to further realize the purpose of rapidness and accuracy, the primers of the miRNA include a primer of hsa-miR-16-2-3p shown as SEQ ID No.1, a primer of hsa-miR-92b-5p shown as SEQ ID No.2, a primer of hsa-miR-130a-3p shown as SEQ ID No.3 and a primer of has-miR-9024-3p shown as SEQ ID No. 4; by limiting the miRNA marker primers and taking the four specifically screened primers as a detection basis, the composite detection primers are constructed, so that the miRNA sequences to be detected can be amplified in a large amount, the detected data sample is large enough, and the purpose of rapidness and accuracy is achieved.
In order to further realize the accurate purpose, the marker sequence of the miRNA including hsa-miR-16-2-3p is shown as SEQ ID No.5, the primer of hsa-miR-92b-5p is shown as SEQ ID No.6, the primer of hsa-miR-130a-3p is shown as SEQ ID No.7, and the primer of has-miR-9024-3p is shown as SEQ ID No. 8; and the DNA sequence of the marker is defined again so as to be matched with the primer sequence, a complete detection tool is constructed, and then the primers are matched to amplify a large amount of miRNA to be detected, so that the aim of rapidness and accuracy is fulfilled.
The oligonucleotide sequences of the above-mentioned miRNAs and their primers are shown in Table 1.
TABLE 1 miRNA sequences and primer sequences thereof
Among them, when synthesizing cDNA, a single miRNA single strand may have a single repetitive nucleotide sequence such as AAAAAAAA added to the 3-terminal of the sequence of miRNA, and thus a primer sequence at the 3-terminal may not be required.
In order to further realize the purpose of rapidness and harmlessness, the primer is applied to the preparation of an auxiliary diagnostic kit for endometrial receptivity; the diagnostic kit is used for detecting serum/plasma miRNA markers related to auxiliary diagnosis of endometrial receptivity; the kit also comprises enzymes and reagents commonly used in PCR reaction; through with the primer of design, construct detect reagent box to the peripheral plasma/serum of parent is direct detection sample object to avoid carrying out the biopsy to the parent, avoid producing the harm to the parent, the holistic detection cycle of PCR is shorter simultaneously, thereby realizes quick harmless purpose.
The invention also provides an application of the miRNA marker for auxiliary diagnosis of endometrial receptivity, which comprises the following steps:
s1, collecting 2ml of sample blood of different parent luteal transformation periods by using a blood collection tube containing EDTA anticoagulant factors, wherein the luteal transformation periods of different parents are a luteal transformation period (LH = 0), a 4 th day after luteal transformation (LH = 4), a 5 th day after luteal transformation (LH = 5) and a 6 th day after luteal transformation (LH = 6), after blood collection is completed, the blood collection tube is gently inverted up and down for 5 times, after standing for 30min at room temperature, the blood collection tube is centrifuged for 10min at 24 ℃ by a centrifugal force of 1200 Xg, collected by using a 1.5ml EP tube, then centrifuged for 15min at 4 ℃ by a centrifugal force of 1200 Xg again, and a pale yellow supernatant after centrifugation is collected and stored at-80 ℃;
s2 for the supernatant, the miRNA in the Plasma sample is extracted by using Qiagen miRNeasy Serum/Plasma Kit (50) Cat No.21784, which comprises the following steps: taking 200 mu L of supernatant stored at room temperature, adding 1000 mu L of QIAzol lysine Reagent for tissue Lysis, repeatedly blowing the supernatant for 10 times by using a pipette gun, standing the supernatant for 5min at room temperature, adding 200 mu L of chloroform solution, mixing the chloroform solution by vortex oscillation for 15s, standing the mixture for 3min at room temperature, centrifuging the mixture for 15min at 4 ℃ by using a centrifugal force of 1200 Xg to obtain mixed supernatant, transferring the mixed supernatant into a 1.5ml EP tube, adding 900 mu L of absolute ethyl alcohol, mixing the mixture uniformly, taking 700 mu L of the mixed supernatant, adding the mixture into an RNeasy MinElute spin column filter tube, centrifuging the mixture for 15s at room temperature by using a centrifugal force of 8000 Xg, carrying out fractional filtration, and discarding filtrate to obtain a miRNA crude product in the centrifugal column filter tube;
s3, opening the cover of the obtained miRNA crude product of the centrifugal column filter tube, placing the miRNA crude product in a brand-new 2ml collecting tube, centrifuging the miRNA crude product for 5min again at room temperature by using the centrifugal force of 14000 Xg, abandoning the 2ml collecting tube, transferring the centrifuged centrifugal column filter tube into the miRNA collecting tube, adding 14 mu l of RNase-free water until the whole filter membrane of the centrifugal column filter tube is immersed in the solution, centrifuging the miRNA crude product for 1min at room temperature by using the centrifugal force of 14000 Xg, and collecting the miRNA sample;
s4, synthesizing 3.75 mul of the obtained miRNA into cDNA by utilizing a Takara Mir-XTM miRNA First-Strand Synthesis (Cat. number 638313) kit, selecting miRNA marker primers of hsa-miR-16-2-3p, hsa-miR-92b-5p, hsa-miR-130a-3p and has-miR-9024-3p, mixing the primers with 2 mul of the obtained cDNA, carrying out a PCR amplification system and a PCR amplification program, detecting the content of the amplified miRNA, and obtaining the content data of the miRNA;
s5 construction of C from the miRNA content data, as shown in FIGS. 1 to 4TDetermining the content of different miRNAs at different periods by a content curve,as can be seen from fig. 1, the differences of the four mirnas at 4 time points, C, when LH =5, i.e. day 5 of luteal phase transformationTThe lower level, i.e. the highest level of miRNA, indicated by a "p" value < 0.05. .
In order to further achieve the purpose of rapidness and accuracy, the hierarchical filtering is as follows:
a. adding 700 mu L Buffer BWT into a centrifugal column filter tube, then adopting 8000 Xg centrifugal force, centrifuging for 15s at room temperature, and abandoning the filtrate;
b. adding 500 μ L Buffer RPE into a centrifugal column filter tube, centrifuging at room temperature for 15s with 8000 Xg centrifugal force, and discarding the filtrate;
c. adding 500 μ L of 80% ethanol into a centrifugal column filter tube, centrifuging at room temperature for 2min by adopting a centrifugal force of 8000 Xg, and discarding the filtrate to finish the fractional filtration;
through carrying out multistage filtration, messenger's miRNA in the sample can fully purify to prevent the interference of other DNA, thereby realize quick accurate purpose.
In order to further realize the purpose of rapidness and accuracy, the specific process of synthesizing the cDNA by the miRNA is that in the kit, 3.75 mu L of miRNA sample, 5 mu L of mRQ Buffer (2 x) and 1.25 mu L of mRQ Enzyme are added into 200 mu L of RNase-free EP tube, after the mixture is mixed uniformly, the EP tube is heated to 37 ℃, the temperature is kept for 1h, then heated to 85 ℃, the temperature is kept for 5min, and then 90 mu L of ddH is added2O is subjected to constant volume to 100 mu L to obtain the cDNA solution; by reverse transcribing miRNA into complementary DNA, PCR amplification is directly carried out by using complementary strands to obtain a large number of samples, thereby achieving the purpose of rapidness and accuracy.
In order to further realize the purpose of rapidness and accuracy, the PCR amplification system is as follows: 2 μ L of cDNA solution, 0.5 μ L of miRNA marker primers, 0.5 μ L of 10 μmol/L mRQ 3' Primer, 12.5 μ L of TB Green Advantage Premix (2X), 0.5 μ L of ROX Dye (50X) and 9 μ L of ddH2O；
The amplification procedure comprises pre-denaturation at 95 ℃ for 10min, 40 cycles of denaturation at 95 ℃ for 15s and annealing at 60 ℃ for 30s, and preparation of a dissolution reaction curve at 95 ℃ for 15s, 60 ℃ for 60s and 95 ℃ for 15 s; the quantity of the cDNA is amplified in large quantity by limiting a PCR amplification system and a PCR amplification program, so that the whole quantity of the amplified cDNA is conveniently measured, the content of miRNA is indirectly deduced, and the aim of rapidness and accuracy is fulfilled.
Example 2
By finally determining different miRNA content data in different periods, constructing an ROC curve of a combination of four miRNA markers by multiple regression analysis, wherein the maximum area of the combination of the four miRNAs (hsa-miR-16-2-3 p, hsa-miR-92b-5p, hsa-miR-130a-3p and has-miR-9024-3 p) under the ROC curve (AUC) is shown in figure 5, the AUC value of the curve in the figure is 0.857, the limit value is 2.51, the sensitivity is 83.33, the specificity is 85.71, and the curve is deduced to obtain a prediction index:
prediction index =0.4167 (miR-16-2-3p) CT value + 0.1651 (miR-92b-5p) CT value + 0.3273 (miR-130a-3p) CT value + 0.0909 (miR-9025-3p) CT value.
The acceptability of the whole sample is predicted through the prediction index, the sample is detected on the day of LH =5, 200 samples are randomly extracted, and the accuracy can reach 85% compared with actual detection data, so that the superiority of the method is clarified.
The foregoing is illustrative of the preferred embodiments of this invention, and it is to be understood that the invention is not limited to the precise form disclosed herein and that various other combinations, modifications, and environments may be resorted to, falling within the scope of the concept as disclosed herein, either as described above or as apparent to those skilled in the relevant art. And that modifications and variations may be effected by those skilled in the art without departing from the spirit and scope of the invention as defined by the appended claims.
SEQUENCE LISTING
<110> Chengdu West darling gynecologic hospital limited company
<120> miRNA marker related to auxiliary diagnosis of endometrial receptivity and application thereof
<130> 2021.02.07
<160> 8
<170> PatentIn version 3.3
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Claims (4)
1. The miRNA marker for auxiliary diagnosis of endometrial receptivity is characterized in that: the miRNA marker consists of hsa-miR-16-2-3p, hsa-miR-92b-5p, hsa-miR-130a-3p, has-miR-9024-3p and primers thereof;
the sequence of the hsa-miR-16-2-3p is 5'-CCAAUAUUACUGUGCUGCUUUA-3'; the sequence of the hsa-miR-92b-5p is 5'-AGGGACGGGACGCGGUGCAGUG-3'; the sequence of the hsa-miR-130a-3p is 5'-CAGUGCAAUGUUAAAAGGGCAU-3'; the sequence of the has-miR-9024-3p is 5'-AGGCUUGGCUCUGAGGGU-3';
the sequence of the primer of the hsa-miR-16-2-3p is 5'-CCAATATTACTGTGCTGCTTTA-3'; the sequence of the primer of the hsa-miR-92b-5p is 5'-AGGGACGGGACGCGGTGCAGTG-3'; the sequence of the primer of the hsa-miR-130a-3p is 5'-AGTGCAATGTTAAAAGGGCAT-3'; the sequence of the primer of has-miR-9024-3p is 5'-AGGCTTGGCTCTGAGGGTAA-3'.
2. The use of miRNA markers for aiding in the diagnosis of endometrial receptivity as claimed in claim 1, wherein: the miRNA marker is applied to the preparation of an auxiliary diagnostic kit for endometrial receptivity.
3. The use of miRNA markers for aiding in the diagnosis of endometrial receptivity as claimed in claim 2, wherein: the diagnostic kit is used for detecting serum/plasma miRNA markers related to auxiliary diagnosis of endometrial receptivity.
4. The use of miRNA markers for aiding in the diagnosis of endometrial receptivity as claimed in claim 2, wherein: the kit also comprises enzymes and reagents commonly used in PCR reaction.
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| US20210002698A1 (en) * | 2019-07-02 | 2021-01-07 | Quark Biosciences Taiwan, Inc. | miRNA Receptivity Analysis of the Endometrium |
| CN110564852A (en) * | 2019-08-06 | 2019-12-13 | 中国医学科学院血液病医院(中国医学科学院血液学研究所) | MiRNAs expression map model related to human multiple myeloma, construction method and application |
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