Nucleic Acid Detection Composition, Detection Device and Method for Detecting Group B Streptococcus

Abstract

The present invention discloses a nucleic acid detection composition, a detection device and a method for detecting group B streptococcus. The composition includes an upstream primer, a downstream primer for thermally amplifying a target gene on the cAMP gene of GBS at a constant temperature, and a probe for detecting the target gene. The sequence of the upstream primer is as shown in SEQ ID NO.1, and the sequence of the downstream primer is as shown in SEQ ID NO.2; the sequence of the probe is as shown in SEQ ID NO.3. The present invention adopts a method combining a multi-enzyme rapid constant temperature amplification technology and an immunochromatographic test strip. Primers and probes are designed for the specific gene sequence of group B streptococcus, and modifications are made on the primers and probes, so that the amplification product can be captured and colored at the T line when flowing to the test strip T line; the whole process only requires manual sample addition and button pressing, the operation is simple and the result is visual, does not depend on detection equipment, environment and personnel, and the result can be easily detected and read.

Description

Technical Field

[0001] The present invention relates to the field of nucleic acid molecule detection, and particularly to a nucleic acid detection composition, a detection device and a method for detecting group B streptococcus. Background Art

[0002] Group B streptococcus (GBS) is a gram-positive coccus colonizing the human reproductive tract and gastrointestinal tract, and partially colonizing the upper respiratory tract of children and adults, threatening the health of pregnant women and neonates. GBS infection is a major risk factor for early neonatal infection. Approximately 50% of GBS-positive pregnant women will vertically transmit it to neonates, and 1% - 2% will develop early-onset GBS infection in neonates, which usually causes sepsis, pneumonia and meningitis, seriously threatening their lives, with a mortality rate as high as 5%. Therefore, before delivery, it is necessary to screen pregnant women in the third trimester of pregnancy for group B streptococcus, and provide targeted prevention and treatment measures according to the screening results, so as to minimize the incidence of premature rupture of membranes, preterm birth and perinatal mortality, and further minimize the harm of maternal and child reproductive tract infections and improve the survival quality of mother and child.

[0003] Molecular point-of-care testing technology (molecular POCT), as a highly sensitive in vitro diagnostic method, has a history of more than ten years. With the continuous progress of multiple technical fields such as nucleic acid isothermal amplification, molecular informatics, microfluidic technology, and remote communication, molecular POCT technology can be used to quickly detect and obtain results, and high-performance nucleic acid detection can also be achieved by ordinary people outside the hospital. It can be applied to multiple scenarios such as home testing, group testing, and in-hospital testing.

[0004] As a simplified nucleic acid amplification method, nucleic acid direct amplification technology can directly amplify using crude samples without nucleic acid extraction and purification steps. Rapid isothermal amplification refers to an amplification method with a constant temperature and can judge the result within 30 minutes. Its simplicity is compatible with nucleic acid direct amplification technology. The combination of the two not only eliminates the need for cumbersome sample processing steps, greatly saving time costs, but also can greatly reduce the structure of the detection device without complex extraction processes and control structures. Patent CN107164493A provides a GBS nucleic acid detection kit that can be used to complete the entire GBS nucleic acid detection process. It can not only get rid of the dependence on a large number of devices for GBS nucleic acid detection, improve the detection efficiency, but also avoid introducing contamination during the detection process and improve the detection accuracy. However, it has deficiencies such as low convenience, complex methods, and high costs. Summary of the Invention

[0005] The technical problem to be solved by the present invention is to provide a nucleic acid detection composition, a detection device and a method for detecting group B streptococcus in view of the above deficiencies in the prior art.

[0006] To solve the above technical problems, the technical solution adopted by the present invention is as follows: In the first aspect of the present invention, a nucleic acid detection composition for detecting group B streptococcus is provided, which includes an upstream primer, a downstream primer for thermally amplifying a target gene on the cAMP gene of GBS, and a probe for detecting the target gene. The sequence of the upstream primer is as shown in SEQ ID NO.1, and the sequence of the downstream primer is as shown in SEQ ID NO.2;

[0007] Preferably, the upstream and downstream primers are each 0.1 - 0.5 μM, and the probe is 0.05 - 0.25 μM.

[0008] In the sequence of the probe, a THF is labeled at the middle position 30 - 35 nt away from the 5' end of the probe as the recognition site for exonuclease. A fluorescein FAM is labeled at the 5' end of the probe, and a C3spacer is labeled at the 3' end of the probe.

[0009] Preferably, the sequence of the probe is as shown in SEQ ID NO.3.

[0010] Preferably, the sequence of the target gene is as shown in SEQ ID NO.4.

[0011] Preferably, the nucleic acid detection composition for detecting group B streptococcus further includes an amplification reaction auxiliary reagent for thermally amplifying, and the amplification reaction auxiliary reagent includes 200 - 400 ng / μL recombinase, 200 - 400 ng / μL recombinase cofactor, 500 - 1000 ng / μL single-stranded DNA binding protein, 100 - 200 ng / μL DNA polymerase, and 200 - 300 ng / μL creatine kinase.

[0012] Preferably, the nucleic acid detection composition for detecting group B streptococcus further includes a dilution buffer for diluting the nucleic acid amplification product, and the dilution buffer includes 50 - 150 mM Tris-HCl and 10 - 100 mM EDTA. The volume of the dilution buffer is 200 - 300 μL.

[0013] Preferably, the nucleic acid detection composition for detecting group B streptococcus further includes a lysis solution for lysing the sample, and the lysis solution includes 50 - 100 mM Tris-HCl, 50 - 80 mM EDTA, 2 wt% - 5 wt% Tween, 0.5 - 2 mM guanidine hydrochloride, 0.5 - 2 mM guanidine isothiocyanate, 0.5 - 1 M NaCl, 2 wt% - 5 wt% sodium dodecyl sulfate, 2 - 5 wt% sodium lauroyl sarcosinate, and deionized water. The volume of the lysis solution is 100 - 150 μL.

[0014] Preferably, the nucleic acid detection composition for detecting group B streptococcus further comprises a lysate for assisting isothermal amplification. The lysate comprises 30 - 50 mM Tris-HCl, 50 - 100 mM potassium acetate, 50 - 100 mM magnesium acetate, 50 - 100 mM creatine phosphate, 1 - 5 mM tris(2-carboxyethyl)phosphine hydrochloride, 100 - 500 μM dNTPs, 1 - 5 mM ATP, 8% - 10% w / v polyethylene glycol 10000, and 1 - 5% w / v polyvinylpyrrolidone. The volume of the lysate is 100 - 200 μL.

[0015] In the second aspect of the present invention, there is provided a nucleic acid detection device for detecting GBS, which realizes the detection of GBS through isothermal amplification by using the composition as described above. The detection device comprises a top cover, a functional layer, and a base that are fitted and arranged in sequence from top to bottom;

[0016] The top cover is provided with a pressing structure, an observation window, a sample addition port, and a sealing cover that cooperates with the sample addition port;

[0017] The functional layer is provided with a liquid storage chamber, a sample addition chamber, an amplification chamber, a buffer chamber, and a detection chamber that are connected in sequence. A heat conducting sheet is arranged at the bottom of the functional layer. The amplification reaction auxiliary reagent is made into a freeze-dried reagent and stored in the amplification chamber;

[0018] The base is provided with a flexible heating film and a heating and temperature control circuit board. The flexible heating film is located directly below the heat conducting sheet.

[0019] Preferably, the internal volume of the amplification chamber is 50 - 150 μL, and more preferably 50 μL.

[0020] Preferably, after the top cover is connected to the functional layer, the sample addition port is communicated with the sample addition chamber through a diversion groove, and the upper part of the amplification chamber is communicated with the buffer chamber through an overflow channel;

[0021] A nucleic acid test strip is arranged in the detection chamber, and the observation window is located above the nucleic acid test strip.

[0022] Preferably, the pressing structure comprises a pressing column with an upper limiting edge at the upper end, a spring sleeved on the pressing column and located between the upper limiting edge and the functional layer, and a lower limiting edge arranged at the bottom of the pressing column and extending into the liquid storage chamber;

[0023] A liquid storage vesicle is arranged in the liquid storage chamber directly below the lower limiting edge of the pressing structure. The liquid storage vesicle stores the dilution buffer solution. The liquid storage vesicle can be ruptured under the extrusion of the pressing structure, so that the stored dilution buffer solution flows out, and successively flows through the sample addition chamber, the amplification chamber, the buffer chamber, and the detection chamber.

[0024] In the third aspect of the present invention, a nucleic acid detection method for detecting GBS is provided. The detection of GBS is achieved by isothermal amplification using the device described above. The method includes the following steps:

[0025] S1. Insert the swab after sampling into the lysis solution for elution and lysis of the sample to be tested;

[0026] S2. Drop the lysis mixture obtained in step S1 into the sample addition port, then drop the dissolution solution into the sample addition port, and seal the sample addition port with the sealing cap;

[0027] S3. Connect the power supply, and keep the amplification chamber at a constant temperature for 10 - 30 min by the flexible heating film, and control the heating temperature at 37 - 42 °C;

[0028] S4. Apply a downward extrusion force to the pressing column of the pressing structure to rupture the liquid storage vesicle and release the dilution buffer solution therein. Under the flow of the dilution buffer solution, the amplification product in the amplification chamber is transferred to the nucleic acid test strip in the detection chamber;

[0029] S5. After 5 - 10 min, observe the T line and C line on the nucleic acid test strip through the detection window. When both the T line and C line show color, it is GBS positive; when the T line does not show color and the C line shows color, it is GBS negative.

[0030] Preferably, the rapid isothermal amplification method can be MIRA, RPA, RAA, ERA, etc., and more preferably MIRA.

[0031] The beneficial effects of the present invention are as follows:

[0032] The present invention adopts a method combining multi - enzyme rapid isothermal amplification technology and immunochromatographic test strip. Primers and probes are designed for the specific gene sequence of group B streptococcus, and modifications are made on the primers and probes so that the amplification product can be captured and show color at the T line of the flow - through test strip; the whole process only requires manual sample addition and button pressing, with simple operation and visual results, not relying on detection equipment, environment and personnel, and the results can be easily detected and read;

[0033] The GBS nucleic acid detection device of the present invention can independently complete the entire GBS detection process, without the need for additional equipment and complex manual intervention, and avoids secondary pollution formed during the operation process. The overall detection time is short, the detection results are easy to interpret, the operation method is simple, the device is portable, and it is suitable for home self - detection of GBS; the GBS nucleic acid detection method of the present invention has high efficiency, high specificity and high sensitivity, and the detection accuracy is relatively high;

[0034] In the detection device of the present invention, after the sample is roughly lysed by the lysis solution and mixed with the dissolution solution, the mixture is added to the sample loading port. Under the action of the diversion groove, the sample solution is diverted to the amplification chamber, and the excess sample enters the buffer chamber; in the amplification chamber, the sample can achieve uniform constant-temperature amplification through the temperature control chip, flexible heating film and heat conduction sheet at the bottom. After the amplification is completed, by pressing the pressing structure on the top of the liquid storage chamber to break the liquid storage bladder, the pre-stored dilution buffer solution is squeezed and released and then flows into the amplification chamber. After that, the amplification product is diluted and after filling the buffer chamber, the finally diluted amplification product will be diverted through the flow channel to the nucleic acid test strip for lateral chromatography, and the detection result can be obtained by observing the C / T line through the observation window. Description of the Drawings

[0035] Figure 1 It is a schematic exploded view of the nucleic acid detection device for detecting GBS in Embodiment 2 of the present invention;

[0036] Figure 2 It is a cross-sectional view of the nucleic acid detection device for detecting GBS in Embodiment 2 of the present invention;

[0037] Figure 3 It is a schematic comparison diagram of the detection results of positive samples (left) and negative samples (right) in the present invention;

[0038] Figure 4 It is the detection results of samples with different concentrations in the test example of the present invention;

[0039] Figure 5 It is the gray scale value of the T line corresponding to different sample concentrations in the test example of the present invention;

[0040] Figure 6 It is the gray scale value of the C line corresponding to different sample concentrations in the test example of the present invention.

[0041] Description of the Reference Numerals:

[0042] 1 - top cover, 2 - functional layer, 3 - base, 4 - liquid storage chamber, 41 - liquid storage bladder, 42 - pressing structure, 420 - pressing column, 421 - upper limit edge, 422 - spring, 423 - lower limit edge, 5 - sample loading chamber, 51 - diversion groove, 52 - sealing cover, 6 - amplification chamber, 61 - overflow channel, 7 - buffer chamber, 8 - detection chamber, 9 - observation window, 10 - heat conduction sheet, 11 - flexible heating film, 12 - heating temperature control circuit board. Detailed Description of the Embodiment

[0043] The following further elaborates on the present invention in conjunction with the embodiments, so that those skilled in the art can implement it with reference to the text of the specification.

[0044] It should be understood that terms such as "having", "comprising", and "including" as used herein do not exclude the presence or addition of one or more other elements or combinations thereof.

[0045] Unless otherwise specified, the test methods used in the following examples are all conventional methods. The materials and reagents used in the following examples can all be obtained commercially unless otherwise specified. For those in the following examples without specific conditions indicated, they are carried out according to conventional conditions or the conditions recommended by the manufacturer. For reagents or instruments without the manufacturer indicated, they are all conventional products that can be obtained through commercial purchase.

[0046] Example 1

[0047] A nucleic acid detection composition for detecting group B streptococcus, comprising an upstream primer, a downstream primer for thermostatic amplification of a target gene in the cAMP gene of GBS, and a probe for detecting the target gene, with the upstream and downstream primers each at 0.4 μM and the probe at 0.1 μM.

[0048] The sequence of the upstream primer is as shown in SEQ ID NO.1, and the sequence of the downstream primer is as shown in SEQ ID NO.2.

[0049] In the sequence of the probe, a THF is labeled at the middle position 30 - 35 nt away from the 5'-end of the probe as the recognition site for exonuclease, a fluorescein FAM is labeled at the 5'-end of the probe, and a C3spacer is labeled at the 3'-end of the probe. Specifically, the sequence of the probe is as shown in SEQ ID NO.3.

[0050] The sequence of the cAMP gene is as shown in SEQ ID NO.4, with a size of 246 bp.

[0051] The specific sequences of SEQ ID NO.1 - SEQ ID NO.4 are shown in Table 1 below:

[0052] Table 1

[0053]

[0054] In this example, the nucleic acid detection composition for detecting group B streptococcus further comprises auxiliary reagents for amplification reaction for thermostatic amplification. The auxiliary reagents for amplification reaction include 300 ng / μL recombinase, 300 ng / μL recombinase cofactor, 600 ng / μL single-stranded DNA binding protein, 200 ng / μL DNA polymerase, and 200 ng / μL creatine kinase.

[0055] In this embodiment, the nucleic acid detection composition for detecting group B streptococcus further includes a dilution buffer for diluting nucleic acid amplification products. The dilution buffer includes 100 mM Tris-HCl and 50 mM EDTA, with a total volume of 250 μL.

[0056] In this embodiment, the nucleic acid detection composition for detecting group B streptococcus further includes a lysis solution for lysing samples. The lysis solution includes 100 mM Tris-HCl, 50 mM EDTA, 5 wt% (mass concentration) Tween, 0.5 mM guanidine hydrochloride, 1 mM guanidine isothiocyanate, 0.5 M NaCl, 2 wt% sodium dodecyl sulfate, 2 wt% sodium lauroyl sarcosinate, and deionized water; the volume of the lysis solution is 100 μL.

[0057] In this embodiment, the nucleic acid detection composition for detecting group B streptococcus further includes a dissolution solution for assisting in isothermal amplification. The dissolution solution includes 50 mM Tris-HCl, 50 mM potassium acetate, 50 mM magnesium acetate, 50 mM creatine phosphate, 2 mM tris(2-carboxyethyl)phosphine hydrochloride, 200 μM dNTPs, 2 mM ATP, 8% w / v (mass-volume concentration) polyethylene glycol 10000, and 2% w / v polyvinylpyrrolidone; the volume of the dissolution solution is 150 μL.

[0058] In this embodiment, the lysis solution and the dissolution solution are pre-sealed in separate lysis solution tubes and dissolution solution tubes. The dissolution solution tubes and lysis solution tubes are disposable plastic tubes with sealing films and droppers.

[0059] Example 2

[0060] Refer to Figure 1 and Figure 2 A nucleic acid detection device for detecting GBS uses the composition of Example 1 to achieve the detection of GBS through isothermal amplification. The detection device includes a top cover 1, a functional layer 2, and a base 3 that are sequentially fitted from top to bottom;

[0061] The top cover 1 is provided with a pressing structure 42, an observation window 9, a sample addition port, and a sealing cover 52 that cooperates with the sample addition port;

[0062] The functional layer 2 is provided with a liquid storage chamber 4, a sample addition chamber 5, an amplification chamber 6, a buffer chamber 7, and a detection chamber 8 that are sequentially connected. A heat conducting sheet 10 is provided at the bottom of the functional layer 2. The amplification reaction auxiliary reagent is made into a freeze-dried reagent and stored in the amplification chamber 6;

[0063] The base 3 is provided with a flexible heating film 11 and a heating temperature control circuit board 12. The flexible heating film 11 is located directly below the heat conducting sheet 10.

[0064] In this embodiment, after the top cover 1 is connected to the functional layer 2, the sample loading port is communicated with the sample loading chamber 5 through the diversion groove 51, and the upper part of the amplification chamber 6 is communicated with the buffer chamber 7 through the overflow channel 61; the overflow channel 61 is U-shaped. After sample loading, the solution can fill the amplification chamber 6, and the excess solution enters the buffer chamber 7 and stays therein. After amplification is completed, the dilution buffer dilutes the amplified solution in the amplification chamber 6 and gradually fills the buffer chamber 7, and finally enters the detection chamber 8.

[0065] A nucleic acid test strip is arranged in the detection chamber 8, and the observation window 9 is located above the nucleic acid test strip. The above-mentioned nucleic acid test strip is a colloidal gold immunochromatographic test strip capable of detecting nucleic acid amplification products. The binding pad of the test strip is coated with colloidal gold particles conjugated with FTIC antibodies, the T line is loaded with streptavidin, and the C line is loaded with goat anti-mouse antibodies. When there is a positive target gene in the sample, one end of the DNA double strand amplified by the specially modified primers and probes has fluorescein and the other end has biotin. When the amplified product flows to the binding pad, the fluorescein on the double strand will bind to the colloidal gold conjugated with FTIC antibodies and continue to chromatograph forward. When it chromatographs to the T line, the biotin on the double strand will bind to streptavidin and fix the colloidal gold at the T line for color development. The excess colloidal gold conjugated with FTIC antibodies continues to chromatograph forward and binds to the polyclonal antibody on the C line for color development. When there is no target gene in the sample, the solution will carry the colloidal gold conjugated with FTIC antibodies on the binding pad to chromatograph forward, and the polyclonal antibody bound at the C line will be captured for color development.

[0066] In this embodiment, the pressing structure 42 includes a pressing column 420 with an upper limit edge 421 at the upper end, a spring 422 sleeved on the pressing column 420 and located between the upper limit edge 421 and the functional layer 2, and a lower limit edge 423 arranged at the bottom of the pressing column 420 and extending into the liquid storage chamber 4;

[0067] A liquid storage vesicle 41 is arranged in the liquid storage chamber 4 directly below the lower limit edge 423 of the pressing structure 42. The liquid storage vesicle 41 stores dilution buffer. The liquid storage vesicle 41 can be ruptured under the extrusion of the pressing structure 42 so that the stored dilution buffer flows out, and successively flows through the sample loading chamber 5, the amplification chamber 6, the buffer chamber 7 and the detection chamber 8. The above-mentioned liquid storage vesicle 41 can optionally use easily squeezable materials such as pierceable plastic films or tinfoil or easily extrudable materials such as cotton and sponge to encapsulate and pre-store the buffer. In this embodiment, a plastic film is used.

[0068] Embodiment 3

[0069] A nucleic acid detection method for detecting GBS uses the device of Embodiment 2 to achieve the detection of GBS through isothermal amplification. The method includes the following steps:

[0070] S1. Insert the swab after sampling into the lysis solution to wash and lyse the sample to be tested;

[0071] S2. Drop the lysis mixture obtained in step S1 into the sample inlet, then drop the dissolution solution into the sample inlet, and seal the sample inlet with the sealing cap 52;

[0072] S3. Connect the power supply, and keep the amplification chamber 6 at a constant temperature of 39 °C for 15 min by the flexible heating film 11;

[0073] S4. Apply a downward extrusion force to the pressing column 420 of the pressing structure 42 to rupture the liquid storage vesicle 41 and release the dilution buffer therein. Under the flow of the dilution buffer, the amplification product in the amplification chamber 6 is transferred to the nucleic acid test strip in the detection chamber 8;

[0074] S5. After 5 min, observe the T line and C line on the nucleic acid test strip through the detection window. When both the T line and C line show color, it is GBS positive; when the T line does not show color and the C line shows color, it is GBS negative.

[0075] The binding pad of the test strip is coated with colloidal gold particles conjugated with FTIC antibody. The T line is loaded with streptavidin, and the C line is loaded with goat anti-mouse antibody. After amplification of the positive sample, a large amount of modified double-stranded DNA is produced. When the colloidal gold combined with it flows through the T line, it is captured, and the excess colloidal gold continues to chromatograph forward to the C line and is captured, so that both the T line and C line show color; the negative sample cannot be amplified. After the probe molecule (single-stranded DNA) binds to the colloidal gold, it chromatographs forward and is captured at the C line, making the C line show color while the T line does not show color. Refer to Figure 3 , which is a schematic diagram of the comparison of the detection results of the positive sample (left) and the negative sample (right).

[0076] Test Example

[0077] The GBS positive plasmid with a known concentration (5 ng / μL) was diluted 10, 100, 1000, and 10000 times respectively, used as samples, and detected by the method of Example 3. The detection results are as shown in the appendix Figure 4 As shown, from left to right in the figure are the results of the above-mentioned positive plasmids diluted 10000 times, 1000 times, 100 times, 10 times, and undiluted. It can be seen that when the dilution factor is within 1000 times, the T line shows obvious color, and when the dilution factor reaches 10000 times, the T line does not show obvious color.

[0078] Refer to Figure 5 , which is the gray value of the T line corresponding to different sample concentrations. The sample numbers therein indicate:

[0079] 1: GBS positive plasmid diluted 10000 times, 2: diluted 1000 times, 3: diluted 100 times, 4: diluted 10 times, 5: undiluted GBS positive plasmid (5 ng / μL). It can be seen that the fitted curve has good linearity.

[0080] Refer toFigure 6 , which is the gray value of the C line corresponding to different sample concentrations.

[0081] Although the embodiments of the present invention have been disclosed above, they are not limited to the applications listed in the specification and embodiments. It can be fully applied to various fields suitable for the present invention. For those familiar with the field, additional modifications can be easily made. Therefore, without departing from the general concept defined by the claims and the equivalent scope, the present invention is not limited to specific details.

Claims

1. A nucleic acid detection device for detecting GBS, characterized in that, it uses the following nucleic acid detection composition to achieve the detection of GBS through isothermal amplification. The nucleic acid detection composition includes an upstream primer, a downstream primer for the target gene on the cAMP gene for isothermal amplification of GBS, and a probe for detecting the target gene on the cAMP gene. The sequence of the upstream primer is as shown in SEQ ID NO.1, and the sequence of the downstream primer is as shown in SEQ ID NO.2; in the sequence of the probe, a THF is marked at the middle position 30-35 nt away from the 5' end of the probe as the recognition site for exonuclease. A fluorescein FAM is marked at the 5' end of the probe, and a C3 spacer is marked at the 3' end of the probe; the sequence of the probe is as shown in SEQ ID NO.3; the sequence of the target gene is as shown in SEQ ID NO.4; the nucleic acid detection composition further includes amplification reaction auxiliary reagents for isothermal amplification. The amplification reaction auxiliary reagents include 200-400 ng / μL recombinase, 200-400 ng / μL recombinase cofactor, 500-1000 ng / μL single-stranded DNA binding protein, 100-200 ng / μL DNA polymerase, and 200-300 ng / μL creatine kinase; the nucleic acid detection composition further includes a lysis solution for lysing the sample. The lysis solution includes 50-100 mM Tris-HCl, 50-80 mM EDTA, 2 wt%-5 wt% Tween, 0.5-2 mM guanidine hydrochloride, 0.5-2 mM guanidine isothiocyanate, 0.5-1 M NaCl, 2 wt%-5 wt% sodium dodecyl sulfate, 2-5 wt% sodium lauroyl sarcosinate, and deionized water; the nucleic acid detection composition further includes a dissolution solution for assisting isothermal amplification. The dissolution solution includes 30-50 mM Tris-HCl, 50-100 mM potassium acetate, 50-100 mM magnesium acetate, 50-100 mM creatine phosphate, 1-5 mM tris(2-carboxyethyl)phosphine hydrochloride, 100-500 μM dNTPs, 1-5 mM ATP, 8%-10% w / v polyethylene glycol 10000, and 1-5% w / v polyvinylpyrrolidone; the detection device includes a top cover, a functional layer, and a base that are sequentially fitted from top to bottom; a pressing structure, an observation window, a sample addition port, and a sealing cover that cooperates with the sample addition port are provided on the top cover; a liquid storage chamber, a sample addition chamber, an amplification chamber, a buffer chamber, and a detection chamber that are sequentially connected are provided on the functional layer. A heat conducting sheet is provided at the bottom of the functional layer. The amplification reaction auxiliary reagents are made into freeze-dried reagents and stored in the amplification chamber; a flexible heating film and a heating and temperature control circuit board are provided on the base. The flexible heating film is located directly below the heat conducting sheet; a nucleic acid test strip is provided in the detection chamber.

2. The nucleic acid detection device for detecting GBS according to claim 1, characterized in that, The described nucleic acid detection composition further includes a dilution buffer for diluting nucleic acid amplification products, and the dilution buffer includes 50 - 150 mM Tris-HCl and 10 - 100 mM EDTA.

3. The nucleic acid detection device for detecting GBS according to claim 2, wherein, after the top cover is connected to the functional layer, the sample addition port is communicated with the sample addition chamber through a diversion groove, and the upper part of the amplification chamber is communicated with the buffer chamber through an overflow channel; The observation window is located above the nucleic acid test strip.

4. The nucleic acid detection device for detecting GBS according to claim 3, wherein, The pressing structure includes a pressing column with an upper limit edge at the upper end, a spring sleeved on the pressing column and located between the upper limit edge and the functional layer, and a lower limit edge provided at the bottom of the pressing column and extending into the liquid storage chamber; A liquid storage vesicle is arranged in the liquid storage chamber directly below the lower limit edge of the pressing structure. The liquid storage vesicle stores the dilution buffer. The liquid storage vesicle can be ruptured under the extrusion of the pressing structure, so that the internally stored dilution buffer flows out, and successively flows through the sample addition chamber, the amplification chamber, the buffer chamber and the detection chamber.

5. The nucleic acid detection device for detecting GBS according to claim 4, wherein, The method for the nucleic acid device to detect GBS includes the following steps: S1. Insert the swab after sampling into the lysis solution for eluting and lysing the sample to be tested; S2. Drop the lysate mixture obtained in step S1 into the sample addition port, then drop the dissolution solution into the sample addition port, and seal the sample addition port with a sealing cover; S3. Connect the power supply, and keep the temperature of the amplification chamber at a constant temperature of 37 - 42 °C for 10 - 30 min by the flexible heating film; S4. Apply a downward extrusion force to the pressing column of the pressing structure to rupture the liquid storage vesicle and release the dilution buffer therein. Under the flow of the dilution buffer, the amplification products in the amplification chamber are transferred onto the nucleic acid test strip in the detection chamber; S5. After 5 - 10 min, observe the T line and C line on the nucleic acid test strip through the detection window. When both the T line and the C line are colored, it is GBS positive. When the T line is not colored and the C line is colored, it is GBS negative.

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