A high-quality extraction method for biological evidence for on-site investigation

Abstract

The invention discloses a high-quality extraction method for biological evidence for on-site investigation, which belongs to the technical field of DNA collection and detection, and comprises the following steps: S1, squeezing an efficient DNA enricher to release the biological evidence collection reagent to the biological trace position of the evidence carrier through the swab of the efficient DNA enricher; wiping the biological trace position with a swab to accelerate the elution of the exfoliated cells and free DNA at the biological trace position; S2, loosening the efficient DNA enricher, sucking back the biological evidence collection reagent containing the exfoliated cells and free DNA; wiping the biological evidence collection reagent remaining on the evidence carrier with a dry cotton swab; S3, extracting and testing the DNA contained in the dry cotton swab, the swab and the biological evidence collection reagent to obtain a DNA typing result diagram. The method of the invention can fully and effectively elute and adsorb all the exfoliated cells and DNA on the surface of the evidence, which is conducive to improving the detection rate of DNA.

Description

Technical Field

[0001] The invention relates to the technical field of DNA collection and detection, and in particular to a high-quality extraction method of biological evidence for on-site investigation. Background Art

[0002] At present, in the extraction of DNA evidence, there are many ways to transfer and extract trace DNA retained on different carriers, such as wiping, sticking, shearing, negative pressure adsorption, and oscillation washing. However, in recent years, due to the influence of environmental factors such as the diversification of crime scenes, the sophisticated means of the perpetrators, and the unsatisfactory investigation conditions, as well as the influence of various objective factors of the evidence itself, such as the large workload of blind extraction, dents on the surface of the evidence, the degradation of the DNA retained by the object, and the poor preservation of the cotton swab after collection, the detection rate of DNA has been greatly reduced. For example, for some evidence with strong permeability or special shapes such as grooves or gaps, it is difficult to extract DNA through conventional adsorption, sticking, shearing or wiping, and the problem of DNA leakage extraction is prone to occur, so that satisfactory DNA test analysis results cannot be obtained. Specifically, for wiping extraction, a dry and wet two-step method is usually used for extraction, that is, first wipe the surface of the object of the test material with a wet cotton swab, and then wipe it dry with a dry cotton swab. The most commonly used reagent for wetting cotton swabs is deionized water. During the above-mentioned collection process, the operator needs to carry a tube of distilled water and two cotton swabs, and the cotton swabs need to be stored in a specific container. The cotton swabs and distilled water are carried separately, and the entire operation process is cumbersome. In addition, only a little distilled water is needed to soak the cotton swabs. Carrying a tube of distilled water may cause the distilled water to be contaminated if the operation is not careful, which is very likely to cause waste. At the same time, if the amount of water is too little, it will be difficult to collect all the biological evidence in the suspected parts of the specimen; and too much water will accelerate the degradation rate of cellular DNA in the biological evidence, increase the difficulty of later DNA extraction, and even lead to the situation where DNA cannot be detected. In addition, biological specimens extracted by wiping with deionized water are easily corrupted by microorganisms, resulting in the degradation of trace amounts of DNA therein.

[0003] Therefore, it is very necessary to develop a high-quality extraction method for biological evidence suitable for on-site investigation. Summary of the invention

[0004] In order to overcome the shortcomings of the prior art, the purpose of the present invention is to provide a high-quality extraction method for biological evidence for on-site investigation, which can fully and effectively elute and adsorb all exfoliated cells and DNA on the surface of the evidence, which is beneficial to improve the detection rate of DNA and improve the extraction quality.

[0005] To solve the above problems, the technical solution adopted by the present invention is as follows:

[0006] A method for high-quality extraction of biological evidence for on-site investigation comprises the following steps:

[0007] S1. Squeeze the high-efficiency DNA enricher to release the biological evidence collection reagent to the biological trace position of the evidence carrier through the swab of the high-efficiency DNA enricher; wipe the biological trace position with the swab to accelerate the elution of the detached cells and free DNA at the biological trace position;

[0008] S2. Release the high-efficiency DNA enricher and suck back the biological evidence collection reagent containing detached cells and free DNA; wipe the biological evidence collection reagent remaining on the evidence carrier with a dry cotton swab;

[0009] S3. Extract and test the DNA contained in the dry cotton swab, swab and biological evidence collection reagent to obtain a DNA typing result diagram.

[0010] As a preferred embodiment of the present invention, the biological evidence collection reagent is composed of the following components: 20-50 mM buffer salt, 0.2%-2% thickener, 0.1%-1% surfactant, 20-50 mM metal ion chelator, 0.1%-0.5% antibacterial agent.

[0011] As a preferred embodiment of the present invention, the thickener is at least one of carbomer, sodium alginate or sodium polyacrylate.

[0012] As a preferred embodiment of the present invention, the concentration of the buffer salt is 5-20 mmol / L, and the buffer salt is one of Tris-HCl, phosphate, NaCl, KCl or MgCl; the surfactant is one of sodium dodecyl sulfate, TritonX-100 or Tween-80; and the antibacterial agent is potassium sorbate or benzoic acid.

[0013] As a preferred embodiment of the present invention, the high-efficiency DNA enricher includes a connecting part, a connecting tube, a pipette tip, a swab, a limiting part and a sealing cover; the first end of the connecting tube is connected to the first end of the connecting part, and a first flow channel is provided in the connecting tube; the first end of the pipette tip is connected to the second end of the connecting part, and an extrusion cavity is provided inside the pipette tip, and the extrusion cavity is filled with a biological evidence collection reagent; the swab is connected to the second end of the connecting tube and is in communication with the first flow channel; the limiting part is located in the extrusion cavity, and the first end of the limiting part is connected to the second end of the connecting part, and the limiting The limiting portion can block the communication between the extrusion chamber and the first flow channel, and the limiting portion can be broken, and after the limiting portion is broken, the extrusion chamber and the first flow channel can be connected to each other; the sealing cover is sleeved on the outer circumference of the connecting tube and the swab, and the first end of the sealing cover is detachably connected to the first end of the connecting portion; wherein, after the limiting portion is broken, the biological evidence collection reagent in the extrusion chamber can be released into the swab by squeezing the suction head, and the biological evidence collection reagent can be recovered into the extrusion chamber through the swab under the action of the negative pressure generated by squeezing the suction head.

[0014] As a preferred embodiment of the present invention, a second flow channel is opened inside the limiting portion, a first end of the second flow channel is connected to the first flow channel, a second end of the second flow channel can be closed by the second end of the limiting portion, and after the limiting portion is broken, the second end of the second flow channel is connected to the extrusion chamber; a third flow channel is opened inside the connecting portion, and both ends of the third flow channel are respectively connected to the first flow channel and the second flow channel.

[0015] As a preferred embodiment of the present invention, the surface of the suction head is marked with scale lines.

[0016] As a preferred embodiment of the present invention, the suction head is made of soft material.

[0017] As a preferred embodiment of the present invention, the step S3 uses a magnetic bead method or a silica gel membrane method DNA extraction kit to extract the DNA contained in the dry cotton swab, the swab and the biological evidence collection reagent.

[0018] As a preferred embodiment of the present invention, the step S3 tests the extracted DNA according to the following steps: after the extracted DNA is subjected to Nanodrop quantitative detection, multi-color fluorescent labeling and PCR reaction multiple composite amplification are performed using a VFP fluorescence detection kit to detect the polymorphism of the STR loci in the DNA and identify individuals; and electrophoresis detection is performed using an ABI genetic analyzer.

[0019] Specifically, the electrophoresis detection includes the following steps: centrifuging the amplified product at a speed of 2000-5000rpm for 1-1min, taking 1-10μL of the product or EX25 Allelic Ladder and mixing it with 0.5-3μL AGCU Marker SI Z-500 and 1-12μL deionized formamide, denaturing at 50-120℃ for 1-10min, and then detecting by ice bath electrophoresis.

[0020] Compared with the prior art, the present invention has the following beneficial effects:

[0021] The high-quality extraction method for biological evidence of the present invention is based on the existing dry-wet two-step wiping method, combines an efficient DNA enricher with a suction head and utilizes the viscosity of the biological evidence collection reagent to wash out the exfoliated cells and DNA on the surface of the evidence carrier as much as possible, and effectively adsorbs and protects the exfoliated cells and DNA on the surface of the evidence carrier, thereby increasing the DNA extraction amount and greatly enhancing the extraction quality, thereby realizing the high-quality collection of on-site biological evidence; then, DNA testing is performed on the biological evidence collection reagent, swab and dry cotton swab adsorbed with the exfoliated cells and DNA, which is beneficial to improving the DNA detection rate, making up for the deficiency of missing DNA extraction for certain special evidence in traditional DNA extraction methods such as wiping, sticking, shearing, scraping, negative pressure adsorption, and oscillation washing, thereby achieving the purpose of high-quality collection of biological evidence on-site, having important promoting significance for enriching the efficient DNA extraction and transfer methods, being beneficial to increasing the probability of identifying criminal suspects, promoting the application of criminal technology and scientific and technological innovation, better serving actual combat, and providing direction for case detection. BRIEF DESCRIPTION OF THE DRAWINGS

[0022] Figure 1 A step diagram of a method for high-quality extraction of biological evidence according to an embodiment of the present invention;

[0023] Figure 2 This is a schematic diagram of the overall structure of a high-efficiency DNA enricher according to an embodiment of the present invention;

[0024] Figure 3 A cross-sectional view of a high-efficiency DNA enricher according to an embodiment of the present invention;

[0025] Figure 4 An exploded view of a high-efficiency DNA enricher according to an embodiment of the present invention;

[0026] Figure 5 The actual pictures of the extraction carrier paper skins of Example 1 and Comparative Example 1 of the present invention;

[0027] Figure 6 In order to adopt the extraction method of the present invention Figure 5 The test result diagram of swabbing and extraction of the carrier shown in FIG;

[0028] Figure 7 To use ordinary cotton swabs to conduct a two-step wet and dry method Figure 5 The test result diagram of swabbing and extraction of the carrier shown in FIG;

[0029] Figure 8 The figures are the physical pictures of the extraction carrier stones of Example 2 and Comparative Example 2 of the present invention;

[0030] Fig. 9 In order to adopt the extraction method of the present invention Figure 8 The test result diagram of swabbing and extraction of the carrier shown in FIG;

[0031] Fig.10 To use ordinary cotton swabs to conduct a two-step wet and dry method Figure 8 The test result diagram of swabbing and extraction of the carrier shown in FIG;

[0032] Description of the accompanying drawings: 100 connecting part, 110 third flow channel, 200 connecting tube, 210 first flow channel, 300 suction head, 310 extrusion chamber, 400 swab, 500 limiting part, 510 second flow channel, 600 sealing cover. DETAILED DESCRIPTION

[0033] The present invention is further described in detail below in conjunction with the accompanying drawings and specific embodiments.

[0034] Reference Figure 1 The method for extracting high-quality biological evidence for on-site investigation provided by the present invention comprises the following steps:

[0035] S1. Squeeze the high-efficiency DNA enricher to release the biological evidence collection reagent to the biological trace position of the evidence carrier through the swab of the high-efficiency DNA enricher; wipe the biological trace position with the swab to accelerate the elution of the detached cells and free DNA at the biological trace position;

[0036] S2. Release the high-efficiency DNA enricher and suck back the biological evidence collection reagent containing detached cells and free DNA; wipe the biological evidence collection reagent remaining on the evidence carrier with a dry cotton swab;

[0037] S3. Extract and test the DNA contained in the dry cotton swab, swab and biological evidence collection reagent to obtain a DNA typing result diagram.

[0038] It can be seen from the above method that the present invention, based on the existing dry-wet two-step wiping method, adopts a high-efficiency DNA enricher that pre-stores biological evidence collection reagents and has the function of squeezing and recovering reagents. The biological evidence collection reagents are released to the biological trace position, and the detached cells and DNA at the biological trace position are wiped and washed off. After the biological evidence collection reagents are sucked back, the residual biological evidence collection reagents are wiped with a dry cotton swab to extract all the detached cells and DNA on the surface of the evidence carrier as much as possible, thereby increasing the amount of DNA extraction, thereby achieving high-quality collection of on-site biological evidence, which is beneficial to improving the DNA detection rate.

[0039] In the above method, the biological evidence collection reagent has a certain viscosity, can effectively adsorb the exfoliated cells and DNA on the surface of the biological evidence, and can better preserve the exfoliated cells and DNA for a certain period of time. Specifically, the biological evidence collection reagent is composed of the following components in molar concentration or mass percentage: 20-50mM (molar concentration) buffer salt, 0.2%-2% (mass percentage) thickener, 0.1%-1% (mass percentage) surfactant, 20-50mM (molar concentration) metal ion chelator, 0.1%-0.5% (mass percentage) antibacterial agent. Among them, the buffer salt can maintain a stable pH value and can better preserve the structure of the collected exfoliated cells; the thickener can give the reagent an appropriate viscosity and can enhance the specific adsorption of the reagent to the organic component, which is conducive to the adsorption and recovery of the exfoliated cells; the surfactant can enhance the wettability and solubility of the reagent, improve the dissolution and recovery rate of the exfoliated cells adhered to the surface of the carrier, and can make the exfoliated cells adhered to some permeable carriers such as cardboard and cotton cloth fully dissolved and easy to recover; the metal ion chelator eliminates the adhesion phenomenon between the exfoliated cells and the carrier, improves the recovery rate, and protects the integrity of the DNA; the antibacterial agent can inhibit the reproduction of microorganisms during the storage of exfoliated cells and DNA samples, and can effectively extend the storage time of the collected samples. Preferably, the thickener is at least one of carbomer, sodium alginate or sodium polyacrylate. The concentration of the buffer salt is 5-20 mmol / L, and the buffer salt is one of Tris-HCl, phosphate, NaCl, KCl or MgCl; the surfactant is one of sodium dodecyl sulfate, Triton X-100 or Tween-80; and the antibacterial agent is potassium sorbate or benzoic acid.

[0040] In the above method, if Figures 2 to 4 As shown, the high-efficiency DNA enricher includes a connecting portion 100, a connecting tube 200, a pipette tip 300, a swab 400, a limiting portion 500 and a sealing cover 600. Figure 3 As shown, the limiting portion 500, the connecting portion 100, the connecting tube 200 and the swab 400 are connected in sequence.

[0041] Reference Figure 3 and Figure 4 The first end of the connecting tube 200 is connected to the first end of the connecting portion 100. The connecting tube 200 can be integrally formed with the connecting portion 100 or can be separately formed. For example, the first end of the connecting tube 200 can be detachably plugged into the first end of the connecting portion 100. Figure 3 A first flow channel 210 is opened in the connecting pipe 200 .

[0042] Reference Figures 2 to 4 , the first end of the suction tip 300 is connected to the second end of the connecting portion 100, and an extrusion cavity 310 is provided inside the suction tip 300, and the extrusion cavity 310 is filled with a biological evidence collection reagent. The portion of the suction tip 300 used for extrusion deformation is made of a flexible and elastic material, such as silicone, so that the suction tip 300 is easily deformed when squeezed, and the extrusion cavity 310 inside it can be deformed, so that the biological evidence collection reagent in the extrusion cavity 310 can be released. After the suction tip 300 is released, the suction tip 300 can be deformed outward and restored, and the negative pressure generated by squeezing the suction tip 300 is used to suck the biological evidence collection reagent back into the extrusion cavity 310 of the suction tip 300. Furthermore, the first end of the suction head 300 can be sleeved on the outer periphery of the second end of the connecting part 100, and the portion of the suction head 300 used for sleeve connection with the connecting part 100 can be made of materials such as PP or PE. The connection between the suction head 300 and the connecting part 100 can be fixed and sealed by glue after sleeve connection, or fixed and sealed by sleeve connection of a sealing ring and then threaded connection.

[0043] Reference Figure 3 and Figure 4 The swab 400 is connected to the second end of the connecting tube 200 and is connected to the first flow channel 210, that is, at least a part of the structure of the swab 400 is exposed at the outlet of the first flow channel 210 to absorb and release the biological evidence collection reagent. The swab 400 is a cotton swab head containing cotton or wrapped with a flocked swab, which can increase the friction of wiping, so that the exfoliated cells or DNA on the evidence carrier are more likely to fall off into the biological evidence reagent, which is convenient for recovery and preservation.

[0044] Continue to refer to Figure 3 and Figure 4, the limiting part 500 is located in the extrusion chamber 310, and the first end of the limiting part 500 is connected to the second end of the connecting part 100. During packaging, transportation and storage, the limiting part 500, as a stopper, can block the communication between the extrusion chamber 310 and the first flow channel 210, and can prevent the biological evidence collection reagent in the extrusion chamber 310 from entering the first flow channel 210 and flowing through the swab 400. The limiting part 500 is made of an easily broken material, and the limiting part 500 can be broken when an external force is applied. After the limiting part 500 is broken, the extrusion chamber 310 and the first flow channel 210 can be connected to each other. When the suction head 300 is squeezed, the biological evidence collection reagent in the extrusion chamber 310 can flow into the first flow channel 210 and then be released into the swab 400. After the suction head 300 is released, the biological evidence collection reagent can be recovered into the extrusion chamber 310 through the swab 400 and the first flow channel 210 under the negative pressure generated by squeezing the suction head 300. Furthermore, an annular indentation may be prefabricated on the outer periphery of the limiting portion 500 so that the limiting portion 500 can be broken along the annular indentation when force is applied.

[0045] Reference Figures 2 to 4 , the sealing cover 600 is sleeved on the outer circumference of the connecting tube 200 and the swab 400, and the first end of the sealing cover 600 is detachably connected to the first end of the connecting part 100. During packaging, transportation and storage, the sealing cover 600 can protect the connecting tube 200 and the swab 400, prevent the swab 400 from being contaminated by the outside world, and prevent the connecting tube 200 from being broken by external forces. When in use, the sealing cover 600 can be removed to expose the swab 400, and then the limiting part 500 is broken and the suction head 300 is squeezed, and the swab 400 is used to wipe the surface of the evidence. After wiping, the sealing cover 600 can be covered again to prevent contamination. Furthermore, the sealing cover 600 can be made of a hard material, and the connection between the sealing cover 600 and the connecting part 100 can be a threaded connection. Specifically, an internal thread can be opened at the first end of the sealing cover 600, and an external thread can be opened at the first end of the connecting part 100, so as to realize the threaded connection between the sealing cover 600 and the connecting part 100, so as to facilitate the installation and disassembly of the sealing cover 600.

[0046] In some embodiments, the connecting tube 200 can be integrally formed with the limiting portion 500, that is, the limiting portion 500 is integrally formed at the first end of the connecting tube 200. During installation, the limiting portion 500 at the first end of the connecting tube 200 can be inserted along the first end of the connecting portion 100 until the limiting portion 500 extends from the second end of the connecting portion 100 into the extrusion cavity 310. When the limiting portion 500 is broken, the first flow channel 210 in the connecting tube 200 can be interconnected with the extrusion cavity 310. In other embodiments, the connecting tube 200 and the limiting portion 500 are separately provided, and the two are respectively connected to the first end and the second end of the connecting portion 100. Both can be integrally formed with the connecting portion 100 or separately provided. For example Figure 3 As shown, the connecting tube 200 is connected to the first end of the connecting part 100, the limiting part 500 is connected to the second end of the connecting part 100, a second flow channel 510 is opened inside the limiting part 500, and a third flow channel 110 is opened inside the connecting part 100. The first end of the second flow channel 510 is connected with the first flow channel 210 through the third flow channel 110, and the second end of the second flow channel 510 can be closed by the second end of the limiting part 500. After the limiting part 500 is broken, the second end of the second flow channel 510 is connected with the extrusion cavity 310, so that the extrusion cavity 310, the second flow channel 510, the third flow channel 110 and the first flow channel 210 are connected in sequence.

[0047] Furthermore, the surface of the pipette tip 300 is marked with scale lines to facilitate the user to identify the reagent content in the enricher. The reagent content in each enricher is 0-5000u l, and the enricher is vacuum packed, sterilized with ethylene oxide, and stored at room temperature away from light to play an antibacterial and anti-pollution role.

[0048] In the above method, step S3 uses a magnetic bead method or a silica gel membrane method DNA extraction kit to extract the DNA contained in the dry cotton swab, swab and biological evidence collection reagent. Step S3 tests the extracted DNA according to the following steps: after the extracted DNA is subjected to Nanodrop quantitative detection, multi-color fluorescent labeling and PCR reaction multiple composite amplification are performed using a VFP fluorescence detection kit to detect the polymorphism of the STR loci in the human genomic DNA in the test material evidence and identify the individual; electrophoresis detection is performed using an ABI genetic analyzer. Specifically, the electrophoresis detection includes the following steps: centrifuging the amplified product at a speed of 2000-5000rpm for 1 to 1min, taking 1 to 10μL of the product or EX25 Allel ic Ladder and mixing it with 0.5 to 3μL AGCU MarkerSI Z-500 and 1 to 12μL deionized formamide, denaturing at 50 to 120℃ for 1 to 10min, and then performing ice bath electrophoresis detection.

[0049] Example 1

[0050] In a scene investigation, a cardboard carrier of physical evidence is used (20 μl of blood diluted 1000 times is dropped on the cardboard with a pipette and dried for 6 hours. Figure 5 A high-quality extraction method for biological evidence on a biological trace located on a paper sheet (a biological trace located on the upper side) is provided, which is achieved by squeezing a high-efficiency DNA enricher with the function of squeezing and re-absorbent reagents, using a suction head to create an air pressure difference, releasing a biological evidence collection reagent, wiping the biological trace position on the evidence carrier to wash off the detached cells and free DNA on the surface of the evidence carrier, and then using the air pressure difference to re-absorb the biological evidence collection reagent containing the detached cells and free DNA, and then using a dry cotton swab to wipe and absorb the residual biological evidence collection reagent, thereby completing the high-quality extraction of biological evidence. The specific operation is as follows:

[0051] S1. Open the package of the consumable high-efficiency DNA enricher. Specifically, open the package of the consumable high-efficiency DNA enricher. Figures 2 to 3 The sealing cover 600 in the structure shown in the figure breaks the limiting part 500 inside the efficient DNA enricher, so that the extrusion chamber 310 and the connecting tube 200 are connected to each other; the suction head 300 is squeezed to release the biological evidence collection reagent through the swab 400 to the Figure 5 At the biological trace position of the evidence carrier shown, the swab 400 on the suction head 300 is used to wipe the biological trace position where the biological evidence needs to be extracted, and the external force of wiping accelerates the elution of the detached cells and free DNA on the evidence;

[0052] S2. After squeezing the suction head 300, place the swab 400 on the biological trace position and release it, and use negative pressure to suck back the biological evidence collection reagent containing the exfoliated cells and free DNA; then use a dry cotton swab to wipe the small amount of biological evidence collection reagent remaining on the evidence; put the dry cotton swab and the high-efficiency DNA enricher containing the biological evidence collection reagent into a preservation tube for storage and bring it back to the laboratory for DNA extraction and testing;

[0053] S3. DNA extraction and testing: Use magnetic bead method or silica gel membrane method DNA extraction kit to extract DNA contained in dry cotton swab, swab 400 and biological evidence collection reagent; After Nanodrop quantitative detection of the extracted DNA, use VFP fluorescence detection kit for multi-color fluorescence labeling and PCR reaction multiple composite amplification to detect the polymorphism of STR loci in human genomic DNA in the sample evidence and identify individuals; Use ABI genetic analyzer for electrophoresis detection: Centrifuge the amplified product at 2000-5000rpm for 1-1min, take 1-10μL product or EX25 Allelic Ladder and mix it with 0.5-3μL AGCU Marker SI Z-500 and 1-12μL deionized formamide, denature at 50-120℃ for 1-10min, and then perform ice bath electrophoresis detection to obtain the following: Figure 6The DNA typing results are shown in Figure 2.

[0054] Depend on Figure 6 It can be seen that the method of Example 1 successfully and effectively detected DNA, with all sites detected and a peak value of more than 10,000, indicating that the DNA detection rate is high and the intensity is large, which is conducive to subsequent analysis and has high extraction quality.

[0055] Comparative Example 1:

[0056] In a field investigation, 20 μl of blood diluted 1000 times was dripped on another location of the cardboard of Example 1 using a pipette and dried for 6 hours. The blood in Comparative Example 1 was consistent with that in Example 1. Figure 5 The conventional wet-dry two-step method for biological evidence on the paperboard (the biological traces located below) includes the following steps:

[0057] Wet a clean cotton swab with pure water, wipe the trace position on the evidence carrier repeatedly, and then use another dry clean cotton swab to wipe the evidence carrier until its surface is dry.

[0058] Get as Figure 7 The DNA typing results are shown in Figure 2.

[0059] Depend on Figure 7 It can be seen that on the relatively flat surface of physical evidence such as cardboard, although the method of comparative example 1 can also detect DNA with all sites, the peak value is below 10,000, and the extraction quality needs to be improved.

[0060] It can be seen from the test results of Example 1 and Comparative Example 1 that the extraction method of the present invention can increase the amount of DNA extracted, improve the detection rate and detection intensity of DNA, be more conducive to subsequent data analysis, and have higher extraction quality.

[0061] Embodiment 2:

[0062] In a scene investigation, a stone carrier of physical evidence is used (20 μl of blood diluted 1000 times is dripped on the stone with a pipette and dried for 6 hours. Figure 8 The method for extracting biological evidence of high quality on the stone located above the sample is to squeeze the high-efficiency DNA enricher with the function of squeezing and sucking back the reagent, use the suction head to create an air pressure difference, release the biological evidence collection reagent, wipe the biological trace position on the evidence carrier to wash off the detached cells and free DNA on the surface of the evidence carrier, and then use the air pressure difference to suck back the biological evidence collection reagent containing the detached cells and free DNA, and then use a dry cotton swab to wipe and absorb the residual biological evidence collection reagent, thereby completing the high-quality extraction of biological evidence. The specific operation steps of this embodiment are basically the same as those of embodiment 1, the only difference is that the evidence carrier is as follows Figure 8 The stone shown in the figure finally Fig. 9The DNA typing results are shown in Figure 2.

[0063] Depend on Fig. 9 It can be seen that the DNA typing result diagram obtained by the method of Example 2 successfully and effectively detected the DNA, and all the sites were detected, indicating that the DNA detection rate was high, which is conducive to subsequent analysis.

[0064] Comparative Example 2:

[0065] In a field investigation, a stone carrying physical evidence (20 μl of blood diluted 1000 times was dripped on the stone using a pipette and dried for 6 hours. The blood and stone in Comparative Example 2 were the same as those in Example 2. The stone was taken from the same large stone. Figure 8 The conventional dry-wet two-step extraction of biological evidence on the stone at the bottom of the specimen includes the following steps:

[0066] Wet a clean cotton swab with pure water, wipe the trace position on the evidence carrier repeatedly, and then use another dry clean cotton swab to wipe the evidence carrier until its surface is dry.

[0067] Get as Fig.10 The DNA typing results are shown in Figure 2.

[0068] Depend on Fig.10 It can be seen that the DNA typing result diagram data obtained by the method of Comparative Example 2 is incomplete, and the sites are not all shown, indicating that the detection rate of DNA is low, which is not conducive to subsequent analysis, and the peak value is significantly lower than that of Example 2, and the extraction quality is obviously inferior to Example 2.

[0069] It can be seen from the test results of Example 2 and Comparative Example 2 that for complex physical evidence surfaces such as stones, the extraction method of the present invention can still effectively extract DNA, increase the amount of DNA extracted, increase the detection rate and detection intensity of DNA, and have a high extraction quality, and its effect is significantly better than that of conventional extraction methods.

[0070] The above-mentioned embodiments are only preferred embodiments of the present invention and cannot be used to limit the scope of protection of the present invention. Any non-substantial changes and substitutions made by technicians in this field on the basis of the present invention shall fall within the scope of protection required by the present invention.

Claims

1. A high-quality extraction method for biological evidence for on-site investigation, characterized in that: The extraction is performed using a high-efficiency DNA enricher, wherein the high-efficiency DNA enricher comprises a connecting portion, a connecting tube, a pipette tip, a swab, a limiting portion, and a sealing cover; The first end of the connecting tube is connected to the first end of the connecting part, and a first flow channel is provided in the connecting tube; the first end of the suction head is connected to the second end of the connecting part, and an extrusion cavity is provided inside the suction head, and the extrusion cavity is filled with a biological evidence collection reagent, and the biological evidence collection reagent is composed of the following components: 20-50mM buffer salt, 0.2%-2% thickener, 0.1%-1% surfactant, 20-50mM metal ion chelator, 0.1%-0.5% antibacterial agent; the swab is connected to the second end of the connecting tube and is connected to the first flow channel; the restriction part is located in the extrusion cavity, and the first end of the restriction part is connected to the second end of the connecting part. One end is connected to the second end of the connecting part, the limiting part can block the communication between the extrusion chamber and the first flow channel, and the limiting part can be broken, and after the limiting part is broken, the extrusion chamber and the first flow channel can be connected to each other; the sealing cover is sleeved on the outer periphery of the connecting tube and the swab, and the first end of the sealing cover is detachably connected to the first end of the connecting part; wherein, after the limiting part is broken, the biological evidence collection reagent in the extrusion chamber can be released into the swab by squeezing the suction head, and the biological evidence collection reagent can be recovered into the extrusion chamber through the swab under the negative pressure generated by squeezing the suction head; The method comprises the following steps: S1. Squeeze the high-efficiency DNA enricher to release the biological evidence collection reagent to the biological trace position of the evidence carrier through the swab of the high-efficiency DNA enricher; wipe the biological trace position with the swab to accelerate the elution of the detached cells and free DNA at the biological trace position; S2. Release the high-efficiency DNA enricher and suck back the biological evidence collection reagent containing detached cells and free DNA; wipe the biological evidence collection reagent remaining on the evidence carrier with a dry cotton swab; S3. Extract and test the DNA contained in the dry cotton swab, swab and biological evidence collection reagent to obtain a DNA typing result diagram.

2. The high-quality extraction method for biological evidence for on-site investigation according to claim 1 is characterized in that: The thickener is at least one of carbomer, sodium alginate or sodium polyacrylate.

3. The high-quality extraction method for biological evidence for on-site investigation according to claim 1 or 2, characterized in that: The concentration of the buffer salt is 5-20 mmol / L, and the buffer salt is one of Tris-HCl, phosphate, NaCl, KCl or MgCl; the surfactant is one of sodium dodecyl sulfate, TritonX-100 or Tween-80; and the antibacterial agent is potassium sorbate or benzoic acid.

4. The high-quality extraction method for biological evidence for on-site investigation according to claim 1 is characterized in that: A second flow channel is provided inside the restriction portion, a first end of the second flow channel is communicated with the first flow channel, a second end of the second flow channel can be closed by the second end of the restriction portion, and after the restriction portion is broken, the second end of the second flow channel is communicated with the extrusion chamber; A third flow channel is provided inside the connecting portion, and two ends of the third flow channel are respectively connected to the first flow channel and the second flow channel.

5. The high-quality extraction method for biological evidence for on-site investigation according to claim 1 is characterized in that: The surface of the suction head is marked with scale lines.

6. The high-quality extraction method for biological evidence for on-site investigation according to claim 1 is characterized in that: The suction head is made of soft material.

7. The high-quality extraction method for biological evidence for on-site investigation according to claim 1 is characterized in that: The step S3 uses a magnetic bead method or a silica gel membrane method DNA extraction kit to extract the DNA contained in the dry cotton swab, the swab and the biological evidence collection reagent.

8. The high-quality extraction method for biological evidence for on-site investigation according to claim 1 is characterized in that: The step S3 tests the extracted DNA according to the following steps: after the extracted DNA is quantitatively detected by Nanodrop, multi-color fluorescent labeling and PCR multiplex amplification are performed using a VFP fluorescence detection kit to detect the polymorphism of the STR loci in the DNA and identify individuals; and electrophoresis detection is performed using an ABI genetic analyzer.

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