Biosensor for sensitively and rapidly detecting activity of Whara syndrome helicase and application

Abstract

The invention relates to the field of biological detection, in particular to a biosensor for sensitively and rapidly detecting Whara syndrome helicase activity and application, the biosensor comprises a substrate chain, crRNA, Cas12a protein and a report chain FQ; according to the invention, a DNA-DNA / DNA-RNA substrate chain with complementary sequences at two ends and a non-complementary'bubble 'structure with a specific length in the middle is constructed, the substrate chain can be specifically unwound by WRN, and released single-stranded DNA further activates trans-cleavage activity and non-specific cleavage fluorescence reporter groups of Cas12a, so that unprecedented high-sensitivity and high-specificity detection on WRN helicase activity is realized; the fluorescent probe can be applied to in-situ and real-time fluorescence imaging and visual analysis of WRN activity in living cells, and has wide application prospects in the fields of cancer mechanism research, WRN-targeted inhibitor high-throughput screening, clinical adjoint diagnosis and the like.

Description

Technical Field

[0001] This invention relates to the field of biological detection, and in particular to a sensitive and rapid biosensor for detecting the activity of helicases in Werner syndrome and its application. Background Technology

[0002] Helicases play a crucial role in nucleic acid metabolism, participating extensively in key life activities such as DNA replication, recombination, transcription, repair, and telomere maintenance. Werner syndrome helicases (WRNs), as key members of the RecQ family, can deconstruct specialized nucleic acid structures formed during various metabolic processes and act as "guardians" in maintaining genome stability.

[0003] Studies have shown that loss of WRN function leads to chromosomal aberrations and telomere loss, resulting in cellular senescence or apoptosis, significantly increasing genomic instability and carcinogenic risk. Furthermore, WRN has been identified as a synthetic lethal target for microsatellite instability (MSI)-type cancers, thus becoming a highly promising biomarker and therapeutic target. Developing potent WRN inhibitors and elucidating their mechanisms of action in living cells is crucial for overcoming clinical drug resistance, guiding new drug development, and improving patient prognosis.

[0004] Currently, methods for detecting WRN helicase activity mainly fall into two categories: in vitro and in vivo. In vitro methods, such as electrophoretic mobility shift assay (EMSA), fluorescence resonance energy transfer assay (FRET), and various single-molecule techniques, generally suffer from problems such as cumbersome operation, long processing times, and difficulty in achieving high-throughput screening. Although single-molecule techniques can reveal details of enzyme action mechanisms, their high technical threshold and strong equipment dependence limit their widespread application in ordinary laboratories. In vivo detection methods mainly include immunofluorescence analysis (IF) and gene-encoded fluorescent probes. While these methods can detect intracellular WRN expression levels, they cannot effectively reflect its helicase activity. Although nucleic acid fluorescent probe methods can detect WRN activity in living cells, their sensitivity is insufficient to support in-depth mechanistic studies.

[0005] Since its discovery in E. coli in the 1980s, the CRISPR / Cas system has been developed as a highly efficient tool for gene editing and molecular detection. The trans-cleavage activity of the Cas12a protein can convert the presence of non-nucleic acid analytes (such as proteins) into detectable nucleic acid signals, providing a new pathway for achieving highly sensitive and specific protein detection. However, there are currently no reports of using this system for the detection of WRN helicase activity.

[0006] In summary, the existing technology lacks a method that can detect WRN helicase activity with high sensitivity and specificity in living cells, especially in terms of detecting both DNA-DNA double-strand activity and DNA-RNA hybrid chain activity, where there is a significant technological gap.

[0007] Therefore, based on the aforementioned technologies, there is an urgent need to develop a sensitive and rapid biosensor for detecting the activity of helicases in Werner syndrome and its applications. Summary of the Invention

[0008] In view of this, the purpose of this invention is to provide a sensitive and rapid biosensor for detecting WRN helicase activity in living cells and its application, in order to solve the problem that there is a lack of methods in the prior art that can detect WRN helicase activity with high sensitivity and high specificity in living cells.

[0009] To achieve the above objectives, the present invention provides a biosensor for the sensitive and rapid detection of helicase activity in Werner syndrome and its application.

[0010] A sensitive and rapid biosensor for detecting the activity of helicases in Werner syndrome, the biosensor comprising a substrate chain, crRNA, Cas12a protein, and reporter chain FQ; The substrate strand is a double-stranded DNA or DNA-RNA hybrid strand composed of complementary region one, non-complementary region two and complementary region two, wherein the non-complementary region is located between complementary region one and complementary region two. The non-complementary region is a sequence consisting of 8 pairs of non-complementary nucleotides; Both complementary regions one and complementary regions two are sequences consisting of complementary 19 / 20 pairs of nucleotides. The reporter chain FQ has a fluorescent group labeled at its 5' end and a quenching group labeled at its 3' end.

[0011] Preferably, the nucleotide sequence of the substrate chain is the sequence shown in SEQ ID NO:8 or SEQ ID NO:9.

[0012] Preferably, the nucleotide sequence of the crRNA is the sequence shown in SEQ ID NO:1.

[0013] Preferably, the molar ratio of Cas12a protein to crRNA is 1:1.25.

[0014] Preferably, the 5' end of the reporter chain FQ is labeled with a fluorescent group FAM, and the 3' end is labeled with a quenching group BHQ2.

[0015] Preferably, the concentration of Cas12a protein in the biosensor is 10 nM.

[0016] Preferably, the concentration of the substrate chain in the biosensor is 20 nM.

[0017] Preferably, the biosensor further includes a reaction buffer, wherein the reaction buffer is NEBuffer 4.

[0018] Preferably, the biosensor further includes adenosine triphosphate.

[0019] Application of a sensitive and rapid biosensor for detecting the activity of Werner syndrome helicase, said biosensor being used to detect the activity of Werner syndrome helicase in unwinding DNA-DNA double strands and DNA-RNA hybrid strands, or to prepare a kit for detecting the activity of Werner syndrome helicase.

[0020] The beneficial effects of this invention are: This invention provides a sensitive and rapid biosensor for detecting WRN helicase activity and its applications. The invention constructs a DNA-DNA / DNA-RNA substrate chain with complementary sequences at both ends and a non-complementary "bubble" structure of a specific length in the middle. This substrate chain can be specifically unwound by WRN, releasing single-stranded DNA that activates the trans-cleavage activity of Cas12a, non-specifically cleaving the fluorescent reporter group to generate a detectable fluorescent signal. Compared with existing technologies, this invention achieves unprecedented high sensitivity and specificity for detecting WRN helicase activity. It can be applied to in situ, real-time fluorescence imaging and visualization analysis of WRN activity within living cells, and has broad application prospects in cancer mechanism research, high-throughput screening of inhibitors targeting WRN, and clinical companion diagnostics. Attached Figure Description

[0021] To more clearly illustrate the technical solutions in this invention or the prior art, the drawings used in the description of the embodiments or the prior art will be briefly introduced below. Obviously, the drawings described below are only for this invention. For those skilled in the art, other drawings can be obtained based on these drawings without creative effort.

[0022] Figure 1 This is a schematic diagram illustrating the design principle of the biosensor of the present invention; Figure 2 This is an optimization of the ratio of Cas12a to crRNA in the two substrate chain unwinding reaction system of WRN in Example 1 of the present invention; Figure 3 This is for the optimization of the Cas12a concentration in the two substrate chain reaction systems for WRN unwinding in Example 2 of the present invention; Figure 4 This is an optimization of the reaction buffer in the WRN unwinding two substrate chain reaction system in Example 3 of the present invention; Figure 5 This is for optimizing the concentration of substrate chains in the WRN unwinding reaction system of the two substrate chains in Example 4 of the present invention; Figure 6 This is a schematic diagram (A) of the reaction system (B) of different lengths of crRNA responding to the substrate DNA-DNA / DNA-RNA in Example 5 of the present invention. Figure 7 This is a schematic diagram (A) of the reaction system (B) showing the fluorescence response of the substrate DNA-DNA / DNA-RNA to crRNA at complementary positions of the activation strand in this invention. Figure 8 This is a schematic diagram (A) of DNA-DNA / DNA-RNA substrate chains with different arm lengths on one side in this invention and their fluorescence response (B). Figure 9 This is a schematic diagram (A) of DNA-DNA / DNA-RNA substrate chains with different sizes of intermediate loops in this invention and their fluorescence response to 35nMWRN, and (B) of the reaction system. Figure 10 The graph shows the fluorescence response of substrate DNA-DNA to different concentrations of WRN (A) and the linear relationship between the concentration of WRN and the reaction rate of substrate DNA-DNA to WRN (B). Figure 11 The graph shows the fluorescence response of substrate DNA-RNA to different concentrations of WRN (A) and the linear relationship between the concentration of WRN and the reaction rate of substrate DNA-RNA to WRN (B). Figure 12 The fluorescence response of the substrate chain DNA-DNA to WRN and other helicases (A) and to other nucleases (B) in this invention are shown. Figure 13 The fluorescence response of the substrate chain DNA-RNA to WRN and other helicases (A) and to other nucleases (B) in this invention are shown. Figure 14 This is a fluorescence imaging diagram of DNA-DNA in different cells in this invention.

[0023] Figure 15 The images (A) and relative fluorescence intensity (B) of the DNA-DNA substrate chain in HCT116 cells treated with H3B968 in this invention are shown. Figure 16 The images (A) and relative fluorescence intensity (B) of the DNA-DNA substrate chain in HCT116 cells treated with HRO761 are shown in this invention. Figure 17The images (A) and relative fluorescence intensity (B) of the DNA-DNA substrate chain in HCT116 cells treated with NSC6171455 are shown in this invention. Detailed Implementation

[0024] To make the objectives, technical solutions, and advantages of this invention clearer, the invention will be further described in detail below with reference to specific embodiments.

[0025] The sources and properties of some of the raw materials used in this invention are as follows: Werner syndrome helicase (WRN), Bloom syndrome helicase (BLM), Beijing Yiqiao Shenzhou Technology Co., Ltd.; Recombinant RecQ1 and RecQ5 proteins with full helicase activity were expressed in the BL21 Escherichia coli system and purified by nickel column (GE Healthcare Technologies Inc., USA).

[0026] EnGen Lba Cas12a (Cpf1) nuclease, exonuclease III (EXO III), lambda exonuclease (Lambda exo), DNase I, endonuclease I (Endo I), lambda exonuclease (Lambda exo), apurinyl / pyrimidine endonuclease 1 (APE1), and uracil-DNA glycosylase (UDG), NEB Corporation, USA; DNA, RNA, Shanghai Jierui Biotechnology Co., Ltd.; Bovine serum albumin (BSA), Shanghai Aladdin Biochemical Technology Co., Ltd.; Hoechst 33342 (100×) blue fluorescent dye, Shanghai Beyotime Biotechnology Co., Ltd. H3B968 (C22H18F6N4O4S, CAS: 2912294-90-5), HRO761 (C31H31ClF3N9O5, CAS: 2869954-34-5), NSC 6171455 (C13H10Cl4N2O4, CAS: 203115-63-3), MCE Corporation, USA; DMEM medium, penicillin / streptomycin antibiotic solution, Opti-MEM I serum-depleted medium, Lipofectamine™ 3000 transfection reagent, Thermo Fisher Scientific (China) Co., Ltd. PM150710, Wuhan Punosei Life Technology Co., Ltd.; Fetal bovine serum, Nanjing Shenghang Biotechnology Co., Ltd.

[0027] Table 1. Enzyme buffer components corresponding to the nucleic acid repair enzymes used in the examples. Preparation example: Preparation of substrate chain S1: Purchase the DNA / RNA single strands shown in Table 2 and dissolve them in TE buffer (10 mM Tris-HCl, 0.1 mM EDTA, pH=8.0) to prepare a stock solution.

[0028] S2: Mix two single chains that can complement each other to form a bubble structure in a 1:1 molar ratio in annealing buffer (10 mM Tris-HCl, 120 mM Na). + In a solution containing pH=7.5, the final concentration was 2 μM. After heating at 95°C for 10 min, the solution was slowly cooled to room temperature to form a stable substrate chain (e.g., B8D19, SEQ ID NO:9; B8D20, SEQ ID NO:8). The solution was then stored at 4°C for later use.

[0029] Table 2. Specific sequences of DNA-DNA / DNA-RNA probes Example 1: Optimization of the Cas12a to crRNA ratio Mix 10×NEBuffer 4, 0.32, 0.4, and 0.48 μL of 625 nM crRNA, 0.8 μL of 25 nM, 0.5 μL of 10 μM FQ, and enzyme-free water in 200 μL centrifuge tubes to achieve Cas12a:crRNA ratios of 1:1, 1:1.25, and 1:1.5, respectively. Incubate at 37°C for 15 min. Then, add 2 μL of 10 mM ATP to the mixing tube and mix well. Spot 2 μL of 200 nM DNA-DNA / DNA-RNA onto the centrifuge tube cap. Take WRN protein from a -80°C freezer, prepare it to 350 nM, and add it to the system to make a final reaction volume of 20 μL. Cap and centrifuge for 20 s. After mixing, start the reaction and immediately use a qPCR instrument to detect the change in fluorescence signal intensity over time. The detection conditions are 37°C, fluorescence is collected every 30 s, and the detection channel is FAM. Channel, results as follows Figure 2 As shown.

[0030] When the Cas12a:crRNA ratio was 1:1.25, the fluorescence difference between the control group and the control group was the largest. Increasing the ratio did not increase the overall fluorescence intensity, so this ratio was selected for subsequent experiments.

[0031] Example 2: Optimization of Cas12a concentration Maintaining a Cas12a:crRNA ratio of 1:1.25, and setting final Cas12a concentrations of 10 nM, 20 nM, and 30 nM respectively, with other conditions remaining the same, the results are as follows. Figure 3 As shown, the signal-to-noise ratio is highest when the Cas12a concentration is 10 nM, so this concentration was selected for subsequent experiments.

[0032] Example 3: Optimization of reaction buffer NEBuffer 4, NEBuffer 2.1, NEBuffer 3.1, and rCutSmart were selected as reaction buffers (see Table 3), and the remaining conditions were the same as the optimal conditions in Example 4. The results are as follows. Figure 4 As shown, when using NEBuffer 4, the experimental group had the strongest fluorescence signal and the control group had the lowest background, so NEBuffer 4 was selected for subsequent experiments.

[0033] Table 3 Composition of each reaction buffer Example 4: Optimization of substrate chain concentration The final concentrations of substrate chains B8D19 and B8D20 were set to 1 nM, 5 nM, 10 nM, 20 nM, 40 nM, and 50 nM, respectively, with other conditions remaining the same as the optimal conditions in Example 1. The results are as follows: Figure 5 As shown, when the substrate chain concentration was 20 nM, the experimental group exhibited a strong signal and low background leakage; therefore, this concentration was selected for subsequent experiments. Example 5: Optimization of crRNA Various crRNA sequences with different numbers (16-20 bases) and complementary positions to the activating strand were designed and investigated (see Table 4). Under the optimal reaction conditions determined in Examples 1-4, the fluorescence response of these crRNAs to 35 nM WRN was tested. Results are as follows: Figure 6-7 As shown, when using crRNA-3 (SEQ ID NO:1), which is 20 bases completely complementary to the activating strand and whose complementary region is not on the non-complementary region of the substrate strand, Cas12a has the best activation effect and the lowest background signal.

[0034] Table 4 crRNA Sequences Performance testing: Substrate chain performance testing A variety of DNA-DNA and DNA-RNA substrate chains with different unilateral complementary arm lengths (10, 15, 19 / 20, 25 bp) and different sizes of the central non-complementary region (6, 8, 10, 12, 14 nt) were designed and investigated (see Table 2). The fluorescence response of these substrate chains to 35 nM WRN was tested under the optimal reaction conditions determined in Example 1. The results are as follows: Figure 8-9 As shown, the experimental group with substrate chains of 19bp / 20bp on both sides exhibited high fluorescence intensity and low signal leakage, indicating that B8D19 / B8D20 performed better and that WRN had better unwinding activity.

[0035] Biosensor sensitivity testing Under the optimal reaction conditions determined in Example 4, the fluorescence response of different concentrations (0.01-5.00 nM) of WRN to substrate chains B8D19 (DNA-DNA) and B8D20 (DNA-RNA) was tested. A graph was plotted against WRN concentration with initial reaction rate. The results are as follows: Figure 10-11 As shown, for DNA-DNA substrates, the limit of detection (LOD) of this biosensor is 5.6 pM, exhibiting good linearity in the range of 0.025–5.00 nM; for DNA-RNA substrates, the LOD is 6.0 pM, exhibiting good linearity in the range of 0.01–1.00 nM.

[0036] Biosensor Selectivity Testing Under the optimal reaction conditions determined in Example 1, substrate chains B8D19 and B8D20 were treated with 35 nM WRN, BLM, RECQ1, RECQ5 helicases, and high concentrations (>1000 nM) of EXO III, EXO I, DNase I, UDG, APE 1 nuclease, and Lambda exonuclease, respectively. The results are as follows: Figure 12-13 As shown, this biosensor exhibits a strong fluorescence response only to WRN, while responding weakly to other tested enzymes, demonstrating extremely high specificity.

[0037] Table 5. Selective responses of biosensors to different enzymes (relative fluorescence intensity %) Intracellular WRN activity imaging 1. Human colon cancer cells HCT116 were seeded at a density of 1.5 × 10^5 cells per well in a 96-well laser confocal microscopy plate and cultured for 12 hours.

[0038] 2. Transfection complex containing 31.25 nM crRNA (SEQ ID NO:1), 25 nM Cas12a protein, 20 nM substrate chain B8D19 (SEQ ID NO:9), and 250 nM FQ (using Lipofectamine) TM Add the 3000 reagent to the cells and incubate at 37°C for 6 hours.

[0039] 3. Discard the culture medium, wash with PBS, and add Hoechst 33342 for nuclear staining. Observe using an OLYMPUS FV3000 laser confocal microscope; the FAM channel (green) shows WRN activity, and the DAPI channel (blue) shows the cell nucleus. Results are as follows: Figure 14 As shown.

[0040] Table 6. Fluorescence response of biosensors under different cell and drug treatments Differentiating WRN activity in different cell lines Human colon cancer cells (HCT116), normal human colon epithelial cells (NCM460), human pancreatic cancer cells (PANC-1), and human umbilical vein endothelial cells (HUVEC) were processed and imaged using the "intracellular WRN activity imaging" method. The results are as follows: Figure 14 As shown, Evaluate the efficacy of WRN inhibitors HCT116 cells were treated with WRN small molecule inhibitors H3B968, HRO761, and NSC6171455 at concentrations of 0, 1, 5, 10, 15, and 20 μM, respectively, with a 1% DMSO control group included. The sensor was then transfected and imaged using the "Intracellular WRN Activity Imaging" method. Results are as follows: Figure 15-17 As shown Data Analysis: from Figure 1-17 As can be seen from Tables 5-6, the biosensor for Werner syndrome helicase activity prepared in this invention exhibits ultra-high sensitivity and excellent specificity. This biosensor achieves a detection limit (LOD) of pM for WRN (DNA-DNA: 5.6 pM; DNA-RNA: 6.0 pM). This sensitivity is 2-3 orders of magnitude higher than that of traditional immunofluorescence methods (which can only perform qualitative or semi-quantitative analysis) and existing nucleic acid fluorescent probe methods (which typically have a sensitivity in the nM range). This makes it possible to detect WRN activity at extremely low concentrations, and to conduct early diagnosis or drug efficacy evaluation. The sensor only responds strongly to WRN, while it only responds weakly (<10%) to helicases BLM, RECQ1, RECQ5, and other nucleases with different functions (EXO III, EXO I, DNase I, APE 1, UDG, Lambda exonuclease). This indicates that the sensor can effectively avoid interference from the complex intracellular enzymatic environment and ensure the accuracy of the detection results. When using crRNA-16 (SEQ ID NO:5), which is only 16 bases complementary, the fluorescence signal intensity was reduced by approximately 70% compared to the example using the optimal crRNA-3 (SEQ ID NO:1). This result confirms that sufficiently long and precise complementarity between crRNA and the activating strand is a structural prerequisite for efficient activation of Cas12a trans-cleavage activity. Insufficient complementarity leads to a significant decrease in the binding affinity and activation efficiency of the Cas12a / crRNA complex to the activating strand, directly affecting the signal amplification effect. When using substrate chain B14D19 (SEQ ID NO:17) with a 14 nt non-complementary region in the middle, the fluorescence signal intensity was reduced by approximately 65% ​​compared to the example using substrate chain B8D19 (SEQ ID NO:9) with the optimal 8 nt bubble structure. This comparison demonstrates that the 8 nt non-complementary region is the "golden size" for optimal loading and efficient unwinding of WRN helicase. Excessively large bubbles may hinder the initial recognition and binding of WRN or affect the processivity (continuous synthesis / unwinding ability) of the unwinding process, thereby reducing unwinding efficiency and the release of fewer activating chains.

[0041] Furthermore, the biosensor successfully generated a clear green fluorescence signal in living cells such as HCT116 and co-localized with the nuclear dye (blue). It enabled in-situ, real-time fluorescence imaging and visualization analysis of WRN unwinding activity inside living cells, overcoming the limitations of existing technologies that can only detect content or in vitro activity. Moreover, the fluorescence intensity in cancer cells HCT116 was 4 times that in normal cells NCM460. This significant difference (p < 0.01) indicates that the sensor can effectively distinguish cancer cells from normal cells based on the level of WRN activity, providing a potential tool for molecular subtyping of cancer and precision treatment strategies based on WRN activity. Treatment of HCT116 cells with 1 μM of the WRN inhibitors H3B968, HRO761, and NSC 6171455 resulted in a 58%, 62%, and 55% decrease in intracellular fluorescence intensity, respectively. This dose-dependent fluorescence attenuation (e.g.) Figure 15-17 As shown in the figure, this biosensor clearly demonstrates that it can quantitatively assess the efficacy and inhibitory efficiency of WRN inhibitors in cells in situ and in real time, providing a powerful tool for high-throughput drug screening.

[0042] Those skilled in the art should understand that the discussion of any of the above embodiments is merely exemplary and is not intended to imply that the scope of the invention is limited to these examples; within the framework of the invention, the technical features of the above embodiments or different embodiments can also be combined, the steps can be implemented in any order, and there are many other variations of the different aspects of the invention as described above, which are not provided in detail for the sake of brevity.

[0043] This invention is intended to cover all such substitutions, modifications, and variations that fall within the broad scope of the appended claims. Therefore, any omissions, modifications, equivalent substitutions, improvements, etc., made within the spirit and principles of this invention should be included within the scope of protection of this invention.

Claims

1. A biosensor for sensitive and rapid detection of Werner syndrome helicase activity, characterized by, The biosensor includes a substrate chain, crRNA, Cas12a protein, and a reporter chain FQ; The substrate strand is a double-stranded DNA or DNA-RNA hybrid strand composed of complementary region one, non-complementary region two and complementary region two, wherein the non-complementary region is located between complementary region one and complementary region two. The non-complementary region is a sequence consisting of 8 pairs of non-complementary nucleotides; Both complementary regions one and complementary regions two are sequences consisting of complementary 19 / 20 pairs of nucleotides. The reporter chain FQ has a fluorescent group labeled at its 5' end and a quenching group labeled at its 3' end.

2. The biosensor for rapid and sensitive detection of Werner syndrome helicase activity according to claim 1, wherein The nucleotide sequence of the substrate chain is the sequence shown in SEQ ID NO:8 or SEQ ID NO:

9.

3. The biosensor for rapid and sensitive detection of Werner syndrome helicase activity according to claim 1, wherein The nucleotide sequence of the crRNA is shown in SEQ ID NO:

1.

4. The biosensor for rapid and sensitive detection of Werner syndrome helicase activity according to claim 1, wherein The molar ratio of Cas12a protein to crRNA is 1:1.

25.

5. The biosensor for sensitive and rapid detection of helicase activity in Werner syndrome according to claim 1, characterized in that, The reporter chain FQ is labeled with a fluorescent group FAM at its 5' end and a quenching group BHQ2 at its 3' end.

6. The biosensor for sensitive and rapid detection of helicase activity in Werner syndrome according to claim 1, characterized in that, The concentration of Cas12a protein in the biosensor is 10 nM.

7. The biosensor for sensitive and rapid detection of helicase activity in Werner syndrome according to claim 1, characterized in that, The concentration of the substrate chain in the biosensor is 20 nM.

8. The biosensor for sensitive and rapid detection of helicase activity in Werner syndrome according to claim 1, characterized in that, The biosensor also includes a reaction buffer, which is NEBuffer 4.

9. A biosensor for sensitive and rapid detection of helicase activity in Werner syndrome according to claim 1, characterized in that, The biosensor also includes adenosine triphosphate.

10. The application of a biosensor for sensitive and rapid detection of helicase activity in Werner syndrome according to any one of claims 1-9, characterized in that, The biosensor is used to detect the activity of Werner syndrome helicase in unwinding DNA-DNA double strands and DNA-RNA hybrid strands, or to prepare a kit for detecting Werner syndrome helicase activity.

Images