Method, System and Device for Implementing Normalization of Detection Signals of Gene Analyzers

Abstract

The present invention provides a method, a system and a device for realizing the normalization of detection signals of a gene analyzer. The method includes: performing internal standard matching on the spectral signals of the internal standard channels of each capillary collected during the electrophoresis process of the gene analyzer, and calculating the single-channel internal standard average value of the internal standard channels of each capillary according to the successfully matched internal standard matching sequences; respectively dividing the preset internal standard standard values by the single-channel internal standard average values of the internal standard channels of each capillary, and taking the obtained calculation results as the normalization coefficients of the corresponding capillaries; respectively performing normalization correction on the spectral signals of other color channels in their respective capillaries according to the normalization coefficients of each capillary to realize the normalization of detection signals. The present invention realizes the adjustment of the peak height of a sample to be detected according to the peak height of the internal standard, can improve the signal differences between different capillaries and the differences in different detection times of the same capillary, and effectively improves the quality of gene analysis results.

Description

Technical Field

[0001] The present invention relates to the technical field of biochemical detection, and in particular, to a method, a system and a device for realizing normalization of detection signals of a gene analyzer. Background Art

[0002] A gene analyzer based on capillary electrophoresis technology can be applied to Sanger sequencing and gene fragment analysis. Capillary electrophoresis uses a quartz capillary as the separation channel, a high-voltage direct current electric field as the driving force, and a porous gel filled as the support medium. The pore size distribution of the gel is ensured by temperature control to match the DNA conformation. When the size of the DNA molecule is comparable to the pore size of the gel, its mobility is related to the size. Short fragments are less hindered and move faster through the capillary, while long fragments are more hindered and move slower through the capillary. Since DNA molecules are negatively charged, after applying a direct current high voltage across the two ends of the capillary, the DNA labeled with a fluorescent group will enter the capillary from the cathode end of the capillary by electro-injection and move towards the anode. DNA molecules of different lengths will pass through the detection window successively. When a certain DNA molecule passes through the optical detection window, the fluorescent group on the DNA is excited by a laser to generate fluorescence, which is then collected by a spectrometer. The spectrometer converts the optical signal into an electrical signal and then into a digital signal. After processing the original digital signal and analyzing it with analysis software, the base sequence or relative fragment length of the DNA molecule can be obtained.

[0003] The signals collected by the gene analyzer are essentially optical signals. The optical signals are further divided into the fluorescence signals on the fluorescent groups of DNA and the Raman signals of the electrophoresis gel. Generally, the Raman signals are used to characterize the state of the optical system, while the DNA fluorescence signals are affected by many factors such as the sampling time, sampling voltage, sampling position, sample concentration, optical system, and the state of the capillary itself. Therefore, when using a gene analyzer for detection, the signal differences between different capillaries and the signal differences of the same capillary for different detection times are always inevitable, resulting in deviations in the detection results. How to solve the signal differences between different capillaries and the signal differences of the same capillary for different detection times is of great significance for improving the quality of gene analysis results. Summary of the Invention

[0004] In view of the above problems, the present invention is proposed to provide a method, a system and a device for realizing normalization of detection signals of a gene analyzer that can overcome or at least partially solve the above problems.

[0005] In one aspect of the present invention, a method for realizing normalization of detection signals of a gene analyzer is provided. The method includes:

[0006] Perform internal standard matching on the spectral signals of the internal standard channels of each capillary collected during the electrophoresis process of the gene analyzer, and calculate the single-channel internal standard average value of the internal standard channels of each capillary according to the successfully matched internal standard matching sequences;

[0007] Divide the preset internal standard standard value by the single-channel internal standard average value of the internal standard channels of each capillary respectively, and use the obtained calculation result as the normalization coefficient of the corresponding capillary;

[0008] Perform normalization correction on the spectral signals of other color channels in each capillary according to the normalization coefficient of each capillary respectively, so as to achieve normalization of the detection signals.

[0009] Optionally, before performing internal standard matching on the spectral signals of the internal standard channels of each capillary collected during the electrophoresis process of the gene analyzer, the method further includes:

[0010] Perform electrophoresis on multiple capillaries filled only with the internal standard of a specified concentration, so as to collect the spectral signals of the internal standard channels of each capillary and perform internal standard matching, calculate the average peak value of each internal standard peak in the internal standard matching sequence corresponding to each capillary that meets the internal standard matching requirements, and obtain the single-channel internal standard average value of the internal standard channels of each capillary;

[0011] Calculate the coefficient of variation of the single-channel internal standard average values of the internal standard channels of each capillary. If the coefficient of variation is less than the preset variation threshold, then use the mean value or a preset multiple of the mean value of the single-channel internal standard average values of the internal standard channels of each capillary as the internal standard standard value.

[0012] Optionally, after using the obtained calculation result as the normalization coefficient of the corresponding capillary, the method further includes:

[0013] Judge whether the normalization coefficient of each capillary is compliant. When the normalization coefficient of the capillary is within the preset range of normalization coefficient values, it is determined that the normalization coefficient of the current capillary is compliant, otherwise it is determined that the normalization coefficient of the current capillary is non-compliant;

[0014] If the proportion of the number of capillaries with non-compliant normalization coefficients in the total number of capillaries in the current electrophoresis detection data is greater than the preset first proportion threshold, or, if the proportion of the number of capillaries with non-compliant normalization coefficients in the total number of capillaries in at least three consecutive electrophoresis detection data is greater than the preset second proportion threshold, then correct the internal standard standard value;

[0015] Wherein, the second proportion threshold is less than the first proportion threshold.

[0016] Optionally, the performing normalization correction on the spectral signals of other color channels in each capillary according to the normalization coefficient of each capillary respectively includes:

[0017] If the normalization coefficient of the capillary is compliant, multiply the spectral signals of other color channels in the current capillary by the normalization coefficient of the corresponding capillary to achieve the normalization correction of the spectral signals;

[0018] If the normalization coefficient of the capillary is non-compliant, multiply the spectral signals of other color channels in the current capillary by the extreme value close to the value range of the normalization coefficient to achieve the normalization correction of the spectral signals.

[0019] Optionally, before performing internal standard matching on the spectral signals of the internal standard channels of each capillary collected during the electrophoresis process of the genetic analyzer, the method further includes:

[0020] Monitoring the Raman signal during the electrophoresis process to obtain the signal value, peak balance data, and / or uniformity data of the Raman signal;

[0021] When any one of the signal value, peak balance data, and / or uniformity data of the Raman signal does not meet the corresponding preset optical parameter standard, perform an optical alignment operation until the signal value, peak balance data, and / or uniformity data all meet the corresponding optical parameter standards.

[0022] Optionally, the internal standard matching of the spectral signals of the internal standard channels of each capillary collected during the electrophoresis process of the genetic analyzer includes:

[0023] S10. Perform data preprocessing on the spectral signals of the internal standard channels of each capillary collected during the electrophoresis process of the genetic analyzer to filter out the background noise of the spectral signals and smooth the spectral signals;

[0024] S11. Perform peak identification on the spectral signals after data preprocessing to screen out the candidate peak sequences included in the current spectral signals;

[0025] S12. Perform internal standard matching on the candidate peaks in the candidate peak sequence in the order of sampling with the standard peak sequence in the standard spectral signals corresponding to the internal standard selected during the electrophoresis process in turn to find the candidate peak combinations that meet the preset internal standard matching conditions in the candidate peak sequence, and the number of candidate peaks in the candidate peak combination is greater than or equal to 3; the internal standard matching conditions include that the absolute value of the difference between the first distance and the second distance is less than the preset distance error threshold, and max{the peak height of the selected candidate peaks in the candidate peak combination, the peak height of the current candidate peak to be matched} / min{the peak height of the selected candidate peaks in the candidate peak combination, the peak height of the current candidate peak to be matched}<the preset relative height threshold, where the first distance is the distance between the peak points of adjacent candidate peaks in the candidate peak sequence, and the second distance is the distance between the peak points of adjacent standard peaks with the same sampling order as the candidate peak for which the first distance is currently calculated in the standard peak sequence;

[0026] S13. Using each candidate peak combination as a matching basis, perform internal standard matching on other candidate peaks in the candidate peak sequence in turn to obtain the matching result corresponding to each candidate peak combination;

[0027] S14. Count the number of candidate peaks included in the matching result corresponding to each candidate peak combination;

[0028] S15. When the maximum value of the number of candidate peaks included in the matching results corresponding to each candidate peak combination is equal to the number of standard peaks included in the standard peak sequence, it is determined that the internal standard matching is successful, and the matching result corresponding to the maximum value is used as the optimal internal standard matching result.

[0029] Optionally, after using the matching result corresponding to the maximum value as the optimal internal standard matching result, the method includes:

[0030] Perform curve fitting on the standard peak sequence and the peak sequence of the optimal internal standard matching result, and use the standard deviation, average residual, and maximum residual after curve fitting as the characteristic data of the current optimal internal standard matching result;

[0031] Input the characteristic data into a preset internal standard matching scoring model for identification to obtain the matching degree score of the current optimal internal standard matching result.

[0032] Optionally, the method further includes:

[0033] If the maximum value of the number of candidate peaks included in the matching results corresponding to each candidate peak combination is not equal to the number of standard peaks included in the standard peak sequence, update the distance error threshold according to a preset first threshold adjustment rule, and return to step S12 until the updated distance error threshold is greater than the maximum value of the distance error threshold;

[0034] When the updated distance error threshold is greater than the maximum value of the distance error threshold, update the relative height threshold according to a preset second threshold adjustment rule, and update the distance error threshold to the corresponding initial value, and return to step S12 until the updated relative height threshold is greater than the maximum value of the relative height threshold;

[0035] When the updated relative height threshold is greater than the maximum value of the relative height threshold, it is determined that the internal standard matching fails.

[0036] In a second aspect, the present invention further provides a system for realizing the normalization of the detection signal of a gene analyzer, and the system includes functional modules for realizing the method for realizing the normalization of the detection signal of a gene analyzer as described above.

[0037] In a third aspect, the present invention also provides a computer device, including a memory, a processor, and a computer program stored on the memory and executable on the processor. When the processor executes the computer program, the steps of the method for normalizing the detection signals of a gene analyzer as implemented above are realized.

[0038] The method, system, and device for normalizing the detection signals of a gene analyzer provided by the embodiments of the present invention calculate the normalization coefficient corresponding to each capillary through a preset internal standard standard value and the single-channel internal standard average value of the internal standard channel of each capillary during the electrophoresis process of the gene analyzer, and perform normalization correction on the spectral signals of other color channels in each capillary according to the normalization coefficient of each capillary, so as to adjust the peak height of the sample to be tested according to the peak height of the internal standard, improve the signal difference between different capillaries, and the difference in different detection times of the same capillary, and effectively improve the quality of gene analysis results.

[0039] The above description is only an overview of the technical solution of the present invention. In order to be able to understand the technical means of the present invention more clearly, it can be implemented according to the content of the description. And in order to make the above and other purposes, features, and advantages of the present invention more obvious and understandable, the specific embodiments of the present invention are specifically described below. BRIEF DESCRIPTION OF THE DRAWINGS

[0040] By reading the detailed description of the preferred embodiments below, various other advantages and benefits will become clear to those of ordinary skill in the art. The drawings are only for the purpose of showing the preferred embodiments and are not considered to be a limitation of the present invention. Moreover, throughout the drawings, the same reference numerals are used to represent the same components. In the drawings:

[0041] Figure 1 is a flowchart of the method for normalizing the detection signals of a gene analyzer provided by the embodiments of the present invention;

[0042] Figure 2 is a schematic diagram of the implementation principle of distance similarity in the internal standard matching conditions provided by the embodiments of the present invention;

[0043] Figure 3 is a flowchart of the internal standard matching method for the detection spectrum of a gene analyzer provided by the embodiments of the present invention;

[0044] Figure 4 is a structural block diagram of the system for normalizing the detection signals of a gene analyzer provided by the embodiments of the present invention. DETAILED DESCRIPTION OF THE EMBODIMENTS

[0045] Exemplary embodiments of the present disclosure will be described in more detail below with reference to the accompanying drawings. Although the exemplary embodiments of the present disclosure are shown in the drawings, it should be understood that the present disclosure can be implemented in various forms and should not be limited by the embodiments set forth herein. On the contrary, these embodiments are provided so that the present disclosure can be more thoroughly understood and the scope of the present disclosure can be fully conveyed to those skilled in the art.

[0046] Those skilled in the art of the present technology can understand that, unless specifically stated otherwise, the singular forms "a", "an", "the", and "said" used herein may also include the plural forms. It should be further understood that the term "comprising" used in the specification of the present invention means the presence of the recited features, integers, steps, operations, elements, and / or components, but does not exclude the presence or addition of one or more other features, integers, steps, operations, elements, components, and / or groups thereof.

[0047] Those skilled in the art of the present technology can understand that, unless otherwise defined, all terms (including technical terms and scientific terms) used herein have the same meaning as the general understanding of those of ordinary skill in the art to which the present invention belongs. It should also be understood that terms such as those defined in a general dictionary should be understood to have a meaning consistent with the meaning in the context of the prior art, and will not be interpreted in an idealized or overly formal sense unless specifically defined.

[0048] When using a gene analyzer for detection, signal differences between different capillaries and differences in the number of detections of the same capillary have always been inevitable, resulting in deviations in the detection results. Factors causing signal deviations include loading time, loading voltage, loading position, sample concentration, optical system, etc. For general instruments, the loading time can be controlled by software, and the loading voltage can be corrected by a gauge. The loading position can be corrected by a loading table, and the optical system can be corrected by optical alignment. However, in addition to factors such as loading time, loading voltage, loading position, sample concentration, and optical system that cause signal deviations, there are also deviations caused by the state of the capillary itself, such as deviations caused by the inner diameter of the capillary group, resistance deviations caused by poor contact between a certain capillary and the electrode, resistance deviations caused by random entry of bubbles into the capillary, and resistance deviations caused by uneven temperatures of different capillaries. These factors cannot be completely avoided. Therefore, in order to avoid affecting the quality of gene analysis results due to signal deviations in detection signals, the present invention proposes a method for normalizing detection signals of a gene analyzer.

[0049] Figure 1 A flowchart of a method for normalizing detection signals of a gene analyzer according to an embodiment of the present invention is schematically shown. Referring to Figure 1 , the method for normalizing detection signals of a gene analyzer according to an embodiment of the present invention specifically includes the following steps:

[0050] S1. Perform internal standard matching on the spectral signals of the internal standard channels of each capillary collected during the electrophoresis process of the gene analyzer, and calculate the single-channel internal standard average value of the internal standard channels of each capillary according to the successfully matched internal standard matching sequences.

[0051] In the embodiments of the present invention, before performing internal standard matching on the spectral signals of the internal standard channels of each capillary collected during the electrophoresis process of the gene analyzer, the method further includes the step of establishing an internal standard standard value, which specifically includes: performing electrophoresis on multiple capillaries filled only with an internal standard of a specified concentration to collect the spectral signals of the internal standard channels of each capillary and perform internal standard matching, calculating the average peak value of each internal standard peak in the internal standard matching sequence corresponding to each capillary that meets the internal standard matching requirements, and obtaining the single-channel internal standard average value of the internal standard channels of each capillary; calculating the coefficient of variation of the single-channel internal standard average values of the internal standard channels of each capillary. If the coefficient of variation is less than a preset variation threshold, for example, 30%, then use the mean value or a preset multiple of the mean value of the single-channel internal standard average values of the internal standard channels of each capillary as the internal standard standard value. If the coefficient of variation is greater than or equal to the preset variation threshold, then re-execute the step of establishing the internal standard standard value. Among them, the preset multiple can be selected from 0.1 - 10 times.

[0052] Specifically, the specific calculation model of the coefficient of variation CV is: CV = standard deviation / average value;

[0053] Where: n is the number of capillary channels, and x i is the single-channel internal standard average value of the i-th capillary channel.

[0054] Specifically, when establishing the internal standard standard value, electrophoresis is performed using an internal standard of a certain concentration, and through steps such as data preprocessing, peak identification, internal standard matching, and scoring for internal standard matching, qualified internal standard matching sequence data for peak normalization is selected. If it is unqualified, that is, the internal standard matching is not successful, then this data is not used. In an alternative embodiment, for a single-channel instrument, the average peak height of all internal standards in a single channel is recorded as the single-channel internal standard average value. For example, if there are 10 peaks in an internal standard, then calculate the average value of the 10 peaks to obtain the single-channel internal standard average value, and use it as the internal standard standard value. In another alternative embodiment, for an 8-channel instrument, if the data of all 8 channels are qualified, then calculate the single-channel internal standard average values of the 8 channels, and then take the average value of the 8 channel peaks, that is, the mean value of the 8 single-channel internal standard average values, to obtain the internal standard standard value.

[0055] Among them, the DNA samples for electrophoresis in a gene analyzer are usually divided into three components: formamide or pure water as the solvent, an internal standard as the scale, and the sample to be tested labeled with a fluorescent group. Among them, generally the concentration of the internal standard is fixed. When establishing the internal standard standard value, the concentration of the internal standard is the same as that of the internal standard in the sample to be tested during detection, and the concentration of the sample to be tested is unknown. Therefore, adjusting the peak height of the sample to be tested according to the peak height of the internal standard can improve the signal deviation within a certain range.

[0056] S2. Divide the preset internal standard standard value by the single-channel internal standard average value of the internal standard channel of each capillary respectively, and use the obtained calculation result as the normalization coefficient of the corresponding capillary.

[0057] S3. Perform normalization correction on the spectral signals of other color channels in each capillary according to the normalization coefficient of each capillary respectively to achieve normalization of the detection signals.

[0058] The method for realizing normalization of detection signals of a gene analyzer provided by the embodiments of the present invention calculates the normalization coefficient of the corresponding capillary through the preset internal standard standard value and the single-channel internal standard average value of the internal standard channel of each capillary during the electrophoresis process of the gene analyzer, and performs normalization correction on the spectral signals of other color channels in each capillary according to the normalization coefficient of each capillary, so as to adjust the peak height of the sample to be tested according to the peak height of the internal standard, improve the signal difference between different capillaries, and the difference in different detection times of the same capillary, and effectively improve the quality of gene analysis results.

[0059] In the embodiments of the present invention, after using the obtained calculation result as the normalization coefficient of the corresponding capillary, the method further includes: judging whether the normalization coefficient of each capillary is compliant. When the normalization coefficient of the capillary is within the preset range of normalization coefficient values, it is determined that the normalization coefficient of the current capillary is compliant, otherwise it is determined that the normalization coefficient of the current capillary is non-compliant; if the proportion of the number of capillaries with non-compliant normalization coefficients in the total number of capillaries in the current electrophoresis detection data is greater than the preset first proportion threshold, or, if the proportion of the number of capillaries with non-compliant normalization coefficients in the total number of capillaries in at least three consecutive electrophoresis detection data is greater than the preset second proportion threshold, then correct the internal standard standard value; wherein, the second proportion threshold is less than the first proportion threshold.

[0060] Specifically, the normalization coefficient of the capillary should be within a certain value range. If it exceeds the range, it is determined that the normalization coefficient of the current capillary is non-compliant. Optionally, the value range of the normalization coefficient can be taken between 0.3 and 3. If during the detection process, it is found that the proportion of the number of capillaries with non-compliant normalization coefficients in the total number of capillaries in the current electrophoresis detection data is greater than the preset first proportion threshold, or, the proportion of the number of capillaries with non-compliant normalization coefficients in the total number of capillaries in at least three consecutive electrophoresis detection data is greater than the preset second proportion threshold, then it is determined that the internal standard standard value needs to be corrected. Among them, the first proportion threshold can be optionally 50%, and the second proportion threshold can be optionally 30%.

[0061] Furthermore, correcting the internal standard standard value includes calculating the coefficient of variation of the normalization coefficients of each capillary. If the coefficient of variation of the normalization coefficients of each capillary is less than the preset second variation threshold, for example, 30%, then the mean or a preset multiple of the mean of the single-channel internal standard means of each capillary internal standard channel of all current capillaries is used as the new internal standard standard value. If the coefficient of variation is greater than or equal to the preset second variation threshold, it is prompted to re-establish the internal standard standard value through electrophoresis. The preset multiple can be optionally 0.1 - 10 times. The coefficient of variation of the normalization coefficient of each capillary is the ratio of the standard deviation of the normalization coefficient of the current capillary to the average of the normalization coefficients of all capillaries. The calculation method refers to the coefficient of variation of the single-channel internal standard mean of the capillary internal standard channel.

[0062] In the embodiment of the present invention, the normalization correction of the spectral signals of other color channels in each capillary according to the normalization coefficient of each capillary includes: if the normalization coefficient of the capillary is compliant, then multiply the spectral signals of other color channels in the current capillary by the normalization coefficient of the corresponding capillary to achieve spectral signal normalization correction; if the normalization coefficient of the capillary is non-compliant, then multiply the spectral signals of other color channels in the current capillary by the extreme value close to the value range of the normalization coefficient of the corresponding capillary to achieve spectral signal normalization correction.

[0063] Specifically, during the testing process, it is possible to select whether to perform normalization correction on the spectral signal according to the testing requirements. If selected, after one electrophoresis is completed, through steps such as data preprocessing, peak identification, internal standard matching, and scoring for internal standard matching, qualified internal standard matching sequence data for peak normalization is selected. If it is unqualified, the data for this time is not calculated. The specific implementation method of normalization correction is to divide the standard value by the average value of the internal standard in a single channel of each capillary to obtain the normalization coefficient for each capillary. In this capillary, the spectral signals of other color channels are then multiplied by this coefficient to achieve the purpose of spectral signal normalization. Among them, the normalization coefficient should be within a certain value range. If there are capillaries that exceed the value range of the normalization coefficient but have not reached the condition for correcting the internal standard standard value, when normalizing the spectral signals of other color channels in the current capillary, the extreme value of the value range of the normalization coefficient closest to the normalization coefficient of the current capillary can be used as the normalization coefficient, and it is multiplied by the spectral signals of other color channels to achieve normalization correction.

[0064] In the embodiment of the present invention, before performing internal standard matching on the spectral signals of the internal standard channels of each capillary collected during the electrophoresis process of the gene analyzer, the method further includes: monitoring the Raman signal during the electrophoresis process to obtain the signal value, peak balance data, and / or uniformity data of the Raman signal; when any one of the signal value, peak balance data, and / or uniformity data of the Raman signal does not meet the corresponding preset optical parameter standard, perform an optical alignment operation until the signal value, peak balance data, and / or uniformity data all meet the corresponding optical parameter standards.

[0065] Specifically, each time the capillary consumable is replaced, automatic optical alignment is performed to reduce the influence of the optical system caused by the deviation in the production and installation of the capillary. If the capillary consumable is not replaced, the optical system will also be affected due to lens contamination, capillary contamination, vibration, etc. At this time, the Raman signal during the electrophoresis process can be monitored to determine whether there is an optical influence. The monitoring of the Raman signal specifically includes: judging the spectral signal value of each channel, peak balance (tilt) judgment, and uniformity judgment. If the spectral signal value of each channel is greater than the standard value, the standard value can be selected from 100 - 10000 according to the test scenario, and the peak balance data is less than the balance standard value, the balance standard value can be selected from 100 - 10000, and the uniformity data is greater than the uniformity standard value, the uniformity standard value can be selected from 0.1 - 1, uniformity = min / max, and the peak balance data is the absolute value of the difference between the average value of the left baseline (such as 560 - 590) and the average value of the right baseline (such as 630 - 660). Among them, min is the minimum value of the spectral signal values of each channel, and max is the maximum value of the spectral signal values of each channel. Each time electrophoresis is performed, if it is found that a certain value is unqualified, the optical alignment operation is automatically performed.

[0066] Further, the implementation process of the optical alignment operation includes:

[0067] Start the optical automatic alignment process, and the alignment motor runs to the optocoupler zero position.

[0068] The alignment motor moves towards the optocoupler limit position, and the movement speed is set by the lower computer, which can be optionally 1000um / s. During the movement of the alignment motor, the spectral data of the spectrometer is read in real time and displayed on the interface.

[0069] Taking four channels as an example, the detection values of the four channels are judged. If they are all greater than the product of the preset spatial correction standard value and the spatial correction optical alignment threshold, the slow scan of the alignment motor starts. Among them, the spatial correction standard value can be optionally 500, and the spatial correction optical alignment threshold can be optionally 50%. The calculation method of the detection value is as follows: find the maximum spectral intensity value between the sampling wavelength lower limit (610) and the sampling wavelength upper limit (620), and subtract the baseline value. The calculation method of the baseline: calculate the average value of the left baseline lower limit and the left baseline upper limit (left baseline, 560 - 590) and the average value of the right baseline lower limit and the right baseline upper limit (right baseline, 630 - 660) respectively, and then calculate the average value of the average value of the left baseline and the average value of the right baseline.

[0070] Each time the alignment motor moves the preset scanning motor single - movement step number threshold, the scanning motor single - movement step number threshold can be optionally 8 subdivisions, and the spectral signal values of the four channels are recorded in the optical automatic alignment file.

[0071] Judge the spectral signal value of one channel. If they are all less than the product of the spatial correction standard value and the spatial correction optical alignment threshold, end recording the spectral signal value of one channel, end the slow scan of the alignment motor, and at the same time start recording the single - step signal value file. If this value has not been found all the time, when the number of spectra taken exceeds 500 (for example, if the spatial correction is set to an integration time of 0.4 seconds, it is 200 seconds), still record the single - step signal value file and prompt "Failed to find the alignment position".

[0072] Calculate the spectral signal values of the four channels recorded in the optical automatic alignment file. Calculate the range coefficient with the spectral signals of the four channels at each position, select the positions with the range coefficient greater than the range coefficient threshold as the alternative positions, and select the one with the largest average value among all the alternative positions as the alignment position. Among them, the range coefficient = min / max, and the range coefficient can be used to evaluate whether the alignment is uniform.

[0073] The alignment motor moves to the alignment position. The number of movement steps is: the difference between the current position where the alignment motor stays and the number of steps of the alignment position.

[0074] Take the spectral signal value, that is, the average detection value within the preset acquisition time. The preset acquisition time can be selected as 10 seconds, and compare it with the spatial correction standard value. If the spectral signal value is greater than the spatial correction standard value, enter the peak lift judgment.

[0075] Peak tilt judgment: The average value of the left baseline lower limit and the left baseline upper limit (i.e., the left baseline, 560-590) minus the average value of the right baseline lower limit and the right baseline upper limit (i.e., the right baseline 630-660), take the absolute value, and compare it with the preset normalized symmetry value. The normalized symmetry value can be selected as 2000. If it is greater than, it is tilted and the judgment fails. If it is less than, it is judged that the spatial correction passes and the normalized value is calculated. If the spatial correction fails, it stays at the position where the sum of the four channels is the largest.

[0076] In the embodiment of the present invention, internal standard matching is performed on the spectral signal of the internal standard channel of each capillary collected during the electrophoresis process of the gene analyzer, specifically including the following steps not shown in the drawings:

[0077] S10, performing data pre-processing on the spectral signal of the internal standard channel of each capillary collected during the electrophoresis process of the gene analyzer to filter out the background noise of the spectral signal and perform smoothing on the spectral signal;

[0078] S11, performing peak recognition on the spectral signal after data pre-processing to screen out candidate peak sequences contained in the current spectral signal.

[0079] Specifically, step S11 specifically includes: identifying the maximum points in the spectral signal, calculating the peak characteristics of the peak signal corresponding to each maximum point, the peak characteristics including one or more characteristics of peak height, bottom peak width, half-height width, peak spacing and adjacent point drop height; screening the peak characteristics of the peak signal corresponding to each maximum point in the order of peak height, half-height width, peak spacing, adjacent point drop height and bottom peak width, and taking the peak signals that meet the corresponding threshold requirements of each peak characteristic as candidate peaks to form a candidate peak sequence.

[0080] In this embodiment, after obtaining the collected fluorescence spectrum signal, the maximum and minimum values ​​of all fluorescence intensities in the fluorescence spectrum signal are identified. The maximum value here is the peak value we are looking for, and the following characteristics of each peak are calculated:

[0081] Peak height: The height of the peak, which refers to the size of the peak;

[0082] Bottom peak width: the distance between the left and right boundaries of the peak;

[0083] Half-height width: Move from the peak to the left and right sides until the fluorescence intensity drops to half the height of the peak, and then measure the horizontal distance between the two positions;

[0084] Peak spacing: the horizontal distance between the highest points of two peaks;

[0085] Height drop of adjacent points: the height difference between the peak and adjacent points.

[0086] Set the threshold requirements for these features according to the test accuracy and the characteristics of the selected internal standard substance. As shown in Table 1, screen the peak points and leave the peak points that meet the requirements to form a candidate peak sequence.

[0087] Table 1 Threshold requirements

[0088]

[0089] In a specific embodiment, the screening rules are as follows:

[0090] Peak identification ==> Peak height ==> Full width at half maximum ==> Peak spacing ==> Height drop of adjacent points ==> Base peak width

[0091] In this embodiment, judge one by one through separate features. The judgment order is related to the data characteristics. For example, full width at half maximum ==> peak spacing. For these two orders, judge the full width at half maximum first to remove abnormal peaks. Some peaks with a small full width at half maximum are abnormal peaks, but the peak height is relatively high; in the judgment of peak spacing, the judgment is carried out in sequence according to the peak height. If abnormal peaks are not removed, some normal peaks will not be recognized at the peak spacing. Therefore, the order here cannot be reversed.

[0092] S12. In the order of sampling sequence, perform internal standard matching on the candidate peaks in the candidate peak sequence and the standard peak sequence in the standard spectral signal corresponding to the internal standard substance selected in the electrophoresis process, so as to find the candidate peak combination that meets the preset internal standard matching conditions in the candidate peak sequence. The number of candidate peaks in the candidate peak combination is greater than or equal to 3; the internal standard matching conditions include that the absolute value of the difference between the first distance and the second distance is less than the preset distance error threshold, and max{the peak height of the selected candidate peaks in the candidate peak combination, the peak height of the current candidate peak to be matched} / min{the peak height of the selected candidate peaks in the candidate peak combination, the peak height of the current candidate peak to be matched}<the preset relative height threshold, where the first distance is the distance between adjacent candidate peak points in the candidate peak sequence, and the second distance is the distance between adjacent standard peak points in the standard peak sequence with the same sampling order as the candidate peak for which the first distance is currently calculated.

[0093] Specifically, step S12 specifically includes: obtaining the distance between any adjacent candidate peak points in the candidate peak sequence, and obtaining the distance between any adjacent standard peak points in the standard peak sequence; matching the corresponding number of candidate peaks that meet the preset internal standard matching conditions from the candidate peak sequence according to the number of candidate peaks included in the preset candidate peak combination in the order of sampling sequence to obtain a candidate peak combination.

[0094] In the embodiments of the present invention, when performing internal standard matching, two indicators, namely an acceptable distance error and a relative height, are introduced, and internal standard matching is performed according to an internal standard matching mode that requires simultaneously meeting distance similarity and height uniformity. Among them, the distance error can be a distance length error or a ratio error ratioD, and the relative height is the relative peak height relativeH. In a specific embodiment, the ratio error ratioD is used as the distance error, and the relative peak height relativeH is used as the relative height for explanation. Optionally, the ratio error generally takes a value less than 2; the relative peak height relativeH takes a value greater than 1.

[0095] The following is an explanation of the ratio error ratioD. Refer to Figure 2 , the internal standard (standard) is known, and the standard peak sequence is as shown in Figure 2 in red, and the candidate peak sequence is as shown in Figure 2 in blue. Calculate the values of a1, a2, ···. In the electrophoresis data of the standard product, that is, the candidate peak sequence, if |ai - bi| < ratioD, it is considered that the distance corresponding to bi meets the requirements. Here, ratioD is the acceptable ratio error, and ai and bi respectively represent the coefficients in the distance relationship between the standard internal standard and the identified internal standard, and "||" represents the absolute value calculation symbol.

[0096] The following is an explanation of the relative peak height relativeH. Each time a new peak (candidate peak) is added or newly matched, it is necessary to satisfy max{the peak height of the selected peaks, the peak height of the candidate peak} / min{the peak height of the selected peaks, the peak height of the candidate peak} < relativeH.

[0097] Among them, the threshold of the ratio error ratioD is matchTH, which represents the maximum acceptable ratio error; the threshold of the relative peak height relativeH is evennessTH, which represents the maximum acceptable relative peak height error. Here, relativeH is obtained from evennessTH through a certain threshold adjustment rule or function conversion, and ratioD is obtained from matchTH through a certain threshold adjustment rule or function conversion.

[0098] S13. Using each candidate peak combination as the matching basis, perform internal standard matching on other candidate peaks in the candidate peak sequence in turn to obtain the matching result corresponding to each candidate peak combination.

[0099] S14. Count the number of candidate peaks included in the matching result corresponding to each candidate peak combination.

[0100] S15. When the maximum value of the number of candidate peaks included in the matching results corresponding to each candidate peak combination is equal to the number of standard peaks included in the standard peak sequence, it is determined that the internal standard matching is successful, and the matching result corresponding to the maximum value is used as the optimal internal standard matching result.

[0101] Regarding the step S15 of using the matching result corresponding to the maximum value as the optimal internal standard matching result, it specifically includes: if there is only one matching result corresponding to the maximum value, then the matching result corresponding to the maximum value is used as the optimal internal standard matching result; if there are more than one matching results corresponding to the maximum value, then the matching result with the largest sampling point position of the first candidate peak among the matching results corresponding to the maximum value is used as the optimal internal standard matching result.

[0102] In the embodiment of the present invention, by performing peak recognition on the spectral signal to screen out the candidate peak sequences that meet the requirements included in the current spectral signal, and then based on two internal standard matching conditions of distance similarity and height uniformity, the candidate peaks in the candidate peak sequence are sequentially subjected to internal standard matching with the standard peak sequence in the standard spectral signal corresponding to the internal standard substance selected during the electrophoresis process in the order of sampling, so as to find the candidate peak combinations that meet the preset internal standard matching conditions in the candidate peak sequence, and use the candidate peak combinations as the initial part of the matching result, and use this as the matching basis to sequentially perform internal standard matching on other candidate peaks in the candidate peak sequence to obtain the complete matching results corresponding to each candidate peak combination. When the number of candidate peaks included in the obtained matching results is equal to the number of standard peaks included in the standard peak sequence, it is determined that the internal standard matching is successful and the current matching result is used as the optimal internal standard matching result. The present invention can quickly and accurately achieve the optimal matching of the internal standard peak, thereby ensuring the accuracy of the spectral detection fragment length.

[0103] In the embodiment of the present invention, after using the matching result corresponding to the maximum value as the optimal internal standard matching result, the method includes an internal standard matching degree scoring operation. The specific implementation is as follows: perform curve fitting on the standard peak sequence and the peak sequence of the optimal internal standard matching result, and use the standard deviation, average residual, and maximum residual after curve fitting as the characteristic data of the current optimal internal standard matching result; input the characteristic data into a preset internal standard matching scoring model for recognition to obtain the matching degree score of the current optimal internal standard matching result.

[0104] Specifically, in the case where the internal standard matching is successful in the present invention, the Local Southern method or the high-order polynomial fitting method can be used to perform curve fitting on the standard peak sequence and the peak sequence of the internal standard matching result, and select the standard deviation, average residual, and maximum residual after curve fitting as the three characteristic data for each matching situation, and then input the characteristic data into a preset internal standard matching scoring model to perform internal standard matching degree scoring.

[0105] Further, the method further includes a training step of an internal standard matching scoring model, specifically including: taking the standard deviation, average residual, and maximum residual after curve fitting of the sample data corresponding to different internal standard matching cases preset as the sample feature data of the corresponding samples, setting the sample data with correct internal standard matching results as the positive class, and setting the sample data with incorrect internal standard matching results as the negative class to obtain a training data set; using the Hinge loss as the loss function for model training, normalizing the classification results using the Sigmoid function, and performing learning and training on the training data set based on a preset machine learning model to obtain a trained internal standard matching scoring model.

[0106] Specifically, the present invention calculates the above three feature data of a large number of samples corresponding to different internal standards, sets the correct matching as the positive class, and the incorrect matching as the negative class to form a training data set, selects the Hinge loss as the loss function for model training, uses a machine learning model such as a support vector machine to train the training data set to obtain model parameters, and finally uses the sigmoid function to further process the results to obtain a trained internal standard matching scoring model.

[0107] In an optional embodiment of the present invention, the method further includes: if the maximum value of the number of candidate peaks included in the matching result corresponding to each candidate peak combination is not equal to the number of standard peaks included in the standard peak sequence, updating the distance error threshold according to a preset first threshold adjustment rule, and returning to step S12 to perform internal standard matching again until the updated distance error threshold is greater than the maximum value of the distance error threshold;

[0108] When the updated distance error threshold is greater than the maximum value of the distance error threshold, updating the relative height threshold according to a preset second threshold adjustment rule, updating the distance error threshold to the corresponding initial value, and returning to step S12 to perform internal standard matching again until the updated relative height threshold is greater than the maximum value of the relative height threshold;

[0109] When the updated relative height threshold is greater than the maximum value of the relative height threshold, it is determined that the internal standard matching fails.

[0110] In the embodiment of the present invention, the dynamic matching of the internal standard is realized by dynamically adjusting the height threshold and the distance error threshold within a preset value range. Specifically, if the internal standard cannot be successfully matched under the state of the initial height threshold and the distance error threshold, in order to avoid the matching failure caused by inappropriate threshold selection, the present invention can dynamically adjust the two types of thresholds respectively. First, the distance error threshold is dynamically increased on the premise of keeping the relative height threshold unchanged, such as adjusting a preset unit step length each time, and the internal standard matching is carried out again under the state of the current height threshold and the adjusted distance error threshold until the internal standard is successfully matched or the updated distance error threshold is greater than the maximum value of the distance error threshold. When the updated distance error threshold is greater than the maximum value of the distance error threshold, the height threshold is dynamically increased, and under the state of the adjusted height threshold, the internal standard matching is restarted with the initial distance error threshold until the internal standard is successfully matched or the updated relative height threshold is greater than the maximum value of the relative height threshold. By using the dynamic programming of the inner and outer two-layer threshold loops to match the identified internal standard peak with the standard internal standard length, the accuracy of the internal standard matching can be further ensured.

[0111] In the embodiment of the present invention, data preprocessing is performed on the spectral signals of the internal standard channels of each capillary collected during the electrophoresis process of the gene analyzer, which specifically includes the following steps:

[0112] S01. Filter the background noise of the spectral signal by using a preset locally adaptive polynomial fitting algorithm.

[0113] Specifically, first segment the data, and then perform polynomial fitting in the following manner:

[0114] initial: k = 1, O0(i) = O(i);

[0115] step1: Fit the signal (1, 2, 3,.....) with a polynomial of appropriate order to obtain P k (i);

[0116] step2: Calculate the residual R k (i) = O k-1 (i) - P i (i);

[0117] step3: Calculate the standard deviation DEV of the residual k ;

[0118] step4: If it is the first iteration, delete the signals that meet the condition O0(i) > P k (i) + DEV k (remove the corresponding subscripts); if it is not the first iteration, assign values according to the following rules:

[0119] if O k+1 (i) < P k (i) + DEV k , O k (i) = O k-1 (i)

[0120] otherwise, O k (i) = P k (i) + DEV k

[0121] step5: Calculate whether it satisfies |(DEV k - DDV k-1 ) / DEV k | < gradient, where gradient = 0.01 and the parameter is adjustable. If it is satisfied, continue to the next step. If not, return to step1 and repeat this process;

[0122] step6: Calculate the baseline P k (i) using the polynomial coefficients, and output the signal after removing the baseline S(i) = O0(i) - P k (i).

[0123] Where: K is the number of iterations; i is the spectral data point, i = 0, 1, 2,..., N - 1; O is the original signal; P is the polynomial fitting signal (baseline); R is the residual between the original signal and the fitting signal; DEV is the standard deviation; O k (i) is the i-th data in the original spectral data at the k-th iteration; P k (i) is the i-th data in the polynomial fitting data at the k-th iteration; R k (i) is the residual of the i-th data between the original spectral data and the fitting data at the k-th iteration; DEV k is the standard deviation at the k-th iteration; gradient is the stop iteration threshold; S(i) is the i-th data of the data after removing the baseline (the data used for the next processing).

[0124] S02. Use the Savitzky Golay polynomial smoothing algorithm to smooth the spectral signal after removing the background noise.

[0125] Specifically, at each data point S i , use a window of length 2n + 1 to perform polynomial fitting on the data within the window. The common polynomial order is p (p = 1, 2, 3,...). Finally, use the value of the center point of the fitting polynomial to replace the original data point S i , thus completing the smoothing process.

[0126] Specifically, the formula for the Savitzky-Golay filter is as follows:

[0127]

[0128] where Hi is the smoothed data point, S i is the original data point, and c j is the coefficient of the filter. The calculation of the filter coefficients involves the process of polynomial fitting and can be solved by methods such as the least squares method.

[0129] Due to problems such as laser background, substrate Raman signal, and spectrometer dark noise, the signal baselines collected in different bands are inconsistent, resulting in the inability to directly determine the type and content of DNA based on the signal intensity. Therefore, the present invention processes the original data through methods such as baseline removal and smoothing, which can ensure the accurate identification of candidate peaks.

[0130] In a specific embodiment of the present invention, the method for realizing the internal standard matching of the detection spectrum of a gene analyzer is as Figure 3 shown. Each identified peak is represented as (pi, H pi ), indicating that the signal intensity of the pi-th data point in the spectral data is H pi , where p0 < p1 < p2 < ··· < pi < ···. All identified peaks are called the candidate peak sequence {(p0, H p0 ), (p1, H p1 ), ···, (pi, H pi ), ···}.

[0131] The peaks corresponding to the standard internal standard length are called the standard peak sequence {s0, s1, s2, ···, si, ···}, where s0 < s1 < s2 < ··· < si < ···.

[0132] 1. Calculate the distance differences between all adjacent data points in the candidate peak sequence, denoted as C = {C0, C1, ···} = {p1 - p0, p2 - p1, ···};

[0133] 2. Calculate the differences between all adjacent lengths in the standard peak sequence, denoted as S = {S0, S1, ···} = {s1 - s0, s2 – s1, ···};

[0134] 3. Initialize the relative height ralativeH;

[0135] 4. Initialize the acceptable proportional error ratioD;

[0136] 5. In the candidate peak sequence, in the order of matching data points from small to large, find a preset number of candidate peaks that meet the internal matching conditions, and find all possible combinations; among them, the preset number can be 3;

[0137] 6. Next, complete all the combined matches. At this time, it is possible that all the detected peaks have been judged, but the internal standard has not been fully matched;

[0138] 7. Calculate the number of peaks matched in each combination and take the maximum value;

[0139] 8. Determine whether this maximum value is equal to the number of internal standards;

[0140] 9. If it is equal to the number of internal standards, the internal standard is successfully matched, and the combination corresponding to the maximum value and with the largest data point of the first peak is output; if not, proceed to step 10

[0141] 10. Update ratioD and determine whether the updated ratioD is less than matchTH. If it is less, return to step 5; otherwise, proceed to step 11;

[0142] 11. Increase the relative height ralativeH by 1. Determine whether the increased ralativeH exceeds the relative peak height threshold. If not, return to step 4 to repeat the matching process; otherwise, the internal standard matching fails, the analysis process ends, and the matching score is equal to 0.

[0143] The present invention adopts a local adaptive fluorescence background noise removal algorithm, segments the data, removes the baseline through polynomial fitting and performs smoothing processing, and then screens the peak points by setting thresholds corresponding to features for the processed data to leave candidate peak points that meet the requirements. Through two matching conditions of distance similarity and height uniformity, dynamic programming is used to match the identified internal standard peaks with the standard internal standard length. In the case of successful internal standard matching, the standard peak sequence and the matched sequence are curve-fitted, and the standard deviation, average residual, and maximum residual after curve fitting are selected as the three characteristic data for each matching case. Then, the characteristic data is input into a preset internal standard matching scoring model to score the degree of internal standard matching, which can not only accurately achieve internal standard matching but also intelligently score the degree of internal standard matching, improving the intelligence and accuracy of the electrophoresis spectrum data analysis process.

[0144] For the method embodiments, for the sake of simple description, they are all expressed as a series of action combinations. However, those skilled in the art should know that the embodiments of the present invention are not limited by the described action sequences, because according to the embodiments of the present invention, certain steps can be performed in other sequences or simultaneously. Secondly, those skilled in the art should also know that the embodiments described in the specification are all preferred embodiments, and the actions involved are not necessarily essential for the embodiments of the present invention.

[0145] In addition, an embodiment of the present invention further provides a system for realizing the normalization of detection signals of a gene analyzer. The system includes functional modules for realizing the method for realizing the normalization of detection signals of a gene analyzer as described above. As Figure 4 shown, the system for realizing the normalization of detection signals of an embodiment of the present invention includes an internal standard matching unit 40, a calculation unit 50, and a normalization unit 60, where:

[0146] The internal standard matching unit 40 is configured to perform internal standard matching on the spectral signals of the internal standard channels of each capillary collected during the electrophoresis process of the gene analyzer, and calculate the single-channel internal standard average value of the internal standard channels of each capillary according to the successfully matched internal standard matching sequences;

[0147] The calculation unit 50 is configured to divide the preset internal standard standard value by the single-channel internal standard average value of the internal standard channels of each capillary respectively, and use the obtained calculation result as the normalization coefficient corresponding to the capillary;

[0148] The normalization unit 60 is configured to perform normalization correction on the spectral signals of other color channels in each capillary respectively according to the normalization coefficient of each capillary, so as to realize the normalization of detection signals.

[0149] In an embodiment of the present invention, the internal standard matching unit is further configured to perform electrophoresis on multiple capillaries filled with only a specified concentration of internal standard before performing internal standard matching on the spectral signals of the internal standard channels of each capillary collected during the electrophoresis process of the gene analyzer, so as to collect the spectral signals of the internal standard channels of each capillary and perform internal standard matching, calculate the average peak value of each internal standard peak in the internal standard matching sequence that meets the internal standard matching requirements corresponding to each capillary, and obtain the single-channel internal standard average value of the internal standard channels of each capillary;

[0150] The calculation unit is further configured to calculate the coefficient of variation of the single-channel internal standard average value of the internal standard channels of each capillary. If the coefficient of variation is less than a preset variation threshold, the average value or a preset multiple of the average value of the single-channel internal standard average value of the internal standard channels of each capillary is used as the internal standard standard value.

[0151] In an embodiment of the present invention, the system further includes a judgment unit and a reset unit. Among them, the judgment unit is configured to judge whether the normalization coefficient of each capillary is compliant after the calculation unit uses the obtained calculation result as the normalization coefficient corresponding to the capillary. When the normalization coefficient of the capillary is within the preset normalization coefficient value range, it is determined that the normalization coefficient of the current capillary is compliant, otherwise it is determined that the normalization coefficient of the current capillary is non-compliant;

[0152] The reset unit is further configured to correct the internal standard standard value if the proportion of the number of capillaries with non-compliant normalization coefficients in the current electrophoresis detection data to the total number of capillaries is greater than a preset first proportion threshold, or if the proportion of the number of capillaries with non-compliant normalization coefficients in at least three consecutive electrophoresis detection data to the total number of capillaries is greater than a preset second proportion threshold;

[0153] Wherein, the second proportion threshold is less than the first proportion threshold.

[0154] Further, the normalization unit 60 is specifically configured to multiply the spectral signals of other color channels in the current capillary by the normalization coefficient of the corresponding capillary to perform spectral signal normalization correction if the normalization coefficient of the capillary is compliant; if the normalization coefficient of the capillary is non-compliant, multiply the spectral signals of other color channels in the current capillary by the extreme value close to the value range of the normalization coefficient to perform spectral signal normalization correction.

[0155] In an embodiment of the present invention, the system further includes an optical alignment unit, configured to monitor the Raman signal during the electrophoresis process, obtain the signal value, peak balance data, and / or uniformity data of the Raman signal before performing internal standard matching on the spectral signals of the internal standard channels of each capillary collected during the electrophoresis process of the gene analyzer; when any one of the signal value, peak balance data, and / or uniformity data of the Raman signal does not meet the corresponding preset optical parameter standard, perform an optical alignment operation until the signal value, peak balance data, and / or uniformity data all meet the corresponding optical parameter standards.

[0156] In an embodiment of the present invention, the internal standard matching unit 40 includes a data preprocessing module 400, a peak identification module 401, an initial matching module 402, an internal standard matching module 403, a statistics module 404, and a determination module 405 not shown in the drawings, wherein:

[0157] The data preprocessing module 400 is configured to perform data preprocessing on the spectral signals of the internal standard channels of each capillary collected during the electrophoresis process of the gene analyzer to filter out the background noise of the spectral signals and smooth the spectral signals;

[0158] The peak identification module 401 performs peak identification on the spectral signals after data preprocessing to screen out the candidate peak sequences included in the current spectral signals;

[0159] The initial matching module 402 performs internal standard matching on the candidate peaks in the candidate peak sequence and the standard peak sequence in the standard spectral signals corresponding to the internal standard substances selected during the electrophoresis process in the order of sampling, so as to find the candidate peak combinations that meet the preset internal standard matching conditions in the candidate peak sequence, and the number of candidate peaks in the candidate peak combination is greater than or equal to 3; the internal standard matching conditions include that the absolute value of the difference between the first distance and the second distance is less than the preset distance error threshold, and max{the peak height of the selected candidate peaks in the candidate peak combination, the peak height of the currently to-be-matched candidate peak} / min{the peak height of the selected candidate peaks in the candidate peak combination, the peak height of the currently to-be-matched candidate peak}<the preset relative height threshold, where the first distance is the distance between the peak value points of adjacent candidate peaks in the candidate peak sequence, and the second distance is the distance between the peak value points of adjacent standard peaks in the standard peak sequence with the same sampling order as the candidate peak for which the first distance is currently calculated;

[0160] The internal standard matching module 403 performs internal standard matching on the other candidate peaks in the candidate peak sequence with each candidate peak combination as the matching basis, and obtains the matching result corresponding to each candidate peak combination;

[0161] The statistics module 404 counts the number of candidate peaks included in the matching result corresponding to each candidate peak combination;

[0162] The determination module 405 determines that the internal standard matching is successful when the maximum value of the number of candidate peaks included in the matching results corresponding to each candidate peak combination is equal to the number of standard peaks included in the standard peak sequence, and takes the matching result corresponding to the maximum value as the optimal internal standard matching result.

[0163] The internal standard matching unit 40 provided by the embodiment of the present invention further includes an intelligent scoring module not shown in the drawings. The intelligent scoring module is used to, after taking the matching result corresponding to the maximum value as the optimal internal standard matching result, perform curve fitting on the standard peak sequence and the peak sequence of the optimal internal standard matching result, and use the standard deviation, average residual, and maximum residual after curve fitting as the characteristic data of the current optimal internal standard matching result; input the characteristic data into a preset internal standard matching scoring model for identification to obtain the matching degree score of the current optimal internal standard matching result.

[0164] Specifically, in the case of successful internal standard matching in the present invention, a high-order polynomial fitting method can be used to implement curve fitting of the standard peak sequence and the peak sequence of the internal standard matching result, and the standard deviation, average residual, and maximum residual after curve fitting are selected as the three characteristic data for each matching case, and then the characteristic data are input into a preset internal standard matching scoring model for internal standard matching degree scoring.

[0165] Furthermore, the internal standard matching unit 40 provided by the embodiment of the present invention further includes a model training module not shown in the drawings, which is used to perform the training operation of the internal standard matching scoring model. Specifically, it includes: using the standard deviation, average residual, and maximum residual after curve fitting of the sample data corresponding to different internal standard matching cases preset as the sample feature data of the corresponding samples, setting the sample data with the correct internal standard matching result as the positive class, and setting the sample data with the incorrect internal standard matching result as the negative class to obtain a training data set; using the Hinge loss as the loss function for model training, normalizing the classification result using the Sigmoid function, and performing learning and training on the training data set based on a preset machine learning model to obtain a trained internal standard matching scoring model.

[0166] Specifically, the present invention calculates the above three feature data of a large number of samples corresponding to different internal standards, sets the correct matching as the positive class, and the incorrect matching as the negative class to form a training data set. It selects the Hinge loss as the loss function for model training, uses a machine learning model such as a support vector machine to train the training data set to obtain model parameters, and finally uses the sigmoid function to further process the results to obtain a trained internal standard matching scoring model.

[0167] The internal standard matching unit 40 provided by the embodiment of the present invention further includes a threshold dynamic adjustment module not shown in the drawings, which is used to update the distance error threshold according to a preset first threshold adjustment rule and return to the initial matching module 402 to perform corresponding operations when the maximum value of the number of candidate peaks included in the matching result corresponding to each candidate peak combination is not equal to the number of standard peaks included in the standard peak sequence, until the distance error threshold updated by the threshold dynamic adjustment module is greater than the maximum value of the distance error threshold;

[0168] The threshold dynamic adjustment module is further used to update the relative height threshold according to a preset second threshold adjustment rule when the updated distance error threshold is greater than the maximum value of the distance error threshold, update the distance error threshold to the corresponding initial value, and return to the initial matching module 402 to perform corresponding operations until the relative height threshold updated by the threshold dynamic adjustment module is greater than the maximum value of the relative height threshold;

[0169] The determination module 405 is further used to determine that the internal standard matching fails when the updated relative height threshold is greater than the maximum value of the relative height threshold.

[0170] For the system embodiment, since it is basically similar to the method embodiment, the description is relatively simple. For the relevant parts, refer to the partial description of the method embodiment, and it has corresponding technical effects.

[0171] In addition, an embodiment of the present invention further provides a computer-readable storage medium, on which a computer program is stored. When the computer program is executed by a processor, the steps of the method for normalizing the detection signal of a gene analyzer are implemented as described above.

[0172] In this embodiment, if the method for normalizing the detection signal of a gene analyzer is implemented in the form of a software functional unit and sold or used as an independent product, it can be stored in a computer-readable storage medium. Based on such an understanding, to implement all or part of the processes in the above-described embodiment methods of the present invention, it can also be completed by a computer program instructing relevant hardware. The computer program can be stored in a computer-readable storage medium. When the computer program is executed by a processor, the steps of the above-described method embodiments can be implemented. Among them, the computer program includes computer program code, and the computer program code can be in the form of source code, object code, executable file, or some intermediate form, etc. The computer-readable medium can include: any entity or device capable of carrying the computer program code, recording medium, USB flash drive, mobile hard disk, magnetic disk, optical disc, computer memory, read-only memory (ROM, Read-Only Memory), random access memory (RAM, Random Access Memory), electrical carrier signal, telecommunication signal, and software distribution medium, etc. It should be noted that the content included in the computer-readable medium can be appropriately increased or decreased according to the requirements of legislation and patent practice in the jurisdiction. For example, in some jurisdictions, according to legislation and patent practice, the computer-readable medium does not include electrical carrier signals and telecommunication signals.

[0173] In addition, an embodiment of the present invention further provides a computer device, including a memory, a processor, and a computer program stored on the memory and executable on the processor. When the processor executes the computer program, the steps of the method for normalizing the detection signal of a gene analyzer are implemented as described above. For example Figure 1 Steps S1 to S3 shown. Alternatively, when the processor executes the computer program, the functions of each module / unit in the above-described system embodiment for normalizing the detection signal of a gene analyzer are implemented. For example Figure 4 The internal standard matching unit 40, calculation unit 50, and normalization unit 60 shown.

[0174] The method, system and device for realizing the normalization of detection signals of a gene analyzer provided by the embodiments of the present invention calculate the normalization coefficient of each capillary according to the preset internal standard standard value and the single-channel internal standard average value of the internal standard channel of each capillary during the electrophoresis process of the gene analyzer, and perform normalization correction on the spectral signals of other color channels in each capillary according to the normalization coefficient of each capillary, so as to realize the adjustment of the peak height of the sample to be tested according to the peak height of the internal standard, improve the signal difference between different capillaries and the difference in different detection times of the same capillary, and effectively improve the quality of gene analysis results.

[0175] In addition, those skilled in the art can understand that although some embodiments herein include certain features included in other embodiments rather than other features, the combination of the features of different embodiments means that it is within the scope of the present invention and forms different embodiments. For example, any one of the claimed embodiments can be used in any combination.

[0176] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention and are not intended to limit them. Although the present invention has been described in detail with reference to the foregoing embodiments, those of ordinary skill in the art should understand that they can still modify the technical solutions described in the foregoing embodiments, or perform equivalent replacements on some of the technical features. And these modifications or replacements do not cause the essence of the corresponding technical solutions to deviate from the spirit and scope of the technical solutions of the embodiments of the present invention.

Claims

1. A method for realizing normalization of detection signals of a gene analyzer, characterized in that: The method comprises: Performing internal standard matching on the spectral signal of the internal standard channel of each capillary collected during the electrophoresis process of the gene analyzer, and calculating the single-channel internal standard average value of each capillary internal standard channel according to the successfully matched internal standard matching sequence; The preset internal standard value is divided by the single channel internal standard average value of the internal standard channel of each capillary, and the calculated result is used as the normalization coefficient of the corresponding capillary; The spectral signals of other color channels in each capillary are normalized and corrected according to the normalization coefficient of each capillary to achieve normalization of the detection signal. The spectral signals of other color channels are obtained by collecting the fluorescence generated by laser excitation of fluorescent groups on DNA.

2. The method according to claim 1, characterized in that: Before performing internal standard matching on the spectral signals of the internal standard channel of each capillary collected during the electrophoresis process of the gene analyzer, the method further comprises: Performing electrophoresis on a multi-channel capillary tube containing only an internal standard substance of a specified concentration to collect the spectral signal of the internal standard substance channel of each capillary tube and perform internal standard matching, calculating the average peak value of each internal standard peak in the internal standard matching sequence corresponding to each capillary tube that meets the internal standard matching requirements, and obtaining the single-channel internal standard average value of each capillary internal standard substance channel; The coefficient of variation of the single-channel internal standard average value of each capillary internal standard channel is calculated. If the coefficient of variation is less than a preset variation threshold, the mean of the single-channel internal standard average values ​​of each capillary internal standard channel or a preset multiple of the mean is used as the internal standard standard value.

3. The method according to claim 1, characterized in that After using the obtained calculation result as the normalization coefficient of the corresponding capillary, the method further includes: Determine whether the normalization coefficient of each capillary is compliant. When the normalization coefficient of the capillary is within the preset normalization coefficient value range, the normalization coefficient of the current capillary is determined to be compliant. Otherwise, the normalization coefficient of the current capillary is determined to be non-compliant. If the ratio of the number of capillaries with non-compliant normalization coefficients in the current electrophoresis detection data to the total number of capillaries is greater than a preset first ratio threshold, or if the ratio of the number of capillaries with non-compliant normalization coefficients in the electrophoresis detection data for at least three consecutive times to the total number of capillaries is greater than a preset second ratio threshold, the internal standard value is corrected; The second ratio threshold is smaller than the first ratio threshold.

4. The method according to claim 3, characterized in that: The normalization correction of the spectral signals of other color channels in the respective capillaries according to the normalization coefficient of each capillary comprises: If the normalization coefficient of the capillary is in compliance, the spectral signals of other color channels in the current capillary are multiplied by the normalization coefficient of the corresponding capillary to achieve spectral signal normalization correction; If the normalization coefficient of the capillary is not compliant, the spectral signals of other color channels in the current capillary are multiplied by the normalization coefficient of the corresponding capillary close to the extreme value within the normalization coefficient value range to achieve spectral signal normalization correction.

5. The method according to claim 1, characterized in that Before performing internal standard matching on the spectral signal of the internal standard channel of each capillary collected during the electrophoresis process of the gene analyzer, the method further comprises: Monitoring the Raman signal during the electrophoresis process, and obtaining the signal value, peak balance data and / or uniformity data of the Raman signal; When any parameter of the signal value, peak balance data and / or uniformity data of the Raman signal does not meet the corresponding preset optical parameter standard, an optical alignment operation is performed until the signal value, peak balance data and / or uniformity data all meet the corresponding optical parameter standard.

6. The method according to any one of claims 1 to 5, characterized in that: The internal standard matching of the spectral signal of the internal standard channel of each capillary collected during the electrophoresis process of the gene analyzer includes: S10, performing data pre-processing on the spectral signal of the internal standard channel of each capillary collected during the electrophoresis process of the gene analyzer to filter out the background noise of the spectral signal and perform smoothing on the spectral signal; S11, performing peak recognition on the spectral signal after data pre-processing to screen out candidate peak sequences contained in the current spectral signal; S12. Perform internal standard matching on the candidate peaks in the candidate peak sequence and the standard peak sequence in the standard spectral signal corresponding to the internal standard selected in the electrophoresis process in sequence according to the sampling order, so as to find the candidate peak combination in the candidate peak sequence that meets the preset internal standard matching conditions, and the number of candidate peaks in the candidate peak combination is greater than or equal to 3; the internal standard matching conditions include that the absolute value of the difference between the first distance and the second distance is less than the preset distance error threshold, and max{peak height of the selected candidate peak in the candidate peak combination, peak height of the current candidate peak to be matched} / min{peak height of the selected candidate peak in the candidate peak combination, peak height of the current candidate peak to be matched}<preset relative height threshold, wherein the first distance is the distance between the peak points of adjacent candidate peaks in the candidate peak sequence, and the second distance is the distance between the peak points of adjacent standard peaks in the standard peak sequence that have the same sampling order as the candidate peak for calculating the first distance currently; S13, using each candidate peak combination as a matching basis, sequentially performing internal standard matching on other candidate peaks in the candidate peak sequence to obtain a matching result corresponding to each candidate peak combination; S14, counting the number of candidate peaks included in the matching results corresponding to each candidate peak combination; S15. When the maximum number of candidate peaks contained in the matching results corresponding to each candidate peak combination is equal to the number of standard peaks contained in the standard peak sequence, the internal standard matching is determined to be successful, and the matching result corresponding to the maximum value is used as the optimal internal standard matching result.

7. The method according to claim 6, characterized in that After taking the matching result corresponding to the maximum value as the optimal internal standard matching result, the method includes: Perform curve fitting on the standard peak sequence and the peak sequence of the optimal internal standard matching result, and use the standard deviation, average residual and maximum residual after the curve fitting as characteristic data of the current optimal internal standard matching result; The characteristic data is input into a preset internal standard matching scoring model for identification to obtain a matching degree score of the current optimal internal standard matching result.

8. The method according to claim 6, characterized in that The method further comprises: If the maximum number of candidate peaks included in the matching results corresponding to each candidate peak combination is not equal to the number of standard peaks included in the standard peak sequence, the distance error threshold is updated according to the preset first threshold adjustment rule, and the process returns to step S12 until the updated distance error threshold is greater than the maximum value of the distance error threshold; When the updated distance error threshold is greater than the maximum value of the distance error threshold, the relative height threshold is updated according to the preset second threshold adjustment rule, and the distance error threshold is updated to the corresponding initial value, and the process returns to step S12 until the updated relative height threshold is greater than the maximum value of the relative height threshold; When the updated relative height threshold is greater than the maximum value of the relative height threshold, it is determined that the internal standard matching fails.

9. A system for realizing normalization of detection signals of a gene analyzer, characterized in that: The system comprises a functional module for implementing the method according to any one of claims 1 to 8.

10. A computer device, characterized in that: The method comprises a memory, a processor and a computer program stored in the memory and executable on the processor, wherein the processor implements the steps of the method according to any one of claims 1 to 8 when executing the computer program.

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