High-throughput method and device for quantifying fecundity of drosophila melanogaster and porous cell culture equipment
By culturing fruit flies in porous cell culture equipment and using image processing technology to identify egg counts, the problems of inefficiency and poor accuracy in traditional methods are solved, and diversified analysis of fruit flies' egg laying ability and high-throughput counting are achieved. It is suitable for fruit flies research in a variety of culture media and genetic backgrounds.
Patent Information
- Application Number
- CN202510488111.X
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- Filing Date
- 2025-04-18
- Publication Date
- 2025-07-22
AI Technical Summary
Traditional fruit fly egg counting methods are inefficient, have poor data accuracy and consistency, and can only determine the egg-laying ability of fruit fly under a single medium.
By setting up multiple male and female fruit fly pairs to be detected in the porous cell culture device to be cultured in different types of culture media, initial egg laying images are obtained, and the target egg number is determined through image preprocessing and recognition technology, diversified analysis of fruit fly egg laying ability is achieved.
It improves the accuracy and consistency of egg data and improves the efficiency of determining egg count. It is suitable for fruit fly reproduction studies in a variety of culture media and genetic backgrounds, supporting genetic, behavioral and physiological research.
Smart Images
Figure CN120355692A_ABST
Abstract
Description
Technical Field
[0001] This application relates to the field of bioinformatics technology, and particularly to a high-throughput method, device and porous cell culture equipment for quantitatively measuring Drosophila fecundity. Background Art
[0002] At present, Drosophila, as a classical experimental animal, has a short life cycle and strong reproductive ability. Therefore, Drosophila reproduction plays an important role in biological research. In the prior art, Drosophila reproduction research is widely applied in multiple research fields such as genetics, pharmacology, and environmental biology. In these studies, the measurement of Drosophila fecundity can not only reflect the health status of Drosophila, but also reveal many influencing factors such as physiological mechanisms, genetic variations, and the environment. Moreover, within the first four weeks after Drosophila adulthood, there is usually a strong egg-laying ability. Therefore, it is particularly important to study the egg-laying ability of Drosophila and accurately determine the number of Drosophila eggs.
[0003] However, the traditional method for counting Drosophila eggs relies on experimenters to observe and count each well under a microscope. This process is not only inefficient but also unable to meet the need for continuous egg data collection. The counting results of the traditional method are easily affected by the experience, judgment of the observer, and environmental factors, and problems such as missed counting and repeated counting are likely to occur, resulting in poor data accuracy and consistency. Moreover, the traditional method for counting Drosophila eggs can only determine the egg-laying ability of Drosophila under a single culture medium. Summary of the Invention
[0004] The embodiments of this application provide a high-throughput method, device and porous cell culture equipment for quantitatively measuring Drosophila fecundity. The embodiments provided in this application solve the technical problems in the prior art that the egg calculation data is affected by environmental factors, resulting in inaccurate base numbers, low efficiency, poor consistency, and the ability to only determine the egg-laying ability of Drosophila under a single culture medium. The embodiments provided in this application realize the diversified analysis of Drosophila egg-laying ability by setting multiple pairs of male and female Drosophila to be detected and culturing them in different types of culture media, and improve the accuracy and consistency of egg data and the efficiency of determining the number of eggs through image recognition of the egg-laying images.
[0005] In the first aspect of the embodiments of this application, the embodiments of this application provide a high-throughput method for quantitatively measuring Drosophila fecundity, and the high-throughput method for quantitatively measuring Drosophila fecundity includes:
[0006] Obtain initial egg-laying images corresponding to multiple pairs of male and female Drosophila to be detected cultured in different types of culture media, wherein different types of the culture media are placed in each pore diameter of a preset porous cell culture plate, and each pair of male and female Drosophila to be detected consists of one female Drosophila and one male Drosophila;
[0007] Perform image preprocessing on the initial spawning image to determine the standard spawning image corresponding to the male and female Drosophila pairs to be detected;
[0008] Perform image recognition on the target standard spawning image to determine the target egg numbers of each of the male and female Drosophila pairs to be detected under different culture media.
[0009] In a feasible implementation manner, the preset multi-well cell culture plate is arranged on a preset multi-well cell culture device, and the pore diameters of the cell culture test tubes of the multi-well cell culture device are adapted to the pore diameters of the respective culture chambers on the preset multi-well cell culture plate.
[0010] In a feasible implementation manner, the image preprocessing includes image grayscale processing and image normalization processing. The performing image preprocessing on the initial spawning image to determine the standard spawning image corresponding to the male and female Drosophila pairs to be detected includes:
[0011] Perform image grayscale processing on the initial spawning image to determine the first candidate spawning image corresponding to the male and female Drosophila pairs to be detected;
[0012] Perform image normalization processing on the first candidate spawning image to determine the standard spawning image corresponding to the male and female Drosophila pairs to be detected.
[0013] In a feasible implementation manner, after performing image preprocessing on the initial spawning image to determine the standard spawning image corresponding to the male and female Drosophila pairs to be detected, the high-throughput method for quantifying Drosophila fecundity further includes:
[0014] Perform image edge detection on the standard spawning image to determine the second candidate spawning image corresponding to the male and female Drosophila pairs to be detected.
[0015] In a feasible implementation manner, the performing image recognition on the target standard spawning image to determine the target egg numbers of each of the male and female Drosophila pairs to be detected under different culture media includes:
[0016] Input the second candidate spawning image into the trained Drosophila egg recognition model for image recognition to determine the initial eggs of each of the male and female Drosophila pairs to be detected under different culture media;
[0017] Screen the initial eggs according to a preset diameter screening rule to determine the target eggs of each of the male and female Drosophila pairs to be detected under different culture media and the target egg numbers of the target eggs.
[0018] In a feasible implementation manner, determine the trained Drosophila egg recognition model by the following method:
[0019] Obtain the oviposition images of male and female Drosophila melanogaster corresponding to multiple samples, and the true number of sample eggs is marked on each of the oviposition images of the samples;
[0020] Input each of the oviposition images of the samples into the initial Drosophila egg recognition model to determine the predicted number of the sample eggs in each of the oviposition images of the samples;
[0021] Train the initial Drosophila egg recognition model based on the predicted number of the sample eggs and the number of the sample eggs to determine the trained Drosophila egg recognition model.
[0022] In the second aspect of the embodiments of the present application, the embodiments of the present application provide a porous cell culture device, and the device includes:
[0023] A base, and a number of first small holes are opened on the base;
[0024] A plurality of cell culture test tubes, the number of the cell culture test tubes is the same as the number of the first small holes, each of the cell culture test tubes is inserted into each of the first small holes, and at least one second small hole is opened at the sealed end of each of the cell culture test tubes;
[0025] Each of the cell culture test tubes is adapted to the aperture of each culture chamber on a preset porous cell culture plate.
[0026] In the third aspect of the embodiments of the present application, the embodiments of the present application provide a high-throughput device for quantitatively measuring Drosophila fecundity, and the high-throughput device for quantitatively measuring Drosophila fecundity includes:
[0027] An acquisition module, configured to acquire the initial oviposition images of multiple pairs of male and female Drosophila to be detected cultured with different types of culture media, wherein different types of the culture media are placed in each aperture of a preset porous cell culture plate, and each pair of male and female Drosophila to be detected consists of a female Drosophila and a male Drosophila;
[0028] A first determination module, configured to perform image preprocessing on the initial oviposition images to determine the standard oviposition images corresponding to the pairs of male and female Drosophila to be detected;
[0029] A second determination module, configured to perform image recognition on the standard oviposition images to determine the target egg numbers of each pair of male and female Drosophila to be detected under different culture media.
[0030] In the fourth aspect of the embodiments of the present application, an electronic device is provided, including: a processor, a memory, and a bus. The memory stores machine-readable instructions executable by the processor. When the electronic device runs, the processor communicates with the memory through the bus. When the machine-readable instructions are run by the processor, the steps of the high-throughput method for quantifying Drosophila fecundity as described above are executed.
[0031] In the fifth aspect of the embodiments of the present application, a computer-readable storage medium is provided. A computer program is stored on the computer-readable storage medium. When the computer program is run by a processor, the steps of the high-throughput method for quantifying Drosophila fecundity as described above are executed.
[0032] Compared with the prior art, the high-throughput method, device, and porous cell culture equipment for quantifying Drosophila fecundity provided by the embodiments of the present application realize diversified and high-throughput analysis of Drosophila egg-laying ability by acquiring initial egg-laying images of multiple pairs of male and female Drosophila to be detected cultured with different types of culture media. Moreover, the present application can perform image preprocessing on the initial egg-laying images, determine the standard egg-laying images corresponding to the pairs of male and female Drosophila to be detected, and then perform image recognition on the target egg-laying images to determine the target egg numbers of each pair of male and female Drosophila to be detected under different culture media. The present application improves the accuracy and consistency of egg data and the efficiency of determining the egg numbers. BRIEF DESCRIPTION OF THE DRAWINGS
[0033] Figure 1 is a flowchart of a high-throughput method for quantifying Drosophila fecundity provided by an embodiment of the present application;
[0034] Figure 2 is a bar chart of the fecundity of multiple strains of Drosophila in a high-throughput method for quantifying Drosophila fecundity provided by an embodiment of the present application;
[0035] Figure 3 is a schematic structural diagram of a porous cell culture equipment provided by an embodiment of the present application;
[0036] Figure 4 is a structural block diagram of a high-throughput device for quantifying Drosophila fecundity provided by an embodiment of the present application;
[0037] Figure 5 is a schematic structural diagram of an electronic device provided by an embodiment of the present application.
[0038] Figures 3 to 5 The corresponding relationship between the reference numerals and the names of the drawings in is as follows:
[0039] 3 Porous cell culture device; 301 Base; 302 Cell culture test tube; 400 High-throughput device for quantifying Drosophila fecundity; 410 Acquisition module; 420 First determination module; 430 Second determination module; 500 Electronic device; 510 Processor; 520 Memory; 530 Bus. Detailed implementation manners
[0040] To better understand the technical solutions provided in the embodiments of this specification, the technical solutions of the embodiments of this specification will be described in detail below through the accompanying drawings and specific embodiments. It should be understood that the specific features in the embodiments of this specification and the embodiments are detailed descriptions of the technical solutions of the embodiments of this specification, rather than limitations on the technical solutions of this specification. Without conflict, the technical features in the embodiments of this specification and the embodiments can be combined with each other.
[0041] In this article, relational terms such as first and second are only used to distinguish one entity or operation from another entity or operation, and do not necessarily require or imply any actual relationship or order between these entities or operations. Moreover, the term "comprising", "including" or any other variant thereof is intended to cover non-exclusive inclusion, so that a process, method, article or device including a series of elements not only includes those elements, but also includes other elements not expressly listed, or also includes elements inherent to such process, method, article or device. Without further limitation, an element defined by the phrase "including a..." does not exclude the existence of additional identical elements in the process, method, article or device including the said element. The term "more than two" includes two or more than two cases.
[0042] First, the applicable application scenarios of this application are introduced. The embodiments provided in this application are applicable to
[0043] Currently, traditional Drosophila egg counting methods rely on experimenters to observe and count each well under a microscope. This process is not only inefficient but also unable to meet the need for continuous egg data collection. The counting results of traditional methods are easily affected by the experience, judgment of observers and environmental factors, and problems such as missed counting and duplicate counting are likely to occur, resulting in poor data accuracy and consistency. Moreover, traditional Drosophila egg counting methods can only determine the egg-laying ability of Drosophila under a single culture medium.
[0044] In the existing method of manually counting the number of Drosophila eggs, Drosophila are kept on standard culture vials or molasses agar plates. After a certain period of time, manual counting is performed using a dissection microscope. However, since the agar medium and yeast paste are soft, Drosophila eggs may sink below the surface, making it difficult to observe, thus underestimating the oviposition rate. To solve this problem, experimenters later began placing Drosophila in multi-well plates instead of culture vials. However, a major drawback of the multi-well plate format is that it is difficult to ensure that Drosophila remain in the same well throughout the experiment, and there are also difficulties in accurately transferring Drosophila when a multi-day process is usually required.
[0045] Based on this, the embodiments of the present application provide a high-throughput method, device, and multi-well cell culture equipment for quantifying Drosophila fecundity. The embodiments provided by the present application solve the technical problems in the prior art, such as inaccurate base numbers, low efficiency, poor consistency in egg calculation data affected by environmental factors, and the ability to determine the oviposition ability of Drosophila only under a single culture medium. The embodiments provided by the present application achieve a diversified analysis of the oviposition ability of Drosophila by setting multiple pairs of male and female Drosophila to be detected in different types of culture media, and improve the accuracy and consistency of egg data and the efficiency of determining the number of eggs through image recognition of oviposition images.
[0046] Please refer to Figure 1 , Figure 1 which is a flowchart of a high-throughput method for quantifying Drosophila fecundity provided by the embodiments of the present application. As Figure 1 shown, the high-throughput method for quantifying Drosophila fecundity includes the following steps:
[0047] S101. Obtain initial oviposition images corresponding to multiple pairs of male and female Drosophila to be detected cultured in different types of culture media. Among them, different types of culture media are placed in each pore of a preset multi-well cell culture plate, and each pair of male and female Drosophila to be detected consists of one female Drosophila and one male Drosophila.
[0048] It should be noted that before determining the initial oviposition images corresponding to multiple pairs of male and female Drosophila to be detected cultured in different types of culture media, the male and female Drosophila to be detected need to be allocated first. In the embodiments of the present application, the male and female Drosophila to be detected are allocated according to pairs of Drosophila, and each pair of male and female Drosophila to be detected consists of one female Drosophila and one male Drosophila, which can ensure the oviposition ability of Drosophila during normal mating.
[0049] In the embodiments provided by the present application, each pair of male and female Drosophila to be detected is placed in a multi-well cell culture device for culturing the Drosophila to be detected. Then, the multi-well cell culture device is sealed by a CO2 generator. At the same time, different types of culture media are added to the respective pore diameters of a preset multi-well cell culture plate, and each cell culture test tube of the multi-well cell culture device is adapted to the pore diameters of the respective culture chambers on the preset multi-well cell culture plate filled with different types of culture media, ensuring seamless docking of the pore diameter with the pore diameter of the culture chamber to prevent the escape of Drosophila. Then, when it is determined that each cell culture test tube of the multi-well cell culture device contains a pair of male and female Drosophila to be detected, an external transfer device is used to flip the multi-well cell culture device to complete the assembly of the device for observing the egg-laying of the male and female Drosophila to be detected, and the male and female Drosophila to be detected in each cell culture test tube can successfully fall onto the surface of the corresponding type of culture media. Then, after the male and female Drosophila to be detected regain their vitality, they are transferred to an external 25°C constant temperature incubator for observation. After the male and female Drosophila to be detected after mating are left standing in the static culture media for 24 hours, the preset multi-well cell culture plate is taken out of the incubator. Then, when it is necessary to immediately collect the initial egg-laying images, the preset multi-well cell culture plate is placed on the platform of a dissection microscope, and a camera is used to accurately focus on each pore diameter and capture the corresponding initial egg-laying images of the male and female Drosophila to be detected. Here, the embodiments provided by the present application adjust the camera focal length as needed to ensure clear imaging and scan all pore diameters.
[0050] Among them, when it is necessary to perform a transfer operation on the male and female Drosophila to be detected, in order to transfer the male and female Drosophila to be detected to a new preset multi-well cell culture plate, the multi-well cell culture device needs to be flipped onto the CO2 generator. After the Drosophila fall onto the mesh lining of the CO2 generator, the original preset multi-well cell culture plate is removed and replaced with a new one, thereby completing the transfer operation on the male and female Drosophila to be detected.
[0051] It can be understood that in the embodiments provided by the present application, when it is not necessary to immediately collect the initial egg-laying images, the preset multi-well cell culture plate can be frozen and stored in an incubator at -20°C according to the actual detection scenario and detection conditions, and taken out for collection when it is necessary to perform imaging or collect the initial egg-laying images corresponding to multiple pairs of male and female Drosophila to be detected.
[0052] Among them, for the porous cell culture device, CO2 generator, and porous cell culture plate provided in the embodiments of the present application, their designs, types, and structures can all be customarily selected and used according to different application scenarios. The CO2 generator in the present application is specifically but not limited to: a CO2 pad; the porous cell culture plate in the present application is specifically but not limited to: a 96-well plate; the porous cell culture device in the present application is specifically but not limited to: a 96-well high-throughput single fly culture device (96-FlyPlate) created based on a preset three-dimensional object creation technology.
[0053] Among them, the preset three-dimensional object creation technology is simply referred to as 3D printing technology. In the present application, a porous cell culture device adapted to a 96-well plate was designed and printed through the 3D printing software of Tinkercad. The embodiments provided in the present application adopt a special geometric chamber design (diameter 6.8 mm × height 15 mm), and triple ventilation holes are configured for the above geometric chamber (the diameter of the ventilation holes can be 1 mm, and the spacing can be 1.2 mm), and the volume of a single ventilation hole is 5.4 mL. The porous cell culture device in the embodiments provided in the present application can ensure air circulation while avoiding cross-well interference, and realizes non-contact Drosophila transfer with the assistance of CO2, with a relatively high success rate (the specific success rate can be: success rate > 95%), and avoids the interference of physical contact on the experiment. This porous cell culture device can handle 96 single Drosophila simultaneously at one time, providing a high-throughput solution for large-scale genetic, nutritional, and pharmacological research.
[0054] Here, each cell culture test tube of the porous cell culture device in the present application corresponds to each pore diameter on the 96-well plate. The diameter and depth of each pore diameter on the 96-well plate have been precisely calculated to ensure that the porous cell culture device can fit with the 96-well plate, and provide reliable support for subsequent automated operations. The present application realizes the customization of the porous cell culture device through 3D printing technology, and the present application can quickly adjust the design according to different experimental requirements, improving the repeatability and efficiency of the experiment.
[0055] Exemplarily, the type of the culture medium in the embodiments provided in the present application can also be customarily set according to different application scenarios and usage conditions. The culture medium in the present application is specifically but not limited to: grape juice culture medium.
[0056] The preparation of the grape juice culture medium in the present application specifically includes the following sub-steps:
[0057] Sub-step 1: Add 200 mL of water, 2 g of agar powder, and 4.8 g of sucrose into a beaker, heat and stir in a microwave oven until completely dissolved, and then boil three times to ensure the uniformity and disinfection effect of the culture medium.
[0058] Sub-step 2: After cooling the above solution to 50 - 60 °C, add 20 mL of grape juice, 5 g of yeast extract, 3 mL of absolute ethanol, and 1.5 mL of glacial acetic acid, and stir well.
[0059] Sub-step 3: Aliquot the mixed grape juice medium into 96-well plates, and after ultraviolet sterilization (for 30 minutes), wait for it to solidify and store it in a 4 °C refrigerator for later use. The maximum storage time is two weeks.
[0060] Among them, the grape juice medium formula in the embodiments provided by this application meets the growth and reproduction requirements of Drosophila melanogaster. By adding appropriate amounts of yeast extract, absolute ethanol, and glacial acetic acid to the medium, the oviposition behavior of Drosophila melanogaster is effectively promoted, and the optimized medium formula improves the fecundity of Drosophila melanogaster, which is suitable for large-scale data screening and experiments. The grape juice medium in the embodiments provided by this application can ensure uniform distribution of nutrients, solve the problem of egg sinking caused by uneven nutrition or soft medium of dead Drosophila melanogaster in traditional media, and significantly improve experimental consistency and throughput.
[0061] In the above, the embodiments provided by this application can replace the 96-well food plates with different formulas according to the actual usage scenario for Drosophila melanogaster reproduction research on transient nutritional intervention.
[0062] S102: Perform image preprocessing on the initial oviposition image to determine the standard oviposition image corresponding to the to-be-detected male and female Drosophila melanogaster pair.
[0063] It should be noted that after obtaining the initial oviposition image in the embodiments provided by this application, image grayscale processing is started on the initial oviposition image to determine the first candidate oviposition image corresponding to the to-be-detected male and female Drosophila melanogaster pair, and then image normalization processing is performed on the first candidate oviposition image to determine the standard oviposition image corresponding to the to-be-detected male and female Drosophila melanogaster pair, and then image edge detection is performed on the standard oviposition image to determine the second candidate oviposition image corresponding to the to-be-detected male and female Drosophila melanogaster pair.
[0064] It can be understood that the image preprocessing of the initial spawning image in the embodiments provided by the present application can be customarily set according to different requirements and usage scenarios. The image preprocessing of the present application can be specific but not limited to: successively performing image grayscale processing (such as image inversion, inversion into an 8-bit grayscale image, etc.) and image normalization processing (such as normalized image pixel enhancement, etc.) on the initial spawning image to determine a clear standard spawning image, and then using a preset Canny edge detection algorithm to perform image edge detection on the standard spawning image, identifying the significant edge graphic features of the corresponding standard spawning image, and minimizing the influence of noise to make its edge positioning clear and accurate, generating a second candidate spawning image corresponding to the to-be-detected male and female Drosophila pairs, and preparing for subsequent image recognition, solving the problems of relying on manual operation and error accumulation in traditional methods, improving the efficiency of egg counting, and being applicable to high-throughput spawning experiments under different experimental conditions.
[0065] S103. Perform image recognition on the target standard spawning image to determine the target egg numbers of each to-be-detected male and female Drosophila pair under different culture media.
[0066] It should be noted that the embodiments provided by the present application determine the second candidate spawning image after edge detection as the target standard spawning image, input the above target standard spawning image into the trained Drosophila egg recognition model for image recognition, determine the initial eggs of each to-be-detected male and female Drosophila pair under different culture media, and then screen the initial eggs according to a preset diameter screening rule to determine the target eggs of each to-be-detected male and female Drosophila pair under different culture media and the target egg numbers of the target eggs.
[0067] It can be understood that the embodiments provided by the present application use a trained Drosophila egg recognition model that performs image recognition using deep learning technology to recognize the Drosophila eggs in the target standard spawning image, and then screen the identified initial eggs according to a preset diameter screening rule to remove an unqualified defect (in the present application, it refers to those shadow features that seemingly are eggs but actually are not), determine the eggs that meet the preset diameter screening as the target eggs under the same culture medium, count the target egg numbers of the target eggs, and also save the segmentation and recognition results of the second candidate spawning image for subsequent analysis and verification.
[0068] Among them, the trained Drosophila egg recognition model in the embodiments provided by the present application can be specific but not limited to a segmentation method that uses flyModel 2 image segmentation for image recognition. More specifically, for example: using a deep learning algorithm (such as a convolutional neural network CNN) to perform egg recognition and egg counting on the target standard spawning image, making the recognition of Drosophila egg numbers in the present application more accurate and high-throughput.
[0069] It can be understood that the embodiments provided in the present application can identify the eggs in the target spawning image according to a preset diameter screening rule and / or other types of preset egg attribute screening rules, such as a preset diameter screening rule, etc., for filtering out those that are in improper positions, too large / small in area, and too large / small in diameter, etc.
[0070] Exemplarily, the present application determines a trained Drosophila egg recognition model through the following sub-steps:
[0071] Sub-step 1: Obtain multiple sample spawning images corresponding to sample male and female Drosophila pairs, and the true number of sample eggs is marked on each sample spawning image.
[0072] It should be noted that before identifying the eggs of the to-be-detected male and female Drosophila pairs, the present application can first obtain the sample spawning images corresponding to the sample male and female Drosophila pairs in the ocean, mark the sample features of the sample eggs in the sample spawning images of each type of sample male and female Drosophila pairs, and determine the true number of the above sample eggs.
[0073] Sub-step 2: Input each sample spawning image into the initial Drosophila egg recognition model to determine the predicted number of sample eggs in each sample spawning image.
[0074] It should be noted that the above collected sample spawning images are input into the initial Drosophila egg recognition model based on a convolutional neural network that can perform deep learning, for generating the predicted number of sample eggs.
[0075] Sub-step 3: Based on the predicted number and the true number of sample eggs, train the initial Drosophila egg recognition model to determine the trained Drosophila egg recognition model.
[0076] It should be noted that according to the preset loss function, the predicted number and the true number of sample eggs, train the initial Drosophila egg recognition model until the loss value of the initial Drosophila egg recognition model converges, and determine the trained Drosophila egg recognition model.
[0077] The embodiments provided in the present application select 12 Drosophila strains to explore the influence of different genotypes on Drosophila fecundity. The experimental design is as follows:
[0078] Experimental object selection: Select 12 to-be-detected male and female Drosophila pairs of different genotypes, covering diverse genetic backgrounds, and set the male-female ratio of each strain to 1:1 to minimize the influence of gender factors on the results in the experiment.
[0079] Experimental conditions: Control the temperature at 25°C ± 1°C to maintain consistent experimental environmental conditions.
[0080] Transfer and mating of male and female Drosophila to be detected: Use a 3D-printed mating device to transfer male and female Drosophila to be detected of each strain to independent wells in a 96-well plate for mating. Place one pair of male and female Drosophila to be detected in each well to ensure no cross-interference. After 24 hours of mating, take out the male and female Drosophila to be detected for egg-laying amount imaging.
[0081] Egg-laying amount imaging and counting: After the mated male and female Drosophila to be detected are left standing in grape juice medium for 24 hours, image each well in the 96-well plate using a microscope and a high-resolution camera, and use an automatic image processing pipeline to segment and count the eggs to obtain the number of eggs in each well. Compare the egg-laying numbers of each Drosophila strain through statistical analysis.
[0082] Data analysis: Compare the egg-laying numbers of female Drosophila to be detected among 12 different strains of male and female Drosophila to be detected, and use statistical methods to analyze the differences in fecundity among different strains, explore how different genotypes affect the fecundity of Drosophila, and then combine the correlation between genetic background and egg-laying number to further speculate on potential gene regulation mechanisms.
[0083] Expected results: By quantitatively comparing the egg-laying abilities of 12 different strains of male and female Drosophila to be detected, it is expected to reveal the significant effects of different genetic backgrounds on the fecundity of Drosophila, providing data support for subsequent research on the association between genes and fecundity.
[0084] Among them, for specific test results, please refer to Figure 2 , Figure 2 which is a bar chart of the fecundity of multiple strains of Drosophila in a high-throughput method for quantitatively measuring the fecundity of Drosophila provided in the embodiments of this application.
[0085] The high-throughput method for quantitatively measuring the fecundity of Drosophila provided in the embodiments of this application, compared with the prior art, through obtaining the initial egg-laying images corresponding to multiple pairs of male and female Drosophila to be detected cultured in different types of culture media, realizes diversified and high-throughput analysis of the egg-laying ability of Drosophila. And this application can perform image preprocessing on the initial egg-laying images, determine the standard egg-laying images corresponding to the pairs of male and female Drosophila to be detected, then perform image recognition on the target egg-laying images to determine the target egg numbers of each pair of male and female Drosophila to be detected under different culture media. This application realizes the automatic recognition of egg numbers, improves the accuracy, consistency and repeatability of egg data, improves the efficiency of determining egg numbers, and can simultaneously process the data of multiple experimental apertures, quickly complete the statistical analysis of large-scale samples, thus meeting the requirements of high-throughput screening experiments. This application can adapt to the diversified experimental needs of more strains of Drosophila under different experimental conditions.
[0086] The high-throughput method for quantifying Drosophila fecundity provided by the embodiments of the present application is not only applicable to the research on Drosophila egg-laying quantification, but also can be widely applied to the fecundity quantification experiments of other insects or model organisms. This method can adapt to the fecundity research of Drosophila on various culture media, with different genetic backgrounds and strains, and has strong versatility and adaptability. It can further explore the relationship between different gene types and fecundity. Moreover, through precise counting and high-throughput screening, the present application can systematically compare the egg-laying behaviors of Drosophila under various treatments, providing strong technical support for the research of genetics, ethology, and physiology.
[0087] Figure 3 It is a schematic structural diagram of a porous cell culture device provided by the embodiments of the present application. As Figure 3 shown, the porous cell culture device 3 includes: a base 301, on which there are a number of first small holes; a plurality of cell culture test tubes 302, the number of cell culture test tubes 302 is the same as the number of first small holes, each cell culture test tube 302 is inserted into each first small hole, and at least one second small hole is opened at the sealed end of each cell culture test tube 302; each cell culture test tube 302 is adapted to the aperture of each culture chamber on the preset porous cell culture plate.
[0088] It should be noted that the number of second small holes at the sealed end of each cell culture test tube 302 in the embodiments provided by the present application can be customarily set according to different application scenarios and the sizes of the male and female Drosophila pairs to be detected. The number of second small holes at the sealed end of each cell culture test tube 302 in the present application is specifically set to 3.
[0089] Figure 4 It is a structural block diagram of a high-throughput device for quantifying Drosophila fecundity provided by the embodiments of the present application. As Figure 4 shown, the high-throughput device 400 for quantifying Drosophila fecundity includes:
[0090] An acquisition module 410, configured to acquire initial egg-laying images corresponding to a plurality of male and female Drosophila pairs to be detected cultured with different types of culture media, wherein different types of culture media are placed in each aperture of a preset porous cell culture plate, and the male and female Drosophila pairs to be detected are composed of one female Drosophila and one male Drosophila.
[0091] A first determination module 420, configured to perform image preprocessing on the initial egg-laying images to determine the standard egg-laying images corresponding to the male and female Drosophila pairs to be detected.
[0092] A second determination module 430, configured to perform image recognition on the standard egg-laying images to determine the target egg numbers of each male and female Drosophila pair to be detected under different culture media.
[0093] Exemplarily, a preset porous cell culture plate is arranged on a preset porous cell culture device, and the cell culture test tubes of the porous cell culture device are adapted to the pore diameters of the respective culture chambers on the preset porous cell culture plate.
[0094] Exemplarily, the image preprocessing includes image grayscale processing and image normalization processing. The first determination module 420 is specifically configured to:
[0095] Perform image grayscale processing on the initial spawning image to determine a first candidate spawning image corresponding to the male and female fruit flies to be detected.
[0096] Perform image normalization processing on the first candidate spawning image to determine a standard spawning image corresponding to the male and female fruit flies to be detected.
[0097] Exemplarily, the first determination module 420 is further specifically configured to:
[0098] Perform image edge detection on the standard spawning image to determine a second candidate spawning image corresponding to the male and female fruit flies to be detected.
[0099] Exemplarily, the second determination module 430 is specifically configured to:
[0100] Input the second candidate spawning image into a trained fruit fly egg recognition model for image recognition to determine the initial eggs of each pair of male and female fruit flies to be detected under different culture media.
[0101] Screen the initial eggs according to a preset diameter screening rule to determine the target eggs of each pair of male and female fruit flies to be detected under different culture media and the target egg quantity of the target eggs.
[0102] Exemplarily, a trained fruit fly egg recognition model is determined by the following method:
[0103] Obtain sample spawning images corresponding to multiple sample pairs of male and female fruit flies to be detected, and the true quantity of sample eggs is marked on each sample spawning image.
[0104] Input each sample spawning image into an initial fruit fly egg recognition model to determine the predicted quantity of sample eggs in each sample spawning image.
[0105] Train the initial fruit fly egg recognition model based on the predicted quantity and the true quantity of the sample eggs to determine the trained fruit fly egg recognition model.
[0106] The high-throughput device 400 for quantifying Drosophila fecundity provided by the embodiments of the present application, compared with the prior art, realizes diversified and high-throughput analysis of Drosophila egg-laying ability by acquiring initial egg-laying images of multiple pairs of male and female Drosophila to be detected cultured with different types of culture media. Moreover, the present application can perform image preprocessing on the initial egg-laying images, determine the standard egg-laying images corresponding to the pairs of male and female Drosophila to be detected, then perform image recognition on the target standard egg-laying images to determine the target egg numbers of each pair of male and female Drosophila to be detected under different culture media. The present application realizes automatic recognition of egg numbers, improves the accuracy, consistency, and repeatability of egg data, enhances the efficiency of determining egg numbers, and can simultaneously process data of multiple experimental apertures, quickly completing statistical analysis of large-scale samples, thereby meeting the requirements of high-throughput screening experiments. The present application can adapt to the diverse experimental needs of more Drosophila strains under different experimental conditions.
[0107] The high-throughput device 400 for quantifying Drosophila fecundity provided by the embodiments of the present application is not only applicable to the quantitative research on Drosophila egg-laying, but also can be widely used in the fecundity quantification experiments of other insects or model organisms. This method can adapt to the fecundity research of Drosophila with various culture media, different genetic backgrounds, and strains, has strong versatility and adaptability, can further explore the relationship between different gene types and fecundity, and through accurate counting and high-throughput screening, the present application can systematically compare the egg-laying behaviors of Drosophila under various treatments, providing strong technical support for the research of genetics, ethology, and physiology.
[0108] Please refer to Figure 5 , Figure 5 which shows a schematic structural diagram of an electronic device provided by the embodiments of the present application. As Figure 5 shown, the electronic device 500 includes a processor 510, a memory 520, and a bus 530.
[0109] The memory 520 stores machine-readable instructions executable by the processor 510. When the electronic device 500 runs, the processor 510 communicates with the memory 520 through the bus 530. When the machine-readable instructions are executed by the processor 510, the steps of the high-throughput method for quantifying Drosophila fecundity in the method embodiments as described above can be executed. The specific implementation manner can refer to the method embodiments and will not be elaborated here. Figures 1 to 2 shown.
[0110] The embodiments of the present application also provide a computer-readable storage medium. A computer program is stored on the computer-readable storage medium. When the computer program is run by a processor, the steps of the high-throughput method for quantifying Drosophila fecundity in the method embodiments as described above can be executed. The specific implementation manner can refer to the method embodiments and will not be elaborated here. Figures 1 to 2 shown.
[0111] Those skilled in the art can clearly understand that for the convenience and conciseness of description, the specific working processes of the systems, devices, and units described above can refer to the corresponding processes in the foregoing method embodiments, and will not be elaborated herein.
[0112] It should be noted that in the above embodiments, the descriptions of the various embodiments have their own emphases. For the parts not described in detail in a certain embodiment, reference can be made to the relevant descriptions of other embodiments.
[0113] Those skilled in the art should understand that the embodiments of the present application can be provided as methods, systems, or computer program products. Therefore, the present application can adopt the form of a complete hardware embodiment, a complete software embodiment, or an embodiment combining software and hardware aspects. Moreover, the present application can adopt the form of a computer program product implemented on one or more computer-readable storage media (including but not limited to disk storage, CD-ROM, optical storage, etc.) containing computer-readable program code.
[0114] The present application is described with reference to the flowcharts and / or block diagrams of methods, apparatuses (systems), and computer program products according to the embodiments of the present application. It should be understood that each process and / or block in the flowchart and / or block diagram can be implemented by computer program instructions, and the combination of processes and / or blocks in the flowchart and / or block diagram can also be implemented by computer program instructions. These computer program instructions can be provided to the processor of a general-purpose computer, a special-purpose computer, an embedded computer, or other programmable data processing devices to generate a machine, so that the instructions executed by the processor of the computer or other programmable data processing devices generate a device for implementing the specified functions in Figure 1 one process or multiple processes and / or blocks Figure 1 one block or multiple blocks.
[0115] These computer program instructions can also be stored in a computer-readable memory that can direct a computer or other programmable data processing device to work in a specific manner, so that the instructions stored in the computer-readable memory generate a manufactured article including an instruction device, and the instruction device implements the specified functions in Figure 1 one process or multiple processes and / or blocks Figure 1 one block or multiple blocks.
[0116] These computer program instructions can also be loaded onto a computer or other programmable data processing device, so that a series of operation steps are executed on the computer or other programmable device to generate a computer-implemented process. Thus, the instructions executed on the computer or other programmable device provide for implementing the specified functions in Figure 1 one process or multiple processes and / or blocks Figure 1Steps of the functions specified in one or more boxes.
[0117] An embodiment of the present application also provides a computer program product, which includes computer software instructions. When the computer software instructions run on a processing device, the processing device is caused to execute the process of a high-throughput method for quantifying Drosophila fecundity.
[0118] The computer program product includes one or more computer instructions. When the computer program instructions are loaded and executed on a computer, the processes or functions according to the embodiments of the present application are wholly or partially generated. The computer may be a general-purpose computer, a special-purpose computer, a computer network, or other programmable devices. The computer instructions may be stored in a computer-readable storage medium, or transmitted from one computer-readable storage medium to another. For example, the computer instructions may be transmitted from a website, a computer, a server, or a data center to another website, a computer, a server, or a data center in a wired manner (such as coaxial cable, optical fiber, digital subscriber line (DSL)) or a wireless manner (such as infrared, wireless, microwave, etc.). The computer-readable storage medium may be any available medium that can be stored by a computer or a data storage device such as a server or a data center that includes one or more integrated available media. The available medium may be a magnetic medium (such as a floppy disk, a hard disk, a magnetic tape), an optical medium (such as a DVD), or a semiconductor medium (such as a solid state disk (SSD)).
[0119] Those skilled in the art can clearly understand that for the convenience and brevity of description, the specific working processes of the systems, devices, and units described above can refer to the corresponding processes in the foregoing method embodiments and will not be elaborated herein.
[0120] In several embodiments provided by the present application, it should be understood that the disclosed devices, apparatuses, and methods can be implemented in other ways. For example, the device embodiments described above are merely illustrative. For example, the division of units is only a logical function division, and there may be other division methods in actual implementation. For example, multiple units or components may be combined or integrated into another system, or some features may be ignored or not executed. Another point is that the displayed or discussed couplings or direct couplings or communication connections to each other may be indirect couplings or communication connections through some interfaces, devices, or units, and may be in electrical, mechanical, or other forms.
[0121] The unit described as a separation component may or may not be physically separated, and the component displayed as a unit may or may not be a physical unit, that is, it may be located in one place or distributed to multiple network units. Some or all of the units can be selected according to actual needs to achieve the purpose of the solution of this embodiment.
[0122] In addition, each functional unit in various embodiments of the present application can be integrated in a processing unit, or each unit can exist physically alone, or two or more units can be integrated in one unit. The above-mentioned integrated unit can be implemented in the form of hardware or in the form of a software functional unit.
[0123] If the integrated unit 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 this understanding, the technical solution of the present application, in essence, or the part that contributes to the prior art, or all or part of this technical solution, can be embodied in the form of a software product. This computer software product is stored in a storage medium and includes several instructions to enable a computer device (which can be a personal computer, a server, or a network device, etc.) to execute all or part of the steps of the methods in various embodiments of the present application. The aforementioned storage medium includes: various media such as USB flash drives, mobile hard disks, read-only memory (ROM), random access memory (RAM), magnetic disks, or optical discs that can store program codes.
[0124] The above embodiments are only used to illustrate the technical solutions of the present application, rather than to limit them; although the present application 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 make the essence of the corresponding technical solutions deviate from the spirit and scope of the technical solutions of various embodiments of the present application.
[0125] Although the preferred embodiments of this specification have been described, those skilled in the art can make additional changes and modifications once they know the basic creative concepts. Therefore, the appended claims are intended to be construed as including the preferred embodiments and all changes and modifications that fall within the scope of this specification.
[0126] Obviously, those skilled in the art can make various changes and modifications to this specification without departing from the spirit and scope of this specification. Thus, if these modifications and variations of this specification fall within the scope of the claims of this specification and their equivalent technologies, this specification is also intended to include these changes and modifications.
Claims
1. A high-throughput method for quantifying Drosophila fecundity, characterized in that, The high-throughput method for quantifying Drosophila fecundity includes: Obtaining initial spawning images corresponding to multiple pairs of male and female Drosophila to be detected cultured in different types of culture media, wherein different types of the culture media are placed in respective pores of a preset multi-well cell culture plate, and each pair of male and female Drosophila to be detected consists of one female Drosophila and one male Drosophila; Performing image preprocessing on the initial spawning images to determine standard spawning images corresponding to the pairs of male and female Drosophila to be detected; Performing image recognition on the standard spawning images to determine the target egg numbers of each pair of male and female Drosophila to be detected under different culture media.
2. The high-throughput method for quantifying Drosophila fecundity according to claim 1, wherein The preset multi-well cell culture plate is arranged on a preset multi-well cell culture device, and each cell culture test tube of the multi-well cell culture device is adapted to the pore diameter of each culture chamber on the preset multi-well cell culture plate.
3. The high-throughput method for quantifying Drosophila fecundity according to claim 1, wherein The image preprocessing includes image grayscale processing and image normalization processing. The performing image preprocessing on the initial spawning images to determine standard spawning images corresponding to the pairs of male and female Drosophila to be detected includes: Performing image grayscale processing on the initial spawning images to determine first candidate spawning images corresponding to the pairs of male and female Drosophila to be detected; Performing image normalization processing on the first candidate spawning images to determine standard spawning images corresponding to the pairs of male and female Drosophila to be detected.
4. The high-throughput method for quantifying Drosophila fecundity according to claim 1, wherein After performing image preprocessing on the initial spawning images to determine standard spawning images corresponding to the pairs of male and female Drosophila to be detected, the high-throughput method for quantifying Drosophila fecundity further includes: Performing image edge detection on the standard spawning images to determine second candidate spawning images corresponding to the pairs of male and female Drosophila to be detected.
5. The high-throughput method for quantifying Drosophila fecundity according to claim 4, wherein The performing image recognition on the standard spawning images to determine the target egg numbers of each pair of male and female Drosophila to be detected under different culture media includes: Inputting the second candidate spawning images into a trained Drosophila egg recognition model for image recognition to determine the initial eggs of each pair of male and female Drosophila to be detected under different culture media; Screening the initial eggs according to a preset diameter screening rule to determine the target eggs of each pair of male and female Drosophila to be detected under different culture media and the target egg numbers of the target eggs.
6. The high-throughput method for quantifying Drosophila fecundity according to claim 5, wherein Determining the trained Drosophila egg recognition model in the following manner: Obtaining sample spawning images corresponding to multiple pairs of male and female Drosophila for sample detection, with the true numbers of sample eggs marked on each sample spawning image; Inputting each sample spawning image into an initial Drosophila egg recognition model to determine the predicted numbers of the sample eggs in each sample spawning image; Training the initial Drosophila egg recognition model based on the predicted numbers and the true numbers of the sample eggs to determine the trained Drosophila egg recognition model.
7. A porous cell culture device is applied to the high-throughput method for quantitatively measuring Drosophila fecundity according to any one of claims 1-6, characterized in that, The device includes: A base, on which there are a number of first small holes; A plurality of cell culture test tubes, the number of the cell culture test tubes being the same as the number of the first small holes, each of the cell culture test tubes being inserted into each of the first small holes, and at least one second small hole being formed in the sealed end of each of the cell culture test tubes; Each of the cell culture test tubes is adapted to the aperture of each culture chamber on a preset multi-well cell culture plate.
8. A high-throughput device for quantitatively measuring Drosophila fecundity, characterized in that, The high-throughput device for quantifying Drosophila fecundity includes: An acquisition module, configured to acquire initial spawning images corresponding to a plurality of pairs of male and female Drosophila to be detected cultured in different types of culture media, wherein different types of the culture media are placed in each aperture of a preset multi-well cell culture plate, and each pair of male and female Drosophila to be detected consists of one female Drosophila and one male Drosophila; A first determination module, configured to perform image preprocessing on the initial spawning images to determine standard spawning images corresponding to the pairs of male and female Drosophila to be detected; A second determination module, configured to perform image recognition on the standard spawning images to determine the target egg numbers of each pair of male and female Drosophila to be detected under different culture media.
9. An electronic device, characterized in that, Including: A processor, a memory and a bus, the memory storing machine-readable instructions executable by the processor. When the electronic device runs, the processor communicates with the memory through the bus. When the machine-readable instructions are run by the processor, the steps of the high-throughput method for quantifying Drosophila fecundity as described in any one of claims 1-7 above are executed.
10. A computer-readable storage medium, characterized in that, A computer program is stored on the computer-readable storage medium. When the computer program is run by the processor, the steps of the high-throughput method for quantifying Drosophila fecundity as described in any one of claims 1-7 above are executed.