High-throughput plasmid construction functional island

Abstract

The utility model discloses a high-throughput plasmid construction functional island, and relates to the technical field of plasmid library construction. Comprising an automatic integration table top, a box body is installed on the top face of the automatic integration table top, a carrying assembly is arranged in the box body, an automatic working assembly is arranged in the circumferential direction of the carrying assembly, and the carrying assembly and the automatic working assembly are both installed on the top face of the automatic integration table top; the carrying assembly comprises a transfer table installed in the center of the automatic integration table top, a first sliding rail robot and a second sliding rail robot are arranged on the two sides of the transfer table correspondingly, and the first sliding rail robot and the second sliding rail robot are used for taking and placing needed objects from the automatic working assembly and the transfer table. According to the utility model, the first slide rail robot and the second slide rail robot are used for respectively taking and placing required articles from the automatic working assembly and the transfer table, so that the one-stop mechanical operation of the whole process of plasmid library construction and extraction can be effectively realized.

Description

Technical Field

[0001] This utility model relates to the field of plasmid library construction technology, and in particular to a high-throughput plasmid construction functional island. Background Technology

[0002] In plant gene research and synthetic biology, plasmid library construction and extraction are core steps in experiments such as gene function verification and high-throughput screening. Traditional experimental procedures heavily rely on manual operation and have the following main drawbacks:

[0003] (1) Low throughput and high time cost: Manual operation requires completing multiple steps such as vector enzyme digestion, Oligo annealing, recombination reaction, heat shock transformation, colony picking, shaking culture and plasmid extraction in each well. A single experiment can only process 1-96 samples, which is difficult to meet the needs of modern scientific research for large-scale (e.g., 500 samples / day) plasmid library screening. In addition, each step requires manual transfer of samples, and a single process takes more than 24 hours.

[0004] (2) Poor quality stability: Manual pipetting and plate transfer operations are prone to introducing errors (such as pipetting volume deviation and cross-contamination), resulting in unreliable plasmid construction success rate (insufficient number of single clones), accuracy (fragment insertion errors), and extraction quality (low yield and endotoxin residue). Actual tests show that the success rate of mixed plasmid library construction under manual operation is less than 60%, and the accuracy is less than 80%; the plasmid yield of a single sample is often less than 3 μg, and the endotoxin content is likely to exceed 100 EU / μg, which seriously affects the efficiency of subsequent gene function verification and screening.

[0005] (3) Insufficient level of automation: Existing technologies mostly use decentralized single-machine equipment (such as liquid workstations, PCR instruments, and nucleic acid extractors). There is a lack of collaborative control between the equipment, and manual intervention is required for sample transfer (such as manually transferring from the PCR instrument to the coating instrument). It is impossible to achieve the closed-loop automation of the entire process of "vector construction-coating-screening-extraction", and it is difficult to support 24-hour uninterrupted experiments.

[0006] The aforementioned problems directly result in a limited range of plasmid library screening (covering only a small number of samples) and low efficiency in target gene screening, failing to meet the needs of the synthetic biology field for high-throughput, high-precision, and standardized experiments.

[0007] Therefore, there is an urgent need for a high-throughput plasmid construction functional island that can effectively realize one-stop mechanized operation of the entire process of plasmid library construction and extraction. Utility Model Content

[0008] The purpose of this invention is to provide a high-throughput plasmid for constructing functional islands, thereby solving the problems existing in the prior art.

[0009] To achieve the above objectives, this utility model provides the following solution: This utility model provides a high-throughput plasmid construction functional island, including an automated integration platform. A box is installed on the top surface of the automated integration platform, and a transport component is disposed inside the box. An automated working component is disposed around the transport component. Both the transport component and the automated working component are mounted on the top surface of the automated integration platform. The transport component includes a transfer platform installed at the center of the automated integration platform. A first slide rail robot and a second slide rail robot are respectively disposed on both sides of the transfer platform. The first slide rail robot and the second slide rail robot are used to pick up and place required items from the automated working component and the transfer platform.

[0010] Preferably, the automated working components include an automated liquid workstation for plasmid library construction, an automated consumables stack, an automated coating instrument, an automated bacterial scraper, an automated liquid dispenser, an automated shaking incubator, an automated microbial colony selector, an automated incubator, a mobile robot interaction station, an automated high-throughput centrifuge, an automated membrane peeler, a fully automated plasmid extractor, and a refrigerator, all installed on the top surface of the automated integrated platform and arranged sequentially around the transport components.

[0011] Preferably, multiple automated consumable stacks are provided, and a first support is provided above the multiple automated consumable stacks. The first support is fixedly connected to the automated integration platform. An electroporator, an automated PCR instrument, and a first automated adhesive sealing instrument are sequentially installed on the top surface of the first support. The electroporator is located near the automated liquid workstation for plasmid library construction, and the first automated adhesive sealing instrument is located near the automated coating instrument.

[0012] Preferably, a second support is provided above the automated liquid dispenser, the second support is fixedly connected to the automated integrated platform, and a second automated adhesive sealing device is installed on the top surface of the second support.

[0013] Preferably, multiple automated shaking incubators are provided.

[0014] Preferably, a third support is provided above the interaction position of the mobile robot, the third support is fixedly connected to the automated integration platform, and an automated multifunctional enzyme-linked immunosorbent assay (ELISA) reader is installed on the top surface of the third support.

[0015] Preferably, the automated liquid workstation for plasmid library construction, the electroporator, the automated PCR instrument, the first automated sealing and sealing instrument, the automated high-throughput centrifuge, the automated membrane tearing machine, the fully automated plasmid extractor, and the refrigerator are arranged around the first sliding rail robot.

[0016] Preferably, the automated coating device, the automated bacterial scraper, the automated liquid dispenser, the automated shaking incubator, the automated microbial colony selector, and the automated incubator are arranged around the second sliding rail robot.

[0017] Preferably, the mobile robot interaction position is arranged opposite to the transfer platform.

[0018] Preferably, the box is equipped with multiple doors, and a fully automated mobile robot is installed on the outside of the box.

[0019] The present invention discloses the following technical effects:

[0020] This invention enables the placement of automated working components around the transport components, allowing the first and second slide rail robots to retrieve and place required items from the automated working components and the transfer platform, respectively. This allows the invention to effectively achieve one-stop mechanized operation for the entire process of plasmid library construction and extraction. Attached Figure Description

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

[0022] Figure 1 This is a schematic diagram of the overall structure of this utility model;

[0023] Figure 2 This is a schematic diagram of the internal structure of the present invention viewed from the left.

[0024] Figure 3 This is a schematic diagram of the internal structure of the present invention from the right side.

[0025] Figure 4 This is a flowchart of the plasmid library construction process for this utility model;

[0026] The components include: 1. Automated liquid workstation for plasmid library construction; 2. Electroporator; 3. Automated PCR instrument; 4. First automated sealing and sealing instrument; 5. Automated film peeling machine; 6. Automated coating instrument; 7. Automated shaking incubator; 8. Automated incubator; 9. Automated bacterial scraper; 10. Automated microbial colony selection instrument; 11. Fully automated plasmid extractor; 12. Automated high-throughput centrifuge; 13. Automated liquid dispenser; 14. Mobile robot interaction station; 15. Automated multifunctional microplate reader; 16. Refrigerator; 17. First sliding rail robot; 18. Fully automated mobile robot; 19. Automated consumables stack; 20. High-efficiency filtration system; 21. Fully tri-color light buzzer module; 22. Automated integrated worktable; 23. Second sliding rail robot; 24. Cabinet. Detailed Implementation

[0027] The technical solutions of the present utility model will be clearly and completely described below with reference to the accompanying drawings of the embodiments. Obviously, the described embodiments are only some embodiments of the present utility model, and not all embodiments. Based on the embodiments of the present utility model, all other embodiments obtained by those of ordinary skill in the art without creative effort are within the protection scope of the present utility model.

[0028] To make the above-mentioned objectives, features and advantages of this utility model more apparent and understandable, the utility model will be further described in detail below with reference to the accompanying drawings and specific embodiments.

[0029] Reference Figures 1 to 4 This utility model discloses a high-throughput plasmid construction functional island, including an automated integration platform 22. A box 24 is installed on the top surface of the automated integration platform 22. A transport component is arranged inside the box 24. An automated working component is arranged around the transport component. Both the transport component and the automated working component are installed on the top surface of the automated integration platform 22. The transport component includes a transfer platform installed at the center of the automated integration platform 22. A first slide rail robot 17 and a second slide rail robot 23 are respectively arranged on both sides of the transfer platform. The first slide rail robot 17 and the second slide rail robot 23 are used to pick up and put away the required items from the automated working component and the transfer platform.

[0030] The automated integrated worktable 22 is constructed from a single piece of 304 stainless steel, offering corrosion resistance and easy cleaning. The bottom of the worktable 22 features a concealed wiring channel with separate channels for power and signal lines, preventing signal interference and keeping wiring concealed. It is equipped with both pull-out (50kg load capacity) and rotating (30kg load capacity) auxiliary platforms for easy consumable replenishment and equipment maintenance. It also integrates an LED lighting module (300 lux brightness, meeting operational lighting needs) and a 254nm UV lamp module, covering 90% of the worktable surface. The disinfection cycle can be set via a central control system to initiate automated disinfection (30 minutes per disinfection cycle), ensuring a clean experimental environment.

[0031] The first and second sliding rail robots 17 and 23 adopt a 6-axis articulated design, which has 360-degree full-space grasping capability. They achieve a positioning accuracy of ±0.05mm through servo motors and ball screw transmission mechanisms. The docking points with each device are equipped with photoelectric sensor switches to ensure the accuracy of sample board grasping and placement. They are mainly responsible for the automated transmission and transfer of consumables and sample boards between devices. The action commands are automatically issued by the central control system according to the experimental process.

[0032] This invention enables the first slide rail robot 17 and the second slide rail robot 23 to pick up and place the required items from the automated working components and the transfer platform, respectively, by setting the automated working components around the transport components; thus, this invention can effectively realize one-stop mechanized operation of the entire process of plasmid library construction and extraction.

[0033] Further optimization of the scheme includes an automated work component consisting of an automated liquid workstation 1 for constructing a plasmid library, an automated consumables stack 19, an automated coating instrument 6, an automated scalpel 9, an automated liquid dispenser 13, an automated shaking incubator 7, an automated microbial colony selector 10, an automated incubator 8, a mobile robot interaction station 14, an automated high-throughput centrifuge 12, an automated membrane peeler 5, a fully automated plasmid extractor 11, and a refrigerator 16.

[0034] The automated liquid workstation for plasmid library construction integrates 45 platform positions (including standard plate positions, reagent tank cooling racks, and deep-well plate reagent cooling racks), equipped with one 96-channel robotic arm, one flexible 8-channel robotic arm, two 360-degree rotating grippers, two cameras, one oscillation module, and eight temperature-controlled incubation modules. Each module is connected in series with the integrated circuit system (power line + signal line) through internal piping (liquid supply / drainage). The robotic arm, combined with a multi-axis transmission mechanism and a grating positioning system, achieves high-precision spatial positioning at the ±0.1mm level. It can independently complete the entire process of carrier construction, coating and liquid addition, and low-temperature temporary storage of samples. The operation instructions are uniformly issued by the central control system.

[0035] The automated film-peeling machine 5 can be used as a standalone system or integrated into automated and robotic workflows via a central control system.

[0036] The automated incubator 8 interacts with other devices via a second sliding rail robot 23. The temperature control range is no narrower than 0℃-70℃, with temperature uniformity ≤±0.3℃ and temperature fluctuation ≤±0.1℃. The door opening position is customizable for more flexible integration, and the plate retrieval speed does not exceed 15 seconds. It can hold 80 microplates, and the chamber material is made of 304 antibacterial stainless steel. It employs ring heating and ring cooling technology. It is equipped with two doors, one for automated integration and one for manual operation.

[0037] Two fully automated plasmid extractors 11 are set up and placed vertically via a fourth support. Each fully automated plasmid extractor 11 is equipped with one 96DW deep-well plate magnetic head, one 96DW hot block, and one 96KF hot block. Based on the principle of magnetic bead extraction, the precise magnetic attraction and release action of the magnetic head is linked with the room temperature-95℃ temperature control of the hot block to achieve fully automated extraction and purification of DNA and RNA. The extracted product plate is directly transferred to the refrigerator 16 by the first sliding rail robot 17 for storage and later use, without the need for intermediate intervention.

[0038] To further optimize the solution, multiple automated consumable stacks 19 are provided. A first support is provided above the multiple automated consumable stacks 19. The first support is fixedly connected to the automated integration platform 22. An electroporator 2, an automated PCR instrument 3, and a first automated adhesive sealing instrument 4 are installed sequentially on the top surface of the first support. The electroporator 2 is located near the automated liquid workstation 1 for plasmid library construction, and the first automated adhesive sealing instrument 4 is located near the automated coating instrument 6.

[0039] The electroporator 2 is composed of an electroporation instrument and an automated sample loading device, and is installed above the automated consumable stack 19 through the first bracket. The electroporator 2 can efficiently introduce the vectors for plasmid library construction into microorganisms, and is equipped with an arc suppression system, which can reduce arcs, ensure the safety of precious samples, and can transform bacteria, yeasts and other microorganisms safely and repeatedly; it can effectively make the transformation efficiency higher than the chemical method.

[0040] The automated PCR instrument 3 consists of a 96-channel PCR instrument and a 384-channel PCR instrument, and is installed side by side above the automated consumable stack 19 through the first bracket, and is connected to the central control system through an Ethernet interface to form a linkage control with the well plate transfer system. It supports the automatic switching and loading of 96-well plates and 384-well plates, can be flexibly scheduled according to the experimental throughput requirements, and meets the nucleic acid amplification requirements of different scales; the sample plate after amplification is automatically transported back to the plasmid library construction automated liquid workstation 1 by the first slide rail robot 17 to perform the next library construction operation.

[0041] In a further optimized solution, a second bracket is provided above the automated dispenser 13. The second bracket is fixedly connected to the automated integration table 22, and a second automated sticky film sealer is installed on the top surface of the second bracket.

[0042] The first automated sticky film sealer 4 and the second automated sticky film sealer adopt a room temperature pressure-sensitive full-automatic film sealer. The room temperature pressure-sensitive full-automatic film sealer can achieve the automatic film sealing of the sticky sealing film without heating, and is suitable for the room temperature reaction plate in vector construction. It seals the film at room temperature without generating any heat and has no adverse effect on biological samples. It occupies a small space, automatically recognizes the plate height: it can automatically recognize the loaded microplate and perform reliable film sealing on it; it can be perfectly integrated into the plasmid library construction system and can automatically cut the film in a parallel manner from left to right. It can be used independently or integrated into the automated experimental process in the central control system.

[0043] The automated dispenser 13 can achieve the automated liquid medium dispensing operation for the liquid culture of 96-well plates after picking bacteria. This instrument uses a detachable and high-temperature sterilizable dispensing box. The dispensing box contains 8 single tubes. Each reagent can be equipped with a separate dispensing box to avoid reagent mixing. As an alternative, if the dispensing box is cleaned between each use, the same dispensing box can be used for multiple reagents.

[0044] In a further optimized solution, multiple automated shaking incubators 7 are provided.

[0045] The automated shaking incubator 7 features a plate shuttle transfer system controlled by a built-in stepper motor, equipped with plate detection functionality. Integrated with automated system software, it offers more than five door selection positions, with an average plate retrieval speed of no more than 25 seconds. The temperature control device is equipped with an external thermostat, with a temperature control range of 4-50℃. It can automatically perform moist heat sterilization, achieving unattended operation and providing effective sterilization even for difficult-to-treat heat-resistant microorganisms. After the sterilization process, it automatically returns to the preset incubation temperature. The shaking rack uses a dual-magnetic-driven shaking system, with the bottom and top of the rack vibrating simultaneously to ensure consistent shaking for different samples. The two racks can be independently set with different shaking speeds; the amplitude is 2mm, and the circumferential shaking speed is 100-1200rpm.

[0046] To further optimize the design, a third support is installed above the mobile robot interaction position 14. The third support is fixedly connected to the automated integration platform 22, and an automated multifunctional microplate reader 15 is installed on the top surface of the third support.

[0047] The Automated Multifunctional Microplate Reader 15 communicates in real-time with the quality control module of the central control system via an RS232 data bus. The Automated Multifunctional Microplate Reader 15 integrates one set each of host filter module and grating module, and is equipped with a 96-channel synchronous detection module. It can automatically detect the A260, A280, and A230 UV absorbance of nucleic acid samples. The detection data is uploaded to the central control system in real time, and the system automatically generates nucleic acid concentration and purity analysis reports, which serve as the core data basis for plasmid library quality control.

[0048] To further optimize the scheme, an automated liquid workstation 1 for plasmid library construction, an electroporator 2, an automated PCR instrument 3, a first automated sealing and sealing instrument 4, an automated high-throughput centrifuge 12, an automated membrane tearing machine 5, a fully automated plasmid extractor 11, and a refrigerator 16 are set around a first sliding rail robot 17.

[0049] Refrigerator 16 interacts with other devices via the first sliding rail robot 17. Refrigerator 16 is internally equipped with three layered automated racks, each with a built-in photoelectric positioning sensor, capable of stably storing 24 standard sample plates at a time. Rack movement and sample retrieval are controlled by a central control system, enabling automated sample plate storage and real-time inventory management, meeting the short-term refrigeration needs of the library samples. An external low-temperature circulating water bath provides a temperature control range of no less than 4℃-25℃. It features two doors, one for automated integration and one for manual operation, facilitating more flexible integration. The plate retrieval speed does not exceed 15 seconds.

[0050] The scheme was further optimized by setting up an automated coating device 6, an automated bacterial scraper 9, an automated liquid dispenser 13, an automated shaking incubator 7, an automated microbial colony selector 10, and an automated incubator 8 around the second sliding rail robot 23.

[0051] The Automated Coating System 6 ensures that, in a clean environment, the covers of standard well plates (8-well plates and single-well plates) can be opened and closed, and that all bacterial suspensions in the wells can be coated to ensure uniform distribution on the agar surface. The Automated Coating System 6 supports standalone use and integration into automated processes.

[0052] The automated scraper 9 can achieve single-septum solid culture medium coating.

[0053] The Automated Microbial Colony Picker 10 can be used to screen and pick microbial clones, picking 1500 colonies per hour. Integrated into a central control system, it is simple to use and can be quickly started and run. It provides comprehensive data tracking from start to finish.

[0054] The automated high-throughput centrifuge 12 can be used for bacterial centrifugation during plasmid extraction, protein and nucleic acid separation, and other operations. It has a maximum speed of 6200 RPM and a maximum centrifugal force (KCF) of 6446. The top cover is used for loading and unloading centrifuged samples. The window opening time is less than 6 seconds. It can be integrated into automated experimental workflows or operated as a standalone unit.

[0055] The design was further optimized so that the mobile robot interaction station 14 was positioned opposite the transfer platform.

[0056] The design was further optimized by installing multiple doors on the housing 24 and setting up a fully automated mobile robot 18 on the outside of the housing 24.

[0057] The fully automated mobile robot 18 operates in a dedicated AGV channel on the outer side of the island, equipped with a 6-axis articulated robotic arm and an intelligent electric gripper. The intelligent electric gripper has a built-in pressure sensor and can adapt to the size of different consumables, from 24-hole plates to 384-hole plates, to achieve rapid gripping and stable fitting. The fully automated mobile robot 18 integrates a dual lidar sensing system, which can monitor environmental obstacles in real time within a 360° range. When an obstacle is encountered, it immediately triggers an audible and visual alarm and pauses its operation. Operation resumes after the obstacle is cleared, ensuring the safety and reliability of cross-island transportation.

[0058] The automated consumables stack 19 adopts a "partitioned storage" mode. The automated consumables stack 19 integrates dedicated storage units for various types of consumables such as deep-hole plates, shallow-hole plates, and pipette tips. It is equipped with an autonomous scanner that can automatically identify the consumable model and inventory quantity. It achieves data interconnection with the central control system through an IoT communication module. Combined with the collaborative operation of the fully automated mobile robot 18 and the first slide rail robot 17, it realizes automated identification, inventory warning, accurate retrieval and replenishment of consumables.

[0059] The interior of the housing 24 is the island, and the exterior is the island. The mobile robot interaction station 14 serves as the connecting node for "island-to-island" transfer, enabling precise docking with the fully automated mobile robot 18. The fully automated mobile robot 18, the first slide rail robot 17, and the second slide rail robot 23 are connected via a standardized industrial Ethernet interface. A built-in signal conversion module ensures cross-device communication protocol compatibility, constructing a consumables transfer network of "island-intra-island slide rail transfer - interaction station handover - AGV cross-island transfer." This completes the consumables transfer and replenishment operations between this island and other automated islands, ensuring the continuity of multi-island collaboration.

[0060] Multiple high-efficiency filtration systems 20 are installed on the top of the enclosure 24, covering the entire space above the carrier construction area and coating culture area. Four sets of HEPA H14 grade high-efficiency air filters are configured. These high-efficiency filtration systems 20 are connected to the island's environmental control system, providing an airflow of 1000 m³ / h. This maintains a slightly positive pressure environment of 0.5 kPa within the island, ensuring air cleanliness meets the Class 100 standard and effectively preventing the intrusion of dust and microorganisms from the external environment, thus guaranteeing the cleanliness of the experimental environment.

[0061] Multiple tri-color LED buzzer modules 21 are installed around the top surface of the enclosure 24. These modules are arranged in a ring around the top edge of the enclosure 24 and are linked to the alarm unit of the central control system via signal cables.

[0062] The tri-color LED buzzer module 21 is equipped with blue, green, and red indicator lights and a buzzer component. The status display of the tri-color LED buzzer module 21 accurately corresponds to the experimental process: it displays blue (brightness 500cd / m²) when the experiment is stationary or paused, green when the experiment is running normally, and red when an error occurs (equipment failure, insufficient consumables, or abnormal samples), and simultaneously triggers an 85dB buzzer alarm, intuitively indicating the equipment operating status and guiding the experimenters to intervene and handle it in a timely manner.

[0063] The following sub-processes are closed-looped by scheduling equipment to operate collaboratively through a central control console:

[0064] (1) Construction and extraction of mixed plasmid libraries: The system automatically completes the dissolution of oligo fragments, configuration of annealing system, recombination reaction, electroconversion, shaking recovery, whole plate coating, static culture, cloning scraping, centrifugation and bacterial collection and plasmid extraction, and can stably complete the construction and extraction of ≥500 libraries within 24 hours.

[0065] (2) One-to-one plasmid library construction and extraction: Automated single clone selection, shaking culture, centrifugation and plasmid extraction, completing ≥500 library constructions and ≥2000 single clone selections and extractions within 24 hours.

[0066] Quality and efficiency assurance mechanisms:

[0067] (1) Contamination control: Efficient filtration system 20 (HEPA-03A) is configured, combined with disposable consumables and UV sterilization module to ensure positive pressure sterility of the operating environment and contamination rate close to 0; (2) Data traceability: The entire process records equipment operating parameters, sample flow path and experimental results (such as cloning coordinates, plasmid yield), supports export of electronic experimental records (ELN), and meets GLP specifications; (3) Precise control: The automated liquid workstation 1 for plasmid library construction adopts a high-precision pipetting module (error <1%), and the automated microbial colony picker 10 achieves a true monoclonal identification accuracy of >98% through high-resolution imaging (10μm / pixel), ensuring experimental repeatability and result reliability.

[0068] Technical solution content:

[0069] (1) Plasmid construction and plating culture:

[0070] ① Annealing: Anneal buffer and Dilution Buffer are manually placed in the automated liquid workstation 1 for plasmid library construction. The first sliding rail robot 17 takes two 96PCR plates, A and B, containing upstream and downstream primers respectively, from the refrigerator 16, and then takes an empty 96PCR plate C from the automated consumables stack. After adding Anneal buffer to the empty PCR plate, the automated liquid workstation 1 adds upstream and downstream primers to PCR plate C. The first sliding rail robot 17 transfers it to the PCR instrument for annealing. At the same time, the first sliding rail robot 17 throws PCR plates A and B into the trash can, and then takes in two 96PCR plates, D and E, from the automated consumables stack 19. After annealing, the first sliding rail robot 17 transfers PCR plate C to the automated liquid workstation 1 for plasmid library construction; Dilution Buffer is added to PCR plate D, and then some sample from PCR plate C is added.

[0071] ② Ligation: The ligation reagent is manually placed in the automated liquid workstation 1 for plasmid library construction. The automated liquid workstation 1 for plasmid library construction automatically prepares the ligation system and adds it to PCRE. Then, a portion of the sample from PCR plate D is added to PCR plate E. The first sliding rail robot 17 transfers PCR plate E to the automated PCR instrument 3 for ligation. At the same time, the first sliding rail robot 17 takes a new 96 PCR plate F from the automated consumable stack 19 and places it in the low-temperature zone of the automated liquid workstation 1 for plasmid library construction.

[0072] ③ Transformation: Competent cells are manually placed into the automated liquid workstation 1 for plasmid library construction. The automated liquid workstation 1 adds the competent cells to PCR plate F. Simultaneously, the first sliding-rail robot 17 transfers PCR plate E from the automated PCR instrument 3 to the automated liquid workstation 1, and adds the sample from PCR plate E to the competent cells in PCR plate F. After pre-cooling for 20 minutes, the first sliding-rail robot 17 discards PCR plate E into the trash can and retrieves a 96-well plate from the automated consumables stack 19, adding LB medium to the automated liquid workstation 1. After pre-cooling, PCR plate F is transferred to the automated PCR instrument 3 for heat shock. The automated liquid workstation 1 then adds all the sample from PCR plate F to a 96-well plate. The first sliding-rail robot 17 transfers the 96-well plate to the transport platform, and the second sliding-rail robot 23 seals the 96-well plate from the transport platform and places it in the automated shaking incubator 7 for 1 hour of incubation.

[0073] ④ Coating: After the 96-well deep plate is cultured in the automated shaking incubator 7, the first sliding rail robot 17 transfers it to the automated liquid workstation 1 for plasmid library construction, while simultaneously retrieving an 8-septum culture dish from the automated consumables stack 19. In the automated liquid workstation 1, the bacterial culture from the 96-well deep plate is added to the culture dish, and the coating head is simultaneously transferred from the automated consumables stack 19 to the automated coating instrument 6. After the liquid addition is complete, the second sliding rail robot 23 places it into the automated incubator 8. This process is repeated until all samples are coated and then placed in the automated incubator 8 for overnight cultivation.

[0074] (2) Single clone selection and culture:

[0075] The second sliding rail robot 23 transfers the culture dish from the automated incubator 8 to the automated microbial colony selector 10, which automatically takes pictures, scans, and identifies single clones. At the same time, with the cooperation of the first sliding rail robot 17 and the second sliding rail robot 23, the culture dish is transported from the automated consumables stack 19 to the automated dispenser 13 to add liquid culture medium, and then transferred to the automated microbial colony selector 10. The automated microbial colony selector 10 automatically selects single clones and places them into the culture plate, which is then placed in the automated shaking incubator 7 for overnight incubation.

[0076] (3) Plasmid extraction:

[0077] After the plasmid extraction reagent is manually placed into the corresponding position in the automated liquid workstation 1 for plasmid library construction, the 48-well plate, which has been cultured overnight in the automated shaking incubator 7, is centrifuged in the automated high-throughput centrifuge 12 and then transferred to the automated liquid workstation 1 for plasmid library construction to remove the supernatant. Plasmid extraction reagent is added, and the plate is shaken, centrifuged, and the precipitate is removed. The supernatant is then transferred to a 96U plate. PW1 and PW2 reagents are then added to the 96U plate, and the plate is shaken and centrifuged. At the same time, the 96V plate is transferred to the automated liquid workstation 1 for plasmid library construction. After centrifugation, the supernatant is transferred from the 96U plate to the 96V plate. The first track-mounted robot 17 throws the 96U base plate into the trash can and places the 96V base plate into the corresponding position of the fully automated plasmid extractor 11. Excess samples are placed in the temporary storage rack. After the automated consumables stack 19 delivers a new 1mL pipette tip box, nucleic acid washing plate, and 96V base plate to the automated liquid workstation 1 for plasmid library construction, washing solution is added to the 96V base plate and elution solution is added to the nucleic acid elution plate. The 96V base plate is stacked under the magnetic rod sleeve, and the eluted nucleic acid plate is placed into the fully automated plasmid extractor 11. The fully automated plasmid extractor 11 starts running. After the operation is completed, the first track-mounted robot 17 throws the plates except for the nucleic acid elution plate into the trash can. The extracted nucleic acid is placed in the refrigerator 16, and the experiment ends.

[0078] In the description of this utility model, it should be understood that the terms "longitudinal", "lateral", "up", "down", "front", "back", "left", "right", "vertical", "horizontal", "top", "bottom", "inner", "outer", etc., indicate the orientation or positional relationship based on the orientation or positional relationship shown in the accompanying drawings. They are only for the convenience of describing this utility model and do not indicate or imply that the device or element referred to must have a specific orientation, or be constructed and operated in a specific orientation. Therefore, they should not be construed as limitations on this utility model.

[0079] The embodiments described above are merely preferred embodiments of the present utility model and are not intended to limit the scope of the present utility model. Various modifications and improvements made to the technical solutions of the present utility model by those skilled in the art without departing from the spirit of the present utility model should fall within the protection scope defined by the claims of the present utility model.

Claims

1. A high-throughput plasmid construction functional island, characterized in that: The application relates to an automatic integrated table (22) comprising a box (24) mounted on the top surface of the automatic integrated table (22), wherein a conveying assembly is arranged in the box (24), and an automatic working assembly is arranged around the conveying assembly. The conveying assembly comprises a transfer table arranged at the center of the automatic integrated table (22), and a first slide rail track robot (17) and a second slide rail track robot (23) are arranged on the two sides of the transfer table respectively, and the first slide rail track robot (17) and the second slide rail track robot (23) are used for taking and placing required articles from the automatic working assembly and the transfer table.

2. The high-throughput plasmid construct functional island of claim 1, wherein: The automatic working assembly comprises a plasmid library construction automatic liquid workstation (1), an automatic consumable stack (19), an automatic coating instrument (6), an automatic bacteria scraping instrument (9), an automatic dispenser (13), an automatic shaking incubator (7), an automatic microbial colony picking instrument (10), an automatic incubator (8), a mobile robot interactive site (14), an automatic high-throughput centrifuge (12), an automatic film tearing machine (5), a full-automatic plasmid extraction instrument (11) and a refrigerator (16) which are arranged on the top surface of the automatic integrated table (22) and sequentially around the conveying assembly.

3. The high-throughput plasmid construct functional island of claim 2, wherein: The automatic consumable stack (19) is provided with a plurality of first supports which are fixedly connected with the automatic integrated table (22), and an electroporation instrument (2), an automatic PCR instrument (3) and a first automatic sealing and sealing film instrument (4) are sequentially arranged on the top surface of the first support, the electroporation instrument (2) is arranged close to the plasmid library construction automatic liquid workstation (1), and the first automatic sealing and sealing film instrument (4) is arranged close to the automatic coating instrument (6).

4. The high-throughput plasmid construct functional island of claim 3, wherein: A second support is arranged above the automatic dispenser (13) and is fixedly connected with the automatic integrated table (22), and a second automatic sealing and sealing film instrument is arranged on the top surface of the second support.

5. The high-throughput plasmid construct functional island of claim 2, wherein: The automatic shaking incubator (7) is provided with a plurality of the automatic shaking incubators.

6. The high-throughput plasmid construct functional island of claim 4, wherein: A third support is arranged above the mobile robot interactive site (14) and is fixedly connected with the automatic integrated table (22), and an automatic multifunctional enzyme label instrument (15) is arranged on the top surface of the third support.

7. The high-throughput plasmid construct functional island of claim 6, wherein: The plasmid library construction automatic liquid workstation (1), the electroporation instrument (2), the automatic PCR instrument (3), the first automatic sealing and sealing film instrument (4), the automatic high-throughput centrifuge (12), the automatic film tearing machine (5), the full-automatic plasmid extraction instrument (11) and the refrigerator (16) are arranged around the first slide rail track robot (17).

8. The high-throughput plasmid construct functional island of claim 4, wherein: The automatic coating instrument (6), the automatic bacteria scraping instrument (9), the automatic dispenser (13), the automatic shaking incubator (7), the automatic microbial colony picking instrument (10), and the automatic incubator (8) are arranged around the second sliding rail track robot (23).

9. The high-throughput plasmid construct functional island of claim 2, wherein: The mobile robot interaction site (14) is arranged opposite to the transfer table.

10. The high-throughput plasmid construct functional island of claim 2, wherein: A plurality of doors are installed on the box (24), and the outside of the box (24) is provided with a full-automatic mobile robot (18).

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