Nucleic acid synthesizer and nucleic acid chain synthesis method

Abstract

The invention relates to the technical field of biological pharmacy, and discloses a nucleic acid synthesizer and a nucleic acid chain synthesis method.The nucleic acid synthesizer comprises a plurality of monomer bottles, a plurality of solvent bottles, a liquid path assembly, a gas path assembly and a control unit; the gas path assembly is provided with a plurality of output ends, the gas input ends of the plurality of single bottles and the gas input ends of the plurality of solvent bottles are in butt joint with the output ends of the gas path assembly one by one, and the control unit is electrically connected with the gas path assembly. And the gas path assembly is used for respectively filling inert gas into the corresponding monomer bottles and the solvent bottles according to instructions of the control unit. The nucleic acid synthesizer has the advantages that liquid mixing is avoided during synthesis of organic solvents, gas mixing is avoided due to the fact that independent gas path systems are arranged between the solvent bottles, current synthesis important parameters can be displayed on the synthesizer in real time in the synthesis process, and the whole synthesizer is simple in structure, high in modular maintenance and the like.

Description

Technical Field

[0001] The present invention relates to the technical field of biosynthesis, in particular to a nucleic acid synthesizer and a nucleic acid chain synthesis method. Background Art

[0002] With the rapid development of molecular biology and biomedicine, small nucleic acid solid phase synthesis technology has become an indispensable and important tool in the field of research and treatment. This technology provides strong support for new drug development, gene therapy, gene regulation, etc., and shows broad application prospects in many fields such as drug research and development, gene therapy, and genetic diagnosis. In addition, small nucleic acid drugs have significant advantages in terms of strong specificity, simple design, short R&D cycle, rich targets, and deep transformation foundation. These advantages make small nucleic acid drugs have broad development prospects and important application value in the field of drug research and development, so corresponding nucleic acid synthesizer equipment has become an urgent need.

[0003] Existing nucleic acid synthesizers have problems such as solvent cross-talk during the synthesis process, gas cross-talk between solvent bottles, inaccurate solvent flow control, and inconvenient local operation and inspection when encountering emergencies during the synthesis process. Summary of the invention

[0004] In view of the deficiencies in the prior art, the present invention provides a nucleic acid synthesizer and a nucleic acid chain synthesis method, which solve the problems mentioned in the above background.

[0005] The present invention provides the following technical solutions: a nucleic acid synthesizer and a nucleic acid chain synthesis method, comprising: a plurality of monomer bottles, a plurality of solvent bottles, a liquid circuit component, a gas circuit component and a control unit;

[0006] The gas circuit assembly has a plurality of output ends, the gas input ends of the plurality of monomer bottles and the gas input ends of the plurality of solvent bottles are connected to the output ends of the gas circuit assembly one by one, the control unit is electrically connected to the gas circuit assembly, and the gas circuit assembly fills the corresponding monomer bottles and solvent bottles with inert gas according to the instructions of the control unit;

[0007] The liquid circuit assembly has a plurality of input ends, the outlet ends of the plurality of monomer bottles and the outlet ends of the plurality of solvent bottles are connected to the input ends of the liquid circuit assembly one by one, the control unit is electrically connected to the liquid circuit assembly, and the liquid circuit assembly transmits the reagents in the corresponding solvent bottles and monomer bottles according to the instructions of the control unit;

[0008] Wherein, the liquid circuit assembly includes a first system pump, a second system pump, a first monomer valve, a second monomer valve, a first solvent valve, a second solvent valve, a circulation valve, a first column position valve, a second column position valve and a synthesis column;

[0009] The first monomer valve, the second monomer valve, the first solvent valve and the second solvent valve all have a plurality of inlet ends, the outlet end of the monomer bottle is connected to the inlet end of the first monomer valve or the inlet end of the second monomer valve, the outlet ends of the first monomer valve and the second monomer valve are both connected to the inlet end of the first solvent valve, and the outlet end of the first solvent valve is connected to the inlet end of the first system pump;

[0010] The outlet end of the solvent bottle is connected to the inlet end of the first solvent valve or the inlet end of the second solvent valve, the outlet end of the second solvent valve is connected to the first inlet end of the circulation valve, and the first outlet end of the circulation valve is connected to the inlet end of the second system pump;

[0011] The outlet of the first system pump and the outlet of the second system pump are connected to a confluence node, the first column position valve, the synthesis column and the second column position valve are sequentially arranged between the confluence node and the second inlet end of the circulation valve, and the first system pump and the second system pump are both provided with exhaust valves.

[0012] Preferably, the liquid circuit assembly further comprises a waste liquid valve, the inlet end of the waste liquid valve is connected to the second outlet end of the circulation valve.

[0013] Preferably, the liquid circuit assembly further comprises: a system pressure sensor, a pre-column pressure sensor, a post-column pressure sensor, a UV detector, a bubble sensor and a conductivity meter detector;

[0014] The system pressure sensor is arranged between the confluence node and the inlet end of the first column position valve, the pre-column pressure sensor is arranged at the inlet end of the first column position valve, the post-column pressure sensor is arranged at the outlet end of the second column position valve, and the UV detector and the conductivity meter detector are arranged between the outlet end of the second column position valve and the second inlet end of the circulation valve.

[0015] Preferably, it also includes a whole machine housing assembly, which includes a housing sheet metal part, a display screen mounting block, a front panel, a module mounting plate, a stress-bearing sheet metal part, a water receiving trough, a side panel and a rear cover;

[0016] The outer shell sheet metal, the front panel, the side panel and the rear cover are all mounted on the stress-bearing sheet metal by means of locking screws, and are enclosed with the stress-bearing sheet metal to form a chassis;

[0017] The display screen mounting block and the module mounting plate are both mounted on the front panel by means of locking screws, a mounting groove is provided at the lower end of the housing sheet metal, and the water receiving trough is placed in the mounting groove;

[0018] A monomer bottle fixing block is installed on the side panel, and the monomer bottle is fixed to the side panel through the monomer bottle fixing block;

[0019] A handle is installed on the side panel, and a plurality of pipeline cable management buckles are installed on the surfaces of the front panel and the side panel.

[0020] Preferably, the gas circuit assembly comprises: a total gas source inlet, a first gas circuit unit and a second gas circuit unit.

[0021] An air circuit connection block is provided inside the chassis, the air circuit connection block has a main air inlet, and the main air source inlet is connected to the main air inlet of the air circuit connection block;

[0022] The first gas circuit unit and the second gas circuit unit are both connected to the total gas source inlet, the first gas circuit unit is used to fill the monomer bottle with inert gas, and the second gas circuit unit is used to fill the solvent bottle with inert gas;

[0023] Wherein, the first gas circuit unit comprises a first pressure gauge, a first pressure regulating valve, a monomer bottle manifold gas source outlet and a monomer bottle manifold, the monomer bottle manifold gas source outlet is connected to the gas inlet of the monomer bottle manifold, the first pressure gauge and the first pressure regulating valve are both arranged at the monomer bottle manifold gas source outlet, and are electrically connected to the control unit, and the gas outlet of the monomer bottle manifold is connected to the gas inlet of the monomer bottle;

[0024] The second gas circuit unit includes a second pressure gauge, a second pressure regulating valve, a solvent bottle manifold gas source outlet and a solvent bottle manifold. The solvent bottle manifold gas source outlet is connected to the gas inlet of the solvent bottle manifold through a gas pipeline. The second pressure gauge and the second pressure regulating valve are both arranged at the solvent bottle manifold gas source outlet and are electrically connected to the control unit. The gas outlet of the solvent bottle manifold is connected to the gas inlet of the solvent bottle.

[0025] Preferably, the monomer bottle and the solvent bottle are both located on the same side of the chassis, and the liquid circuit assembly is installed on the front panel of the chassis.

[0026] A method for synthesizing a nucleic acid chain comprises the following steps:

[0027] Step S1: system cleaning, switching the first solvent valve and the second solvent valve to the cleaning solvent inlet pipeline, and opening the first solvent valve, the second solvent valve, the first system pump and the second system pump. After the cleaning solvent is pumped by the second system pump and the first system pump, it is merged through a three-way joint and enters the system pressure sensor, the pre-column pressure sensor, the first column position valve, the synthesis column, the second column position valve, the post-column pressure sensor, the conductivity meter detector, the UV detector, the circulation valve and the waste valve. During the process of conveying the cleaning solvent, the circulation valve is in a discharge state;

[0028] Step S2: Deprotection, the first solvent valve and the second solvent valve are switched to the deprotection solvent inlet pipeline, and the deprotection solvent is delivered according to the liquid path in the above step S1. When the deprotection solvent passes through the synthesis column, the DMT group on the CPG is removed to expose the 5' hydroxyl group for the next coupling. After the deprotection is completed, the first solvent valve and the second solvent valve are switched to the cleaning solvent inlet pipeline, and the system cleaning step is performed again;

[0029] Step S3: Activation, switching the first solvent valve to the monomer reagent inlet pipeline, switching the second solvent valve to the activation solvent inlet pipeline, transporting the monomer reagent and the activation solvent according to the liquid path in the above step S1, after completing the activation, switching the first solvent valve and the second solvent valve to the cleaning solvent inlet pipeline, and performing the system cleaning step again;

[0030] Step S4: coupling, switching the circulation valve 18 to the circulation pumping state, and closing the second system pump, switching the first solvent valve to the monomer reagent inlet pipeline, the monomer reagent is pumped by the first system pump, and then passes through the system pressure sensor, the pre-column pressure sensor, the first column position valve, the synthesis column, the second column position valve, the post-column pressure sensor, the conductivity meter detector, the UV detector, and the circulation valve and returns to the inlet end of the first system pump again. After the coupling is completed, the first solvent valve and the second solvent valve are switched to the cleaning solvent inlet pipeline, and the system cleaning step is performed again;

[0031] Step S5: oxidation, switching the first solvent valve to the oxidant inlet pipeline, and turning on the first system pump to deliver the oxidant according to the liquid path in the above step S1. After the oxidation is completed, switching the first solvent valve and the second solvent valve to the cleaning solvent inlet pipeline, and performing the system cleaning step again;

[0032] Step S6: Capping, switch the solvent valve A and the second solvent valve to the capping solvent inlet pipeline, and open the first solvent valve, the second solvent valve, the first system pump and the second system pump, and the capping solvent enters the liquid circuit in the above step S1. After the capping is completed, switch the first solvent valve and the second solvent valve to the cleaning solvent inlet pipeline, and perform the system cleaning step again.

[0033] Preferably, it also includes:

[0034] Prior to a system cleaning step, setting a preset flow rate of a cleaning solvent via a control unit; and

[0035] During the system cleaning step, the flow rate of the cleaning solvent is monitored. When the flow rate value of the cleaning solvent reaches a preset flow rate, the system cleaning step is determined to be completed.

[0036] Preferably, it also includes:

[0037] The flow rate of the deprotection solvent is set by a control unit before the activation step, and the flow rate of the deprotection solvent is monitored during the activation and washing step. When the flow rate value of the deprotection solvent reaches a preset flow rate, the activation step is judged to be completed;

[0038] The flow rate of the oxidant is set by a control unit before the oxidation step, and the flow rate of the oxidant is monitored during the oxidation step. When the flow rate value of the oxidant reaches a preset flow rate, it is determined that the oxidation step is completed;

[0039] Before the capping step, the flow rate of the capping solvent is set by the control unit, and the flow rate of the capping solvent is monitored during the capping step. When the flow rate value of the capping solvent reaches the preset flow rate, it is determined that the capping step is completed;

[0040] Before the deprotection step, a target UV value is set by a control unit, and during the deprotection process, the UV value is monitored by a UV detector. When the UV value reaches the target UV value, the deprotection step is judged to be completed; and before the coupling step, a preset coupling time is set by the control unit.

[0041] Preferably, it also includes:

[0042] The above steps are repeated until the oligonucleotide chain is extended to a predetermined length.

[0043] Compared with the prior art, the present invention has the following beneficial effects:

[0044] The present invention, by arranging the whole machine housing component, the liquid circuit component and the gas circuit system, synthesized organic solvents do not mix liquids, independent gas circuit systems are provided between solvent bottles to prevent gas mixing, and during the synthesis process, the current important synthesis parameters can be displayed in real time on the instrument. The whole machine has a simple structure, and the instrument can be modularly installed and disassembled, which is convenient for maintenance and repair. BRIEF DESCRIPTION OF THE DRAWINGS

[0045] Figure 1 It is a schematic diagram of the three-dimensional structure of the present invention;

[0046] Figure 2 This is a schematic diagram of the structure of the housing assembly of the whole machine of the present invention;

[0047] Figure 3 It is a schematic diagram of the structure of the liquid circuit assembly of the present invention;

[0048] Figure 4 It is a side view structural schematic diagram of the present invention;

[0049] Figure 5 It is a schematic diagram of the structure of the gas circuit system of the present invention;

[0050] Figure 6 This is the liquid circuit diagram of the present invention.

[0051] In the figure: 1. Shell sheet metal; 2. Display screen mounting block; 3. Front panel; 4. Module mounting plate; 5. Force-bearing sheet metal; 6. Water tank; 7. Side panel; 8. Pipeline cable clip; 9. Handle; 10. Rear cover; 11. First system pump; 12. Exhaust valve; 13. System pressure sensor; 14. Pre-column pressure sensor; 15. Post-column pressure sensor; 16. Synthetic column; 17. First column position valve; 18. Circulation valve; 19. UV detector; 20. Waste valve; 21. First monomer valve ; 22. Monomer bottle; 23. Second solvent valve; 24. Acetonitrile manifold; 25. Bubble sensor; 26. Conductivity meter detector; 27. Second column valve; 28. Second system pump; 29. ​​Second monomer valve; 30. First solvent valve; 31. First pressure gauge; 32. First pressure regulating valve; 33. Second pressure gauge; 34. Second pressure regulating valve; 35. Total gas source inlet; 36. Solvent bottle manifold gas source outlet; 37. Monomer bottle manifold gas source outlet; 38. Solvent bottle manifold; 39. Monomer bottle manifold. DETAILED DESCRIPTION

[0052] The following will be combined with the drawings in the embodiments of the present invention to clearly and completely describe the technical solutions in the embodiments of the present invention. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0053] See also Figure 1-6 , a nucleic acid synthesizer, comprising: a plurality of monomer bottles 22, a plurality of solvent bottles (not shown in the figure), a liquid circuit assembly, a gas circuit assembly and a control unit;

[0054] The gas circuit assembly has multiple output ends, and the gas input ends of the multiple monomer bottles 22 and the gas input ends of the multiple solvent bottles are connected to the output ends of the gas circuit assembly one by one. The control unit is electrically connected to the gas circuit assembly, and the gas circuit assembly fills the corresponding monomer bottles 22 and solvent bottles with inert gas according to the instructions of the control unit.

[0055] The liquid circuit assembly has multiple input ends, and the outlet ends of multiple monomer bottles 22 and the outlet ends of multiple solvent bottles are connected to the input ends of the liquid circuit assembly one by one. The control unit is electrically connected to the liquid circuit assembly, and the liquid circuit assembly transmits the reagents in the corresponding solvent bottles and monomer bottles 22 according to the instructions of the control unit;

[0056] The liquid circuit assembly includes a first system pump 11, a second system pump 28, a first monomer valve 21, a second monomer valve 29, a first solvent valve 23, a second solvent valve 30, a circulation valve 18, a first column position valve 17, a second column position valve 27 and a synthesis column 16;

[0057] The first monomer valve 21, the second monomer valve 29, the first solvent valve 23 and the second solvent valve 30 all have a plurality of inlet ends, the outlet end of the monomer bottle 22 is connected to the inlet end of the first monomer valve 21 or the inlet end of the second monomer valve 29, the outlet ends of the first monomer valve 21 and the second monomer valve 29 are both connected to the inlet end of the first solvent valve 23, and the outlet end of the first solvent valve 23 is connected to the inlet of the first system pump 11;

[0058] The outlet end of the solvent bottle is connected to the inlet end of the first solvent valve 23 or the inlet end of the second solvent valve 30, the outlet end of the second solvent valve 30 is connected to the first inlet end of the circulation valve 18, and the first outlet end of the circulation valve 18 is connected to the inlet of the second system pump 28;

[0059] The outlet of the first system pump 11 and the outlet of the second system pump 28 are connected to a junction node, the first column position valve 17, the synthesis column 16 and the second column position valve 27 are sequentially arranged between the junction node and the second inlet end of the circulation valve 18, and the first system pump 11 and the second system pump 28 are both provided with an exhaust valve 12. Preferably, the first system pump 11 and the second system pump 28 are both plunger pumps, and the plunger pumps have a metering function. In this way, during the synthesis process, the first system pump 11 and the second system pump 28 can accurately control the flow rate of the reagent.

[0060] Specifically, the monomer bottle 22 is used to hold monomer reagents, and the solvent bottle is used to hold synthesis reaction reagents, which include but are not limited to cleaning solvents, deprotection solvents, activation solvents, oxidants, and capping solvents.

[0061] The synthesis column 16 has a reaction chamber, and the monomer reagent and the synthesis reaction reagent transmitted by the liquid circuit component undergo a synthesis reaction in the reaction chamber of the synthesis column 16. A carrier is provided in the reaction chamber, and the carrier is used to collect the synthesized product. The control unit is used to control the switching of the pipeline, and the reagents transmitted by the liquid circuit component are switched according to the steps of the synthesis process.

[0062] The liquid circuit assembly further includes a waste liquid valve 20 , the inlet end of the waste liquid valve 20 is connected to the second outlet end of the circulation valve 18 .

[0063] The liquid circuit assembly also includes: a system pressure sensor 13, a pre-column pressure sensor 14, a post-column pressure sensor 15, a UV detector 19, a bubble sensor 25 and a conductivity meter detector 26;

[0064] Among them, the conductivity meter detector 26 is used to monitor the conductivity of the organic solvent. The operator assists in judging whether the organic solvent is correct based on the conductivity detection result. The UV detector 19 is used to detect the removal of the DMT group on the CPG. When the value displayed by the UV detector 19 reaches a certain value, it is judged that the deprotection process is completed. The bubble sensor 25 is used to monitor whether the liquid circuit is cut off and prompt the operator to replenish the organic solvent.

[0065] The system pressure sensor 13 is arranged between the confluence node and the inlet end of the first column position valve 17, the pre-column pressure sensor 14 is arranged at the inlet end of the first column position valve 17, the post-column pressure sensor 15 is arranged at the outlet end of the second column position valve 27, and the UV detector 19 and the conductivity meter detector 26 are arranged between the outlet end of the second column position valve 27 and the second inlet end of the circulation valve 18.

[0066] The whole machine housing assembly is also included, and the whole machine housing assembly includes a housing sheet metal part 1, a display screen mounting block 2, a front panel 3, a module mounting plate 4, a stress-bearing sheet metal part 5, a water receiving trough 6, a side panel 7 and a rear cover 10;

[0067] The outer shell sheet metal part 1, the front panel 3, the side panel 7 and the rear cover 10 are all mounted on the stress-bearing sheet metal part 5 by means of locking screws, and are enclosed with the stress-bearing sheet metal part 5 to form a chassis;

[0068] The display screen mounting block 2 and the module mounting plate 4 are both mounted on the front panel 3 by means of locking screws. A mounting groove is provided at the lower end of the housing sheet metal part 1, and the water receiving tank 6 is placed in the mounting groove.

[0069] A monomer bottle fixing block is installed on the side panel 7, and the monomer bottle 22 is fixed on the side panel 7 through the monomer bottle fixing block;

[0070] A handle 9 is installed on the side panel 7, and a plurality of pipeline cable buckles 8 are installed on the surface of the front panel 3 and the side panel 7. Preferably, the handle 9 is a hidden handle, which is convenient for moving the instrument and beautiful.

[0071] In this embodiment, the liquid circuit assembly is installed on the front panel 3 of the chassis and a water receiving tank is provided at the bottom. The circuit system is provided on the inner side of the chassis, which prevents the liquid circuit assembly from leaking liquid in an emergency and causing the circuit system to fail, thus achieving water-electricity separation. In addition, the components of the liquid circuit assembly are arranged on the front panel 3, so that the liquid circuit condition can be directly observed in the face of an emergency during the early process synthesis exploration.

[0072] In this embodiment, the nucleic acid synthesizer is provided with 8 solvent bottles and 12 monomer bottles 22. The monomer bottles 22 are fixed on the right side wall of the chassis and arranged in three rows and four columns. The solvent bottles are placed on the right side of the chassis.

[0073] In this embodiment, the monomer bottle 22 and the solvent bottle are both located on the right side of the chassis, so that the pipeline arrangement does not need to bypass the components, making the pipeline arrangement more streamlined and beautiful. In addition, the pipeline connection between each component of the liquid circuit assembly is streamlined and cross-connected, which can reduce the dead volume of the liquid circuit, thereby reducing the waste of solvent and greatly saving costs.

[0074] In this embodiment, the solvent valves and the solvent inlet positions are arranged in sequence, and the solvents are switched in coordination with the synthesis process, which can prevent the occurrence of liquid cross-talk during the synthesis process.

[0075] The gas circuit assembly includes: a total gas source inlet 35, a first gas circuit unit and a second gas circuit unit.

[0076] An air circuit connection block (not shown) is provided inside the chassis, and the air circuit connection block has a main air inlet, and the main air source inlet 35 is connected to the main air inlet of the air circuit connection block;

[0077] The first gas circuit unit and the second gas circuit unit are both connected to the main gas source inlet 35, the first gas circuit unit is used to fill the monomer bottle with inert gas, and the second gas circuit unit is used to fill the solvent bottle with inert gas;

[0078] Among them, the first gas circuit unit includes a first pressure gauge 31, a first pressure regulating valve 32, a monomer bottle manifold gas source outlet 37 and a monomer bottle manifold 39, the monomer bottle manifold gas source outlet 37 is connected to the air inlet of the monomer bottle manifold 39, the first pressure gauge 31 and the first pressure regulating valve 32 are both arranged at the monomer bottle manifold gas source outlet 37, and are electrically connected to the control unit, and the gas outlet of the monomer bottle manifold 39 is connected to the gas inlet of the monomer bottle 22;

[0079] The second gas circuit unit includes a second pressure gauge 33, a second pressure regulating valve 34, a solvent bottle manifold gas source outlet 36 and a solvent bottle manifold 38. The solvent bottle manifold gas source outlet 36 is connected to the gas inlet of the solvent bottle manifold 38 through a gas pipeline. The second pressure gauge 33 and the second pressure regulating valve 34 are both arranged at the solvent bottle manifold gas source outlet 36 and are electrically connected to the control unit. The gas outlet of the solvent bottle manifold 38 is connected to the gas inlet of the solvent bottle.

[0080] The gas circuit assembly also includes an acetonitrile manifold 24 having an inlet end and multiple outlet ends. The solvent bottle manifold 38 is connected to the solvent bottle manifold gas source outlet 36 through the acetonitrile manifold 24, and the monomer bottle manifold 39 is connected to the monomer bottle manifold gas source outlet 37 through the acetonitrile manifold 24.

[0081] In this embodiment, the total gas source inlet 35, the solvent bottle manifold gas source outlet 36 and the monomer bottle manifold gas source outlet 37 are arranged on one side of the chassis, and the solvent bottle manifold 38 and the monomer bottle manifold 39 are arranged on the other side of the chassis.

[0082] A method for synthesizing a nucleic acid chain comprises the following steps:

[0083] Step S1: System cleaning, switching the first solvent valve 30 and the second solvent valve 23 to the cleaning solvent inlet pipeline, and opening the first solvent valve 30, the second solvent valve 23, the first system pump 11 and the second system pump 28. After the cleaning solvent is pumped by the second system pump 28 and the first system pump 11, it is combined through the three-way joint and enters the system pressure sensor 13, the pre-column pressure sensor 14, the first column position valve 17, the synthesis column 16, the second column position valve 27, the post-column pressure sensor 27, the conductivity meter detector 26, the UV detector 19, the circulation valve 18 and the waste valve 20. During the process of conveying the cleaning solvent, the circulation valve 18 is in a discharge state;

[0084] Step S2: Deprotection, the first solvent valve 30 and the second solvent valve 23 are switched to the deprotection solvent inlet pipeline, and the deprotection solvent is transported according to the liquid path in the above step S1. When the deprotection solvent passes through the synthesis column 16, the DMT group on the CPG is removed to expose the 5' hydroxyl group for the next coupling. After the deprotection is completed, the first solvent valve 30 and the second solvent valve 23 are switched to the cleaning solvent inlet pipeline, and the system cleaning step is performed again;

[0085] Step S3: Activation, switching the first solvent valve 30 to the monomer reagent inlet pipeline, switching the second solvent valve 23 to the activation solvent inlet pipeline, transporting the monomer reagent and the activation solvent according to the liquid path in the above step S1, after the activation is completed, switching the first solvent valve 30 and the second solvent valve 23 to the cleaning solvent inlet pipeline, and performing the system cleaning step again;

[0086] Step S4: coupling, switching the circulation valve 18 to the circulation pumping state, and closing the second system pump 28, switching the first solvent valve 30 to the monomer reagent inlet pipeline, the monomer reagent is pumped by the first system pump 11, and then passes through the system pressure sensor 13, the pre-column pressure sensor 14, the first column position valve 17, the synthesis column 16, the second column position valve 27, the post-column pressure sensor 27, the conductivity meter detector 26, the UV detector 19, and the circulation valve 18 and returns to the inlet end of the first system pump 11 again. After completing the coupling, the first solvent valve 30 and the second solvent valve 23 are switched to the cleaning solvent inlet pipeline, and the system cleaning step is performed again;

[0087] Step S5: oxidation, switching the first solvent valve 30 to the oxidant inlet pipeline, and turning on the first system pump 11, delivering the oxidant according to the liquid path in the above step S1, after the oxidation is completed, switching the first solvent valve 30 and the second solvent valve 23 to the cleaning solvent inlet pipeline, and performing the system cleaning step again;

[0088] Step S6: capping, switching the solvent valve A30 and the solvent valve B23 to the capping solvent inlet pipeline, and opening the first solvent valve 30, the second solvent valve 23, the first system pump 11 and the second system pump 28, so that the capping solvent enters the liquid circuit in the above step S1. After the capping is completed, the first solvent valve 30 and the second solvent valve 23 are switched to the cleaning solvent inlet pipeline, and the system cleaning step is performed again;

[0089] The synthesis method provided in this embodiment is suitable for synthesizing oligonucleotide chains.

[0090] Also includes:

[0091] Prior to a system cleaning step, setting a preset flow rate of a cleaning solvent via a control unit; and

[0092] During the system cleaning step, the flow rate of the cleaning solvent is monitored. When the flow rate value of the cleaning solvent reaches a preset flow rate, the system cleaning step is determined to be completed.

[0093] Also includes:

[0094] The flow rate of the deprotection solvent is set by a control unit before the activation step, and the flow rate of the deprotection solvent is monitored during the activation and washing step. When the flow rate value of the deprotection solvent reaches a preset flow rate, the activation step is judged to be completed;

[0095] The flow rate of the oxidant is set by a control unit before the oxidation step, and the flow rate of the oxidant is monitored during the oxidation step. When the flow rate value of the oxidant reaches a preset flow rate, it is determined that the oxidation step is completed;

[0096] Before the capping step, the flow rate of the capping solvent is set by the control unit, and the flow rate of the capping solvent is monitored during the capping step. When the flow rate value of the capping solvent reaches the preset flow rate, it is determined that the capping step is completed;

[0097] Before the deprotection step, a target UV value is set by the control unit, and during the deprotection process, the UV value is monitored by the UV detector 19. When the UV value reaches the target UV value, the deprotection step is judged to be completed; and before the coupling step, a preset coupling time is set by the control unit.

[0098] Also includes:

[0099] The above steps are repeated until the oligonucleotide chain is extended to a preset length;

[0100] During the entire reaction process, the monomer bottle and the solvent bottle need to be filled with argon or nitrogen for protection to achieve the purpose of isolating from the air.

[0101] Although embodiments of the present invention have been shown and described, it will be appreciated by those skilled in the art that various changes, modifications, substitutions and variations may be made to the embodiments without departing from the principles and spirit of the present invention, and that the scope of the present invention is defined by the appended claims and their equivalents.

Claims

1. A nucleic acid synthesizer, characterized in that: include: A plurality of monomer bottles (22), a plurality of solvent bottles, a liquid circuit assembly, a gas circuit assembly and a control unit; The gas circuit assembly has a plurality of output ends, the gas input ends of the plurality of monomer bottles (22) and the gas input ends of the plurality of solvent bottles are connected to the output ends of the gas circuit assembly one by one, the control unit is electrically connected to the gas circuit assembly, and the gas circuit assembly fills the corresponding monomer bottles (22) and solvent bottles with inert gas according to the instructions of the control unit; The liquid circuit assembly has a plurality of input ends, the outlet ends of the plurality of monomer bottles (22) and the outlet ends of the plurality of solvent bottles are connected one by one to the input end of the liquid circuit assembly, the control unit is electrically connected to the liquid circuit assembly, and the liquid circuit assembly transmits the reagents in the corresponding solvent bottles and monomer bottles (22) according to the instructions of the control unit; The liquid circuit assembly comprises a first system pump (11), a second system pump (28), a first monomer valve (21), a second monomer valve (29), a first solvent valve (23), a second solvent valve (30), a circulation valve (18), a first column position valve (17), a second column position valve (27) and a synthesis column (16); The first monomer valve (21), the second monomer valve (29), the first solvent valve (23) and the second solvent valve (30) all have a plurality of inlet ends, the outlet end of the monomer bottle (22) is connected to the inlet end of the first monomer valve (21) or the inlet end of the second monomer valve (29), the outlet ends of the first monomer valve (21) and the second monomer valve (29) are both connected to the inlet end of the first solvent valve (23), and the outlet end of the first solvent valve (23) is connected to the inlet of the first system pump (11); The outlet end of the solvent bottle is connected to the inlet end of the first solvent valve (23) or the inlet end of the second solvent valve (30), the outlet end of the second solvent valve (30) is connected to the first inlet end of the circulation valve (18), and the first outlet end of the circulation valve (18) is connected to the inlet end of the second system pump (28); The outlet of the first system pump (11) and the outlet of the second system pump (28) are connected to a junction node, the first column position valve (17), the synthesis column (16) and the second column position valve (27) are arranged in sequence between the junction node and the second inlet end of the circulation valve (18), and the first system pump (11) and the second system pump (28) are both provided with an exhaust valve (12).

2. The nucleic acid synthesizer according to claim 1, characterized in that: The liquid circuit assembly further comprises a waste liquid valve (20), the inlet end of the waste liquid valve (20) being connected to the second outlet end of the circulation valve (18).

3. The nucleic acid synthesizer according to claim 1, characterized in that: The liquid circuit assembly further comprises: a system pressure sensor (13), a pre-column pressure sensor (14), a post-column pressure sensor (15), a UV detector (19), a bubble sensor (25) and a conductivity meter detector (26); The system pressure sensor (13) is arranged between the confluence node and the inlet end of the first column position valve (17), the pre-column pressure sensor (14) is arranged at the inlet end of the first column position valve (17), the post-column pressure sensor (15) is arranged at the outlet end of the second column position valve (27), and the UV detector (19) and the conductivity meter detector (26) are arranged between the outlet end of the second column position valve (27) and the second inlet end of the circulation valve (18).

4. The nucleic acid synthesizer according to claim 1, characterized in that: It also includes a whole machine housing assembly, which includes a housing sheet metal part (1), a display screen mounting block (2), a front panel (3), a module mounting plate (4), a stress-bearing sheet metal part (5), a water receiving trough (6), a side panel (7) and a rear cover plate (10); The outer shell sheet metal part (1), the front panel (3), the side panel (7) and the rear cover (10) are all mounted on the load-bearing sheet metal part (5) by means of locking screws, and are enclosed with the load-bearing sheet metal part (5) to form a chassis; The display screen mounting block (2) and the module mounting plate (4) are both mounted on the front panel (3) by means of locking screws; a mounting groove is provided at the lower end of the housing sheet metal component (1), and the water receiving groove (6) is placed in the mounting groove; A monomer bottle fixing block is installed on the side panel (7), and the monomer bottle (22) is fixed on the side panel (7) via the monomer bottle fixing block; A handle (9) is installed on the side panel (7), and a plurality of pipeline cable management buckles (8) are installed on the surfaces of the front panel (3) and the side panel (7).

5. The nucleic acid synthesizer according to claim 4, characterized in that: The gas circuit assembly comprises: a total gas source inlet (35), a first gas circuit unit and a second gas circuit unit, An air circuit connection block is arranged inside the chassis, the air circuit connection block has a main air inlet, and the main air source inlet (35) is connected to the main air inlet of the air circuit connection block; The first gas circuit unit and the second gas circuit unit are both connected to the total gas source inlet (35), the first gas circuit unit is used to fill the monomer bottle with inert gas, and the second gas circuit unit is used to fill the solvent bottle with inert gas; Wherein, the first gas circuit unit comprises a first pressure gauge (31), a first pressure regulating valve (32), a monomer bottle manifold gas source outlet (37) and a monomer bottle manifold (39); the monomer bottle manifold gas source outlet (37) is connected to the gas inlet of the monomer bottle manifold (39); the first pressure gauge (31) and the first pressure regulating valve (32) are both arranged at the monomer bottle manifold gas source outlet (37) and are electrically connected to the control unit; the gas outlet of the monomer bottle manifold (39) is connected to the gas inlet of the monomer bottle (22); The second gas circuit unit comprises a second pressure gauge (33), a second pressure regulating valve (34), a solvent bottle manifold gas source outlet (36) and a solvent bottle manifold (38); the solvent bottle manifold gas source outlet (36) is connected to the gas inlet of the solvent bottle manifold (38) via a gas pipeline; the second pressure gauge (33) and the second pressure regulating valve (34) are both arranged at the solvent bottle manifold gas source outlet (36) and are electrically connected to the control unit; the gas outlet of the solvent bottle manifold (38) is connected to the gas inlet of the solvent bottle.

6. The nucleic acid synthesizer according to claim 4, characterized in that: The monomer bottle and the solvent bottle are both located on the same side of the chassis, and the liquid path assembly is installed on the front panel (3) of the chassis.

7. A method for synthesizing a nucleic acid chain, characterized in that: The following steps are involved: Step S1: System cleaning, switching the first solvent valve (30) and the second solvent valve (23) to the cleaning solvent inlet pipeline, and opening the first solvent valve (30), the second solvent valve (23), the first system pump (11) and the second system pump (28); after the cleaning solvent is pumped by the second system pump (28) and the first system pump (11), it is combined through a three-way joint and enters the system pressure sensor (13), the pre-column pressure sensor (14), the first column position valve (17), the synthesis column (16), the second column position valve (27), the post-column pressure sensor (27), the conductivity meter detector (26), the UV detector (19), the circulation valve (18) and the waste valve (20); during the process of conveying the cleaning solvent, the circulation valve (18) is in a discharge state; Step S2: Deprotection, the first solvent valve (30) and the second solvent valve (23) are switched to the deprotection solvent inlet pipeline, and the deprotection solvent is transported according to the liquid path in the above step S1. When the deprotection solvent passes through the synthesis column (16), the DMT group on the CPG is removed to expose the 5' hydroxyl group for the next coupling. After the deprotection is completed, the first solvent valve (30) and the second solvent valve (23) are switched to the cleaning solvent inlet pipeline, and the system cleaning step is performed again; Step S3: Activation, switching the first solvent valve (30) to the monomer reagent inlet pipeline, switching the second solvent valve (23) to the activation solvent inlet pipeline, and transporting the monomer reagent and the activation solvent according to the liquid path in the above step S1. After the activation is completed, the first solvent valve (30) and the second solvent valve (23) are switched to the cleaning solvent inlet pipeline, and the system cleaning step is performed again; Step S4: coupling, switching the circulation valve 18 to the circulation pumping state, and closing the second system pump (28), switching the first solvent valve (30) to the monomer reagent inlet pipeline, the monomer reagent is pumped by the first system pump (11), passes through the system pressure sensor (13), the pre-column pressure sensor (14), the first column position valve (17), the synthesis column (16), the second column position valve (27), the post-column pressure sensor (27), the conductivity meter detector (26), the UV detector (19), the circulation valve (18), and then returns to the inlet end of the first system pump (11). After the coupling is completed, the first solvent valve (30) and the second solvent valve (23) are switched to the cleaning solvent inlet pipeline, and the system cleaning step is performed again; Step S5: oxidation, switching the first solvent valve (30) to the oxidant inlet pipeline, and turning on the first system pump (11), delivering the oxidant according to the liquid path in the above step S1, after the oxidation is completed, switching the first solvent valve (30) and the second solvent valve (23) to the cleaning solvent inlet pipeline, and performing the system cleaning step again; Step S6: Capping, switching the first solvent valve (30) and the second solvent valve (23) to the capping solvent inlet pipeline, and opening the first solvent valve (30), the second solvent valve (23), the first system pump (11) and the second system pump (28), so that the capping solvent enters the liquid circuit in the above step S1. After the capping is completed, switching the first solvent valve (30) and the second solvent valve (23) to the cleaning solvent inlet pipeline, and performing the system cleaning step again.

8. The method for synthesizing a nucleic acid chain according to claim 7, characterized in that: Also includes: Prior to a system cleaning step, a preset flow rate of the cleaning solvent is set by the control unit; as well as During the system cleaning step, the flow rate of the cleaning solvent is monitored. When the flow rate value of the cleaning solvent reaches a preset flow rate, the system cleaning step is determined to be completed.

9. The method for synthesizing a nucleic acid chain according to claim 7, characterized in that: Also includes: The flow rate of the deprotection solvent is set by a control unit before the activation step, and the flow rate of the deprotection solvent is monitored during the activation and washing step. When the flow rate value of the deprotection solvent reaches a preset flow rate, the activation step is judged to be completed; The flow rate of the oxidant is set by a control unit before the oxidation step, and the flow rate of the oxidant is monitored during the oxidation step. When the flow rate value of the oxidant reaches a preset flow rate, it is determined that the oxidation step is completed; Before the capping step, the flow rate of the capping solvent is set by the control unit, and the flow rate of the capping solvent is monitored during the capping step. When the flow rate value of the capping solvent reaches the preset flow rate, it is determined that the capping step is completed; Before the deprotection step, a target UV value is set by a control unit, and during the deprotection process, the UV value is monitored by a UV detector (19), and when the UV value reaches the target UV value, the deprotection step is judged to be completed; as well as Prior to the coupling step, a preset time for coupling is set by the control unit.

10. The method for synthesizing a nucleic acid chain according to claim 7, characterized in that: Also includes: The above steps are repeated until the oligonucleotide chain is extended to a predetermined length.

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