WO2013075263A1 - 热对流聚合酶连锁反应装置 - Google Patents

热对流聚合酶连锁反应装置 Download PDF

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Publication number
WO2013075263A1
WO2013075263A1 PCT/CN2011/001941 CN2011001941W WO2013075263A1 WO 2013075263 A1 WO2013075263 A1 WO 2013075263A1 CN 2011001941 W CN2011001941 W CN 2011001941W WO 2013075263 A1 WO2013075263 A1 WO 2013075263A1
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Prior art keywords
test tube
polymerase chain
chain reaction
thermal convection
heat sink
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PCT/CN2011/001941
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English (en)
French (fr)
Inventor
苏城
邓秉华
Original Assignee
瑞基海洋生物科技股份有限公司
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Priority to KR1020147002887A priority Critical patent/KR101691466B1/ko
Priority to CN201180072273.1A priority patent/CN103649301B/zh
Priority to CA2856345A priority patent/CA2856345C/en
Priority to IN3868CHN2014 priority patent/IN2014CN03868A/en
Priority to PCT/CN2011/001941 priority patent/WO2013075263A1/zh
Priority to EP11876152.7A priority patent/EP2784150B1/en
Publication of WO2013075263A1 publication Critical patent/WO2013075263A1/zh

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    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L7/00—Heating or cooling apparatus; Heat insulating devices
    • B01L7/52—Heating or cooling apparatus; Heat insulating devices with provision for submitting samples to a predetermined sequence of different temperatures, e.g. for treating nucleic acid samples
    • B01L7/525—Heating or cooling apparatus; Heat insulating devices with provision for submitting samples to a predetermined sequence of different temperatures, e.g. for treating nucleic acid samples with physical movement of samples between temperature zones
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L2300/00—Additional constructional details
    • B01L2300/08—Geometry, shape and general structure
    • B01L2300/0832—Geometry, shape and general structure cylindrical, tube shaped
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L2300/00—Additional constructional details
    • B01L2300/08—Geometry, shape and general structure
    • B01L2300/0832—Geometry, shape and general structure cylindrical, tube shaped
    • B01L2300/0838—Capillaries
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L2300/00—Additional constructional details
    • B01L2300/18—Means for temperature control
    • B01L2300/1805—Conductive heating, heat from thermostatted solids is conducted to receptacles, e.g. heating plates, blocks
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L2300/00—Additional constructional details
    • B01L2300/18—Means for temperature control
    • B01L2300/1861—Means for temperature control using radiation
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L2400/00—Moving or stopping fluids
    • B01L2400/04—Moving fluids with specific forces or mechanical means
    • B01L2400/0403—Moving fluids with specific forces or mechanical means specific forces
    • B01L2400/0442—Moving fluids with specific forces or mechanical means specific forces thermal energy, e.g. vaporisation, bubble jet
    • B01L2400/0445—Natural or forced convection
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L9/00—Supporting devices; Holding devices
    • B01L9/06—Test-tube stands; Test-tube holders
    • B01L9/065—Test-tube stands; Test-tube holders specially adapted for capillary tubes

Definitions

  • the invention relates to a polymerase chain reaction, and in particular to a thermal convection polymerase chain reaction device. current technology
  • PCR polymerase chain reaction
  • the temperature of the denaturation reaction is typically between 90 and 97 ° C; the temperature of the primer bonding reaction is selected depending on the melting temperature of the primer used, typically between 35 and 65 ° C; typical of the extension reaction The temperature is 72 °C.
  • the thermal convection polymerase chain reaction method mainly embeds the bottom of the test tube containing the PCR mixture in hot water, and the rest of the test tube is exposed to room temperature air to dissipate heat, so that the bottom of the PCR mixture is gradually reduced to the surface.
  • the temperature gradient is 9, for example. C to 35 °C to induce thermal convection, allowing the PCR mixture to undergo different reaction steps through different temperature zones.
  • a primary object of the present invention is to provide a thermal convection polymerase chain reaction apparatus which ensures that the temperature of the surface of the mixed solution during the PCR reaction is lower than the temperature required for the bonding reaction.
  • the present invention provides a thermal convection polymerase chain reaction device for holding a test tube to carry out a polymerase chain reaction in the test tube, characterized in that the device comprises: a heat dissipation
  • the seat has a body, and a passage is disposed through the body for receiving the test tube; wherein the passage has a large diameter section and a small diameter section located below the large diameter section.
  • a test tube rack is disposed above the heat sink, and the test tube rack has a receiving hole for inserting the test tube.
  • the receiving hole of the test tube rack has a large diameter portion, a shoulder portion and a small diameter portion from top to bottom, wherein the shoulder portion abuts against the first step portion of the test tube.
  • the bottom end of the test tube rack is inserted into the channel of the heat sink.
  • the inner diameter of the small diameter section of the passage is gradually reduced from the top to the bottom.
  • the heat sink is made of metal.
  • the present invention ensures that the temperature of the surface of the mixed solution during the PCR reaction is lower than the temperature required for the adhesion reaction of the primer.
  • the invention is applicable to the thermal convection polymerase chain reaction in the field of biotechnology.
  • Figure 1 is a schematic view of a preferred embodiment of the present invention
  • Figure 2 is a perspective exploded view of a preferred embodiment of the present invention.
  • the thermal convection polymerase chain reaction apparatus 10 mainly comprises a heat sink 20 and a test tube rack 30.
  • the heat sink 20 has a body 22, and a channel 24 is disposed through the body 22 for receiving the test tube 12, wherein the passage 24 has a large diameter section 241 and a small diameter section 242 located below the large diameter section 241.
  • the heat sink 20 Since the middle section 123 to the upper section 122 of the test tube 12 are disposed in the passage 24 of the heat sink 20, and the heat sink 20 is made of a material having a high thermal conductivity such as an aluminum alloy, the heat energy of the PCR mixture can be conducted to the surrounding air to The heat sink 20 is more efficiently dissipated, so that the temperature of the surface of the mixed liquid is maintained below the primer bonding temperature during the whole reaction process, for example, 10 to 55 ° C, to avoid the PCR reaction due to the excessive surface temperature of the mixed liquid. Interrupted. Furthermore, the mixed solution located in the large diameter section 241 dissipates heat slowly, and the mixed solution located in the small diameter section 242 dissipates heat rapidly. After experimental tests, it is found that the heat dissipation effect of the structure of the heat sink 20 is most favorable for the PCR reaction.
  • a test tube rack 30 is disposed above the heat sink 20, and the test tube rack 30 has a receiving hole 32 for inserting the test tube 12, so that the test tube 12 can be stably positioned.
  • the outer wall of the test tube 12 is prevented from coming into contact with the inner edge of the heat sink 20, so that the temperature of the mixture is lowered too quickly.
  • the receiving hole 32 of the test tube rack 30 can be designed to be provided with a large diameter portion 321 , a shoulder portion 322 and a small diameter portion 323 from top to bottom, wherein the shoulder portion 322 is used for one of the test tubes 12 .
  • the step 121 abuts, whereby the test tube 12 can be stably positioned.
  • a support base 40 is disposed under the heat sink 20, and the test tube bottom 124 is heated by a heat source (not shown) to maintain the temperature of the PCR mixture in the bottom portion 124 at 90 to 97 °C.
  • the bottom end 301 of the test tube rack 30 is inserted into the passage 24 of the heat sink 20, so that the test tube rack 30 is firmly disposed above the heat sink 20.
  • the inner diameter of the small diameter section 242 of the passage 24 is gradually reduced from the top to the bottom.
  • the structure of the device may also be varied without departing from the spirit of the invention.
  • the heat sink 20 may be provided with a plurality of channels 24 for simultaneous reaction of a plurality of test tubes 12 (shown in Figure 2). All such structural changes that are easily conceivable are covered by the scope of patent protection of the present invention.
  • the thermal convection polymerase chain reaction device provided by the invention is suitable for the thermal convection polymerization enzyme chain reaction in the biotechnology field, and ensures that the temperature of the surface of the mixed solution is lower than the temperature required for the adhesion reaction of the primer during the PCR reaction.

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  • Health & Medical Sciences (AREA)
  • Chemical & Material Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Biochemistry (AREA)
  • General Health & Medical Sciences (AREA)
  • Molecular Biology (AREA)
  • Clinical Laboratory Science (AREA)
  • Chemical Kinetics & Catalysis (AREA)
  • Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)
  • Apparatus Associated With Microorganisms And Enzymes (AREA)

Abstract

本发明提供了一种热对流聚合酶链锁反应（PCR）装置，用于承置一试管，使PCR在所述试管内进行，所述装置包含：一散热座，其具有一本体以及一通道穿设于所述本体并用于供所述试管穿置；其中所述通道具有一大径段，以及一位于所述大径段下方的小径段。由此，可确保PCR进行中混合液表面温度低于引物退火反应所需温度。

Description

热对流聚合酶连锁反应装置 技术领域
本发明与聚合酶连锁反应有关，特别是指一种热对流聚合酶连锁反应装置。 现有技术
在生技领域中，聚合酶连锁反应 (PCR)已成为一项扩增特定核酸序列的成熟 技术， 此反应过程需要三个主要步骤： "变性反应"、 "引子黏合反应"及 "延展 反应"，其各需不同的反应温度。变性反应的温度典型地为介于 90至 97°C之间； 引子黏合反应的温度为依据所用引子的解链温度而选择，典型地为介于 35至 65 °C之间； 延展反应的典型温度为 72°C。
热对流聚合酶连锁反应方法主要是将装有 PCR混合液的试管底部埋置于热 水中， 试管的其余部分则暴露在室温空气中而可散热， 使 PCR混合液底部至表 面呈渐减的温度梯度如 9，。C至 35 °C， 以诱发热对流， 使 PCR混合液经历不同 的温度区段而进行不同的反应步骤。
然而， 在习知的反应装置中， 由于试管的其余部分暴露在室温空气中散热， 散热速度不够快， 随着加热时间渐长 PCR混合液表面的温度越高， 常常反应未 及完成， PCR混合液表面的温度便已高于引子黏合反应所需的温度， 使 PCR反 应无法继续进行， 而无法得到足量的特定核酸序列。 发明内容
本发明的主要目的是提供一种热对流聚合酶连锁反应装置， 其可确保 PCR 反应进行中混合液表面的温度低于弓 I子黏合反应所需温度。
为达到上述目的， 本发明所提供的一种热对流聚合酶连锁反应装置， 用于 承置一试管， 使聚合酶连锁反应在所述试管内进行， 其特征在于所述装置包含 有： 一散热座， 具有一本体， 以及一通道穿设于所述本体并用于供所述试管穿 置； 其中所述通道具有一大径段， 以及一位于所述大径段下方的小径段。
上述本发明的技术方案中， 还包含有一试管架设置于所述散热座上方， 且 所述试管架具有一供所述试管插置的容置孔。
所述试管架的容置孔由上而下依序具有一大径部、一肩部， 以及一小径部， 其中所述肩部供所述试管的一阶部抵靠。 所述试管架底端插置在所述散热座的通道中。
所述通道的小径段内径由上往下逐渐缩小。
其中所述散热座由金属制成。
本发明的优点： 采用上述技术方案， 本发明可确保 PCR反应进行中混合液 表面的温度低于引子黏合反应所需温度。
本发明适用于生技领域中的热对流聚合酶连锁反应。 附图说明
图 1 是本发明一较佳实施例的示意图
图 2 是本发明一较佳实施例的立体分解图 本发明最佳实施方式
现举以下实施例并结合附图对本发明的结构及功效进行详细说明。
如图 1所示， 本发明的较佳实施例所提供的热对流聚合酶连锁反应装置 10 主要包含有一散热座 20以及一试管架 30。 散热座 20具有一本体 22， 以及一通 道 24穿设于本体 22并用于供试管 12穿置，其中通道 24具有一大径段 241， 以 及一位于大径段 241下方的小径段 242。 由于试管 12的中段 123至上段 122为 设置于散热座 20的通道 24中， 且散热座 20是以高热传导系数的材质如铝合金 所制成， 因此 PCR混合液的热能可以通过周遭空气传导至散热座 20而更有效 率地逸散， 使混合液表面的温度在整个反应进行过程中均维持在引子黏合温度 以下， 例如 10〜55°C， 避免因混合液表面温度过高而使 PCR反应中断。 再者， 位于大径段 241的混合液散热较慢， 位于小径段 242的混合液散热较快， 经过 实验测试后发现， 此种散热座 20结构的散热效果最有利于 PCR反应的进行。
为使试管 12稳固地插置于散热座 20，可在散热座 20上方设置一试管架 30， 试管架 30具有一用于供试管 12插置的容置孔 32，使试管 12可稳固地定位，避 免试管 12外壁与散热座 20内缘接触而使混合液温度太快降低。
实际应用时， 试管架 30的容置孔 32可设计成由上而下依序设有一大径部 321、一肩部 322，以及一小径部 323，其中肩部 322用于供试管 12的一阶部 121 抵靠， 由此， 试管 12能够稳固地定位。 散热座 20下方设有一支撑座 40， 并利 用一热源 （图中未示） 对试管底部 124加热， 使底部 124内的 PCR混合液温度 维持在 90至 97°C。 再者， 试管架 30底端 301插置在散热座 20的通道 24中， 使试管架 30稳 固地设置于散热座 20上方。
此外， 在本实施例中， 通道 24的小径段 242的内径为由上往下逐渐缩小， 经过多次实验与不断改良， 发现以此结构散热时 PCR反应效率最高。 实验分别 在 10至 40°C间的 7个不同环境温度下进行 PCR反应，将试管底部 124内的 PCR 混合液温度加热到 93至 97°C， 散热座 20的温度经过测量为 36至 53 °C， 混合 液表面的温度则在 36〜53 °C之间， 使 PCR反应顺利进行。
在不违背本发明精神的前提下， 此装置的结构也可加以变化， 例如： 散热 座 20可设有多个通道 24供多个试管 12同时进行反应 （如图 2所示）。 凡此等 易于思及的结构变化， 均应被本发明的专利保护范围所涵盖。 工业应用性
本发明提供的热对流聚合酶连锁反应装置适用于生技领域中的热对流聚合 酶连锁反应， 可确保 PCR反应进行中混合液表面的温度低于引子黏合反应所需 温度。

Claims

权利要求
1、 一种热对流聚合酶连锁反应装置， 用于承置一试管， 使聚合酶连锁反应 在所述试管内进行， 其特征在于所述装置包含有：
一散热座， 具有一本体， 以及一通道穿设于所述本体并用于供所述试管穿 置； 其中所述通道具有一大径段， 以及一位于所述大径段下方的小径段。
2、 如权利要求 1所述的热对流聚合酶连锁反应装置， 其特征在于： 还包含 有一试管架设置于所述散热座上方， 且所述试管架具有一供所述试管插置的容 置孔。
3、 如权利要求 2所述的热对流聚合酶连锁反应装置， 其特征在于： 所述试 管架的容置孔由上而下依序具有一大径部、 一肩部， 以及一小径部， 其中所述 肩部供所述试管的一阶部抵靠。
4、 如权利要求 2所述的热对流聚合酶连锁反应装置， 其特征在于： 所述试 管架底端插置在所述散热座的通道中。
5、 如权利要求 1至 4中任一项所述的热对流聚合酶连锁反应装置， 其特征 在于： 所述通道的小径段内径由上往下逐渐缩小。
6、 如权利要求 1所述的热对流聚合酶连锁反应装置， 其特征在于： 其中所 述散热座由金属制成。
PCT/CN2011/001941 2011-11-22 2011-11-22 热对流聚合酶连锁反应装置 WO2013075263A1 (zh)

Priority Applications (6)

Application Number Priority Date Filing Date Title
KR1020147002887A KR101691466B1 (ko) 2011-11-22 2011-11-22 열대류 중합효소 연쇄반응 장치
CN201180072273.1A CN103649301B (zh) 2011-11-22 2011-11-22 热对流聚合酶连锁反应装置
CA2856345A CA2856345C (en) 2011-11-22 2011-11-22 Device for thermal convection polymerase chain reaction
IN3868CHN2014 IN2014CN03868A (zh) 2011-11-22 2011-11-22
PCT/CN2011/001941 WO2013075263A1 (zh) 2011-11-22 2011-11-22 热对流聚合酶连锁反应装置
EP11876152.7A EP2784150B1 (en) 2011-11-22 2011-11-22 Device for polymerase chain reaction via thermal convection

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PCT/CN2011/001941 WO2013075263A1 (zh) 2011-11-22 2011-11-22 热对流聚合酶连锁反应装置

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CN109554295A (zh) * 2019-01-21 2019-04-02 武汉理工大学 远洋船员的pcr扩增与疾病检测装置

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CN105670924A (zh) * 2016-03-01 2016-06-15 上海理工大学 自然对流聚合酶链反应微系统
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CN108410688A (zh) * 2017-02-09 2018-08-17 克雷多生物医学私人有限公司 一种热对流式聚合酶链式反应的装置
CN109401948A (zh) * 2018-11-01 2019-03-01 福建省博凯科技有限公司 一段式热对流pcr仪
CN109294901A (zh) * 2018-11-01 2019-02-01 福建省博凯科技有限公司 一段式热对流pcr仪及控制方法

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CN101735944A (zh) * 2008-11-18 2010-06-16 霍夫曼-拉罗奇有限公司 毛细管

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CN108949545A (zh) * 2018-08-16 2018-12-07 上海海洋大学 一种新型核酸等温扩增反应组件
CN109554295A (zh) * 2019-01-21 2019-04-02 武汉理工大学 远洋船员的pcr扩增与疾病检测装置

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