JPH0361852A - Gel-electrophoresis analyzing system - Google Patents

Abstract

PURPOSE:To make it possible to use a general-purpose electron multiplier tube and to improve profitability by winding a motor-driven endless belt having a slit around an electrophoresis plate, and detecting fluorescence emitted through the slit with a photomultiplier. CONSTITUTION:Fluorescence is generated in response to gel electrophoresis from an electrophoresis plate 3 based on the projection of laser light from a laser light source 1. The fluorescence emitted through a slit 9 of a motor- driven endless belt 5 that is wound around the plate 3 is detected with a photomultiplier 15. Therefore, only the fluorescence of a migrating lane where the slit has passed is inputted into the photomultiplier. A line sensor and highly sensitive multichannels are not required. The economical efficiency is enhanced by the use of a general-purpose electron multiplier tube.

Description

DETAILED DESCRIPTION OF THE INVENTION [Field of Industrial Application] The present invention relates to an electrophoresis system. More specifically, the present invention relates to an analysis method in a low-cost gel electrophoresis system that generates fluorescence by irradiating laser light.

[Prior Art] Gel electrophoresis is widely used as a method for determining the base sequence of DNA and the like.

Conventionally, samples were labeled with radioisotopes and analyzed during electrophoresis, but this method had the disadvantage of being laborious and time-consuming. For this reason, recently, a method of labeling the sample with a fluorescent material is being considered.

In the method using light, fluorescently labeled DNA fragments are migrated in a gel, but a light excitation part and a photodetector are provided for each migration path 15 to 20 cm below the migration start part, and the DNA passing through them is Measure the pieces in sequence. The migration speed, ie, the size of the DNA fragment, can be determined from the detection time.

Japanese Unexamined Patent Publication No. 63-21556 discloses that the line on the gel of an electrophoresis device irradiated with a laser and the array direction of the photodiode array are configured to be perpendicular to the direction of migration of DNA fragments in the electrophoresis device. A DNA base sequencing device has been disclosed.

FIG. 3 is a schematic diagram illustrating the configuration of the device.

As shown in the figure, the laser light emitted from the light source 70 is bent by a mirror 72 and irradiates the migration plate 74 from the side. When the fluorescent label DNA 76 reaches the irradiation area, it emits fluorescence. The fluorescence from each migration path is imaged by the lens 78 on the light receiving section 82 of the fluorescence detection 380. This signal is amplified, converted into an electrical signal by a photodiode array 84, and subjected to arithmetic processing to obtain an analysis result.

[Problem to be solved by the invention] In conventional devices, a licensor or a high-sensitivity multichannel photodetector (manufactured by Hamamatsu Photonics) is used as a fluorescence detector.
etc. are used. However, these detectors are generally very expensive and cause a significant increase in the manufacturing cost of the entire electrophoresis device.

Therefore, an object of the present invention is to provide a gel electrophoresis analysis method having a fluorescence detection system that is inexpensive but capable of efficiently detecting fluorescence.

[Means for Solving the Problems] In order to achieve the above object, the present invention provides a gel electrophoresis system that generates fluorescence by irradiating laser light with a motor-driven type having at least one slit at a predetermined location. A gel electrophoresis analysis method is provided in which an endless belt is wound around a migration plate at a position corresponding to the irradiation line of a laser beam, and fluorescence emitted from the slit is detected by a photomultiplier tube.

It is preferable that the endless belt has two slits and is driven by a DC motor, and that an encoder is attached to the motor.

[Function] As described above, in the system of the present invention, an inexpensive photomultiplier tube is used in place of a conventional expensive fluorescence detector, so that the manufacturing cost of the entire system can be kept low.

Furthermore, since an endless belt having slits is wrapped around the electrophoresis plate, only the fluorescence in the electrophoresis lane through which the slits pass enters the photomultiplier tube.

Therefore, even if a photomultiplier tube is used instead of a conventional expensive licenser or high-sensitivity multichannel device, highly reliable detection is possible.

[Example] Hereinafter, the gel electrophoresis analysis method of the present invention will be explained in more detail with reference to the drawings.

FIG. 1 is a conceptual diagram showing an example of the configuration of an apparatus used to carry out the gel electrophoresis analysis method of the present invention.

As shown in the figure, in the apparatus used to carry out the analysis method of the present invention, a laser beam emitted from a laser light source 1 is incident on a gel electrolyte layer 4 from right beside a migration plate 3, and a fluorescent material is irradiate the labeled specimen with

An endless belt 5 is placed on the irradiation line of the laser beam on the migration plate 3.
to confuse. The belt 5 may be in contact with the migration plate or may be in non-contact. The belt 5 has a slit 9 having a width equal to or equal to the width of the migration lane 7 at the upper end of the migration plate.
is installed. At least 1 slit 9 on the belt
Although it is sufficient to arrange a plurality of them at equal intervals, in order to improve the efficiency of analysis, it is preferable to arrange a plurality of them at equal intervals. For example, the first slit is at the edge of the electrophoresis plate (i.e., the end position of the scanning line).
Once , another slit is placed at the starting point of the scanning line, allowing continuous analysis. Although the size of the slit itself is not an essential requirement of the present invention, for example, if the width of the migration lane is 2 mm, it is preferable that the width of the slit is also 2 mm. Further, the height of the slit is preferably about 2 to 3-1.

The belt can be moved at a constant speed by a driving means 11 such as a DC motor, for example. The direction of rotation of the belt is not particularly limited. In order to match and record the position of the migration lane 7 and the position of the slit, it is preferable to attach an encoder 13 to the DC motor.

A photomultiplier tube 15 is arranged so as to be perpendicular to the scanning line of the laser beam. It is preferable to provide a condensing lens 17 on the front surface of the multiplier tube 15 in order to increase the condensing efficiency of the fluorescence emitted from the slit 9. Further, a filter 19 may be provided in front of the condenser lens to transmit only fluorescence of a specific wavelength and cut out disturbance light of other wavelengths.

The encoder 13 and photomultiplier tube 15 are each connected to a signal processing circuit 21. Photomultiplier tube 15
Since the received light signal is analog, it is necessary to pass it through an A/D converter 23 to convert it to digital.

Signal from A/D converter 23 and encoder 13
The signal is first transmitted to the interface 25 in the signal processing circuit 21, then transmitted to the CPU 29 via the bus 27 for arithmetic processing, and then stored in the memory 31 via the bus 27 again. Alternatively, the data may be temporarily stored in the memory 31, sent to the CPU 29 via the bus for arithmetic processing, and returned to the memory again. An appropriate output device 35 such as a printer and/or display device 37 such as a CRT can be connected to the CPU 29 via an interface 33.

FIG. 2 is a characteristic diagram showing an example of the output waveform. The lower row shows the position of each migration lane recorded by the encoder, and the upper row shows a peak curve showing the change in fluorescence intensity of each lane. Therefore, by simultaneously referring to the data in the lower row and the first row, it is possible to determine, for example, the base sequence of DNA or the like.

- The migration speed of a sample such as DNA within the membrane is slower than the speed at which the slit moves in the lateral direction, so even if multiple bases exist at both ends of the scanning line at the same time, the migration speed of the sample such as DNA is slower than the speed at which the slit moves in the lateral direction. Each can be detected before electrophoresis. - For example, if the moving speed of the slit is 1 second/half revolution of the electrophoresis plate, it will only take about 1 second to detect fluorescence in the electrophoresis lane from one end of the electrophoresis plate, so one base
Unless electrophoresis is performed at a speed of 20 per second or more, if the slit height is two steps, both the bases in the leftmost electrophoresis lane and the bases in the rightmost electrophoresis lane can be detected even if they are on the same scanning line.

The gel electrophoresis analysis method of the present invention can be used for the analysis of: 1) not only NA but any target as long as it can be labeled with a fluorescent substance;

[Effects of the Invention] As explained below, in the gel electrophoresis system used to carry out the analysis method of the present invention, an inexpensive photomultiplier tube is used in place of the conventional expensive fluorescence detector. Therefore, the manufacturing cost of the entire system can be kept low.

Furthermore, since an endless belt having slits is wound around the electrophoresis plate, only the fluorescence in the electrophoresis lane through which the slits pass enters the photomultiplier tube.

Therefore, even if a photomultiplier tube is used instead of a conventional expensive licenser or high-sensitivity multichannel, highly reliable detection within -1·min is possible.

[Brief explanation of drawings]

FIG. 1 is a conceptual diagram showing an example of the configuration of an apparatus used to carry out the gel electrophoresis analysis method of the present invention, and FIG. 2 shows a diagram corresponding to each electrophoresis lane obtained by the apparatus. FIG. 3 is a characteristic diagram showing a fluorescence peak waveform, and FIG. 3 is a schematic configuration diagram of a DNA base sequencing apparatus disclosed in Japanese Patent Application Laid-Open No. 63-21556. 1... Laser light source, 3... Migration plate, 5... Endless belt. 7...Migration lane, 9...Slit, 11...Motor. 13... Encoder, 15... Photomultiplier tube. 17... Condensing lens, 19... Filter. 21... Arithmetic processing circuit, 23... A/D converter. 25...Interface, 27...Bus. 29...CPU, 31...Memory.

Claims (2)

[Claims] (1) In a gel electrophoresis system that generates fluorescence by irradiating laser light, a motor-driven endless belt with at least one slit at a predetermined location is wound around the electrophoresis plate at a position corresponding to the irradiation line of the laser beam. A gel electrophoresis analysis method characterized in that the fluorescence emitted from the slit is detected by a photomultiplier tube. (2) The gel electrophoresis analysis method according to claim 1, wherein the endless belt has a plurality of slits and is driven by a DC motor, and an encoder is attached to the motor.