CN110066789B - Improved HPLC purification method of long-chain DNA primer - Google Patents
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Abstract
The invention discloses an improved HPLC purification method of a long-chain DNA primer, which comprises the following steps: firstly, synthesizing a DNA primer on a full-automatic DNA synthesizer, and automatically removing a DMTr protecting group on a first basic group 5' -OH group on a monomer; secondly, injecting an ion exchange high performance liquid chromatography system for coarse detection; thirdly, removing impurity short-chain DNA; fourthly, removing a phosphoramidite monomer; fifthly, further purifying; and sixthly, detecting the purity. The invention firstly carries out HPLC purification on DNA with DMT to remove short fragments, then carries out DMT removal step, and then further purifies, the DNA with DMT has very weak polarity and very strong hydrophobicity, the short fragments without DMT can be efficiently separated by the HPLC purification, then 10% acetic acid is added for treatment after the target molecular weight with DMT is drained, the DMT is completely removed, and the treated primer is continuously carried out HPLC purification, thus obtaining the DNA primer with high quality, wherein the purity reaches more than 99%.
Description
Technical Field
The invention belongs to the technical field of DNA primer purification, and particularly relates to an improved HPLC purification method of a long-chain DNA primer.
Background
Currently, primer synthesis is mainly performed by a solid-phase phosphoramidite triester method. The DNA fragment synthesized by the solid phase phosphoramidite triester method has the characteristics of high efficiency, quick coupling and relatively stable initial reactant. The method is to complete the synthesis of a DNA chain on a solid phase carrier, and the chemical synthesis of DNA is different from the enzymatic DNA synthesis process, wherein the DNA synthesis process extends from the 5 → 3 direction, and the DNA synthesis process starts from the 3 'end, and adjacent nucleotides are connected by 3' → 5 phosphodiester bonds.
There are many DNA synthesizers based on this method, and a high-throughput DNA automatic synthesizer manufactured by ABI/PE company is widely used. The principle of primer synthesis performed by each synthesizer is basically the same, and the main differences are high and low synthesis yield, reagent consumption, single cycle time and the like. In addition, ammonium salts generated during the short fragments and deprotection groups generated during the DNA synthesis reaction require that the primer molecules be cleaved from the CPG after the synthesis is completed, and then further purified. Based on the principles and procedures of the above synthesis, several purification methods such as C18 column, OPC or HAP, PAGE, HPLC are now common.
High Performance Liquid chromatography (hplc) is a high Performance separation method based on instrumental methods that separate products based on the hydrophobicity of DNA, unlike classical Liquid chromatography. When the method is used for separation and purification, most of N-short fragments can be effectively removed due to a series of advantages of a high-performance chromatographic column, a high-precision infusion pump, a high-sensitivity detector and the like, so that failed sequences or unbound markers can be removed, and the DNA fragments can be purified. Meanwhile, the qualitative analysis is carried out on the DNA by matching with mass spectrum, so as to ensure the correctness of the recovered fragment.
Because the purity of HPLC products is very high, the analysis time is short, can be widely applied to the fields of medicine, environmental protection, food industry, life science and the like, and the HPLC is a good purification method for the purification of unmodified oligonucleotide primers and modified primers with less than 40 basic groups if the requirement on the purity of products is very high.
Disclosure of Invention
The invention aims to provide an improved method for HPLC purification of long-chain DNA primers, which is characterized in that DMT-bearing DNA is firstly subjected to HPLC purification to remove short fragments, then subjected to DMT removal step and further purified, the DMT-bearing DNA has weak polarity and strong hydrophobicity and can be well adsorbed on a C18 column, and the short fragments without DMT can be efficiently separated through HPLC purification. Then, the target molecular weight of DMT is pumped to dryness, 10% acetic acid is added for treatment for 20-30min, DMT is completely removed, and the treated primer is continuously subjected to HPLC purification, so that the high-quality DNA primer can be obtained, and the purity is more than 99%.
The purpose of the invention can be realized by the following technical scheme:
an improved method for HPLC purification of long-chain DNA primers, comprising the steps of:
firstly, synthesizing a DNA primer on a full-automatic DNA synthesizer, and automatically removing a 4,4 '-dimethoxytrityl protecting group on a first base 5' -OH group on a monomer to obtain a crude product of DNA synthesis;
secondly, injecting the DNA synthesis crude product obtained in the last step into an ion exchange high performance liquid chromatography system with extremely strong separation performance, carrying out crude detection, and confirming the position of a main peak and the content of target DNA;
thirdly, increasing the injection amount, recovering the head part of the main peak flat head, and removing impurity short-chain DNA to obtain a DNA primer with a phosphoramidite monomer;
fourthly, after the DNA primer with the phosphoramidite monomer is drained, adding acetic acid aqueous solution to treat for 20-30min, and removing the phosphoramidite monomer;
fifthly, injecting the DNA primer without the phosphoramidite monomer in the last step into an ion exchange high performance liquid chromatography system again for further purification treatment;
and sixthly, detecting the purity of the DNA recovered in the last step, and confirming that the purity reaches more than 99 percent to finish purification.
Further, the amount of purification of the DNA primer at a time was 1 OD.
Further, the mass fraction of the acetic acid aqueous solution in the fourth step is 10%, and the mass ratio of the DNA primer after being dried and the added acetic acid aqueous solution is 1: 12-14.
Furthermore, the vibration processor in the fourth step comprises a reaction frame and vibration devices arranged on the reaction frame, wherein the two vibration devices are symmetrically arranged on two sides of the reaction frame;
the two end side surfaces of the reaction frame are provided with mounting cylinders;
the vibrating device comprises a supporting bottom plate, a sliding groove is formed in the surface of the supporting bottom plate, a sliding block is installed in the sliding groove, and the sliding block is in sliding fit with the sliding groove; a second mounting through hole is formed in the surface of the sliding block;
a supporting vertical plate is vertically fixed at the edge position of the surface of the supporting bottom plate, a first mounting through hole is formed in the surface of the supporting vertical plate, a first shaft lever is mounted in the first mounting through hole in a penetrating manner through a bearing, a first disc is fixed on the first shaft lever, a second disc and a third disc which are sequentially connected with each other are arranged on the first disc, and the first disc is connected with the second disc through a connecting rod, and the second disc is connected with the third disc through a connecting rod;
a second shaft lever is fixed at the center of the surface of the third disc and movably arranged in a second circular through hole on the surface of the sliding block;
the side surface vertical fixation of sliding block has the connecting rod, and the connecting rod cooperates with the installation section of thick bamboo, and the reaction frame is installed on the connecting rod through the installation section of thick bamboo.
Further, the supporting vertical plates are arranged in parallel to the sliding grooves.
Furthermore, fixed shafts are uniformly distributed on the surfaces of the first disc, the second disc and the third disc along the circumferential direction and are movably connected with the connecting rod.
The invention has the beneficial effects that:
the invention aims at the improvement of a long-chain DNA primer purification method, and mainly aims at carrying out HPLC purification on DMT-bearing DNA to remove short fragments, then carrying out DMT removal step and further purifying. HPLC purification is based on the different size fragment with payload separation of products; the different length of DNA fragments in the crude product of the synthesis determines that it has different net charges, and longer fragments with high charges flow slower than the shorter fragments with low charges in the ion exchange column, because DMT-bearing DNA is very polar and hydrophobic and adsorbs well on C18 column, and the short fragments without DMT can be separated efficiently by HPLC purification. Then, after the target molecular weight of DMT is pumped, 10% acetic acid aqueous solution is added into a specially-made vibration processor for vibration treatment for 20-30min, the DNA primers and the acetic acid aqueous solution which are pumped are placed in a reaction frame, and the reaction frame moves back and forth, so that the DNA primers and the acetic acid aqueous solution are more fully contacted, the reaction is more complete, the DMT is completely removed in a short time, the realization mode is automatic, the working efficiency can be effectively improved, and the manpower can be saved; the primers after the oscillation treatment are continuously purified by HPLC, so that high-quality DNA primers can be obtained, and the purity reaches more than 99%.
Drawings
In order to facilitate understanding for those skilled in the art, the present invention will be further described with reference to the accompanying drawings.
FIG. 1 is a schematic diagram of a vibration processor according to the present invention;
FIG. 2 is a schematic diagram of the structure of the reaction frame of the vibration processor of the present invention;
fig. 3 is a schematic structural diagram of a vibration device of the vibration processor of the present invention.
Detailed Description
The technical solutions of the present invention will be described clearly and completely with reference to the following embodiments, and it should be understood that the described embodiments are only a part of the embodiments of the present invention, and not all of the embodiments. All other embodiments, which can be derived by a person skilled in the art from the embodiments given herein without making any creative effort, shall fall within the protection scope of the present invention.
An improved method for HPLC purification of long-chain DNA primers, comprising the steps of:
firstly, synthesizing a DNA primer on a full-automatic DNA synthesizer, and automatically removing a DMTr (4,4 '-dimethoxytrityl) protecting group on a first base 5' -OH group on a monomer to obtain a crude product of DNA synthesis;
secondly, injecting the DNA synthesis crude product obtained in the last step into an ion exchange high performance liquid chromatography system with extremely strong separation performance, carrying out crude detection, and confirming the position of a main peak and the content of target DNA;
thirdly, increasing the injection amount, and recovering the head part of the main peak flat head, so that the aim of removing impurity short-chain DNA can be fulfilled, and the DNA primer with DMT (phosphoramidite monomer) is obtained; preferably, in order to ensure the purity of the DNA primer, the purification amount at one time is generally 1 OD;
fourthly, after the DNA primer with DMT is drained, the DNA primer is placed in a reaction frame 1 of a vibration processor, 10 percent (mass fraction) of acetic acid aqueous solution is added into the DNA primer, and the DNA primer is vibrated in the vibration processor for 20 to 30min to achieve the aim of removing the DMT;
wherein the mass ratio of the DNA primer after being dried and the added acetic acid aqueous solution is 1: 12-14;
fifthly, injecting the DNA primer subjected to DMT removal in the last step into an ion exchange high performance liquid chromatography system for further purification treatment;
and sixthly, detecting the purity of the recovered DNA, and confirming that the purity reaches more than 99 percent to finish purification.
Referring to fig. 1-3, the vibration processor in the fourth step, as shown in fig. 1, includes a reaction frame 1 and vibration devices 2 mounted on the reaction frame 1, wherein the two vibration devices 2 are symmetrically mounted on two sides of the reaction frame 1;
as shown in fig. 2, the reaction frame 1 is used for containing the DNA primer after being dried and 10% acetic acid aqueous solution, the two are oscillated in the reaction frame 1, and the two end side surfaces of the reaction frame 1 are provided with mounting cylinders 101;
as shown in fig. 3, the vibration device 2 includes a supporting base plate 201, a sliding groove 202 is formed on the surface of the supporting base plate 201, a sliding block 203 is installed in the sliding groove 202, and the sliding block 203 is in sliding fit with the sliding groove 202; the surface of the sliding block 203 is provided with a second mounting through hole;
a supporting vertical plate 204 is vertically fixed at the edge position of the surface of the supporting base plate 201, the supporting vertical plate 204 is arranged in parallel to the sliding groove 202, a first mounting through hole is formed in the surface of the supporting vertical plate 204, a first shaft lever 205 is mounted in the first mounting through hole in a penetrating manner through a bearing, one end of the first shaft lever 205, which extends out of the supporting vertical plate 204, is connected with a motor, and the motor drives the first shaft lever 205 to rotate; a first disc 206 is fixed on the first shaft rod 205, a second disc 207 and a third disc 208 which are connected with each other in sequence are arranged on the first disc 206, and the first disc 206 and the second disc 207, the second disc 207 and the third disc 208 are connected through a connecting rod 209;
specifically, fixed shafts are uniformly distributed on the surfaces of the first disc 206, the second disc 207 and the third disc 208 along the circumferential direction, and the fixed shafts are movably connected with a connecting rod 209;
a second shaft rod 210 is fixed at the center of the surface of the third disc 208, and the second shaft rod 210 is movably arranged in a second circular through hole on the surface of the sliding block 203;
it should be noted that, a connecting rod 211 is vertically fixed on the side surface of the sliding block 203, the connecting rod 211 is matched with the mounting cylinder 101, and the reaction frame 1 is mounted on the connecting rod 211 through the mounting cylinder 101;
the working principle and the mode of the vibration device are as follows:
after a motor connected with the first shaft lever 205 is started, the first shaft lever 205 drives the first disc 206 to rotate, in the rotating process, the second disc 207 rotates around the first disc 206, the third disc 208 rotates around the second disc 207, and finally the sliding block 203 moves back and forth along the sliding groove due to the limiting effect of the sliding groove on the sliding block 203, so that the reaction frame 1 is driven to move back and forth;
the DNA primer and the acetic acid aqueous solution which are drained are placed in the reaction frame 1, and the reaction frame 1 moves back and forth, so that the DNA primer and the acetic acid aqueous solution are contacted more fully, the reaction is more complete, the DMT is completely removed in a short time, the realization mode is automatic, the working efficiency can be effectively improved, and the labor can be saved;
example 1: using general CPG, removing DMTr protecting group from long-chain PHO modified primer (73mer, 5' PHO) on a DNA synthesizer, removing short segment by HPLC purification method, pumping out target molecular weight with DMT, adding 10% acetic acid for 20 min, completely removing DMT, purifying by HPLC, inspecting, detecting with high purity, basically no impurity peak in electrophoresis spectrogram, and purity up to 99.5%.
Example 2
Using general CPG, removing DMTr protecting group from long-chain PHO modified primer (73mer, 5' PHO) on a DNA synthesizer, removing short segment by HPLC purification method, pumping out target molecular weight with DMT, adding 10% acetic acid for 25 min, completely removing DMT, purifying by HPLC, inspecting, detecting with high purity, basically no impurity peak in electrophoresis spectrogram, and purity up to 99.8%.
Example 3
Using general CPG, removing DMTr protecting group from long-chain PHO modified primer (73mer, 5' PHO) on a DNA synthesizer, removing short segment by HPLC purification method, pumping out target molecular weight with DMT, adding 10% acetic acid for 30min to completely remove DMT, purifying by HPLC again, inspecting, detecting with high purity, basically no impurity peak in electrophoresis spectrogram, and purity up to 99.6%.
Comparative example 1
Using general CPG, the DMTr protecting group was removed from the long-chain PHO modified primer (73mer, 5' PHO) on a DNA synthesizer, the short fragment was removed by HPLC purification, and then 10% acetic acid was added for 30 minutes after draining the target molecular weight with DMT, as shown in fig. 2, the electrophoretogram showed more miscellaneous peaks with a purity of 73.49%.
Comparative example 2
Using general CPG, removing DMTr protecting group from long-chain PHO modified primer (73mer, 5' PHO) on DNA synthesizer, purifying by HPLC method, and obtaining purity 70.25% with more miscellaneous peaks in the electrophoretogram.
The preferred embodiments of the invention disclosed above are intended to be illustrative only. The preferred embodiments are not intended to be exhaustive or to limit the invention to the precise embodiments disclosed. Obviously, many modifications and variations are possible in light of the above teaching. The embodiments were chosen and described in order to best explain the principles of the invention and the practical application, to thereby enable others skilled in the art to best utilize the invention. The invention is limited only by the claims and their full scope and equivalents.
Claims (3)
1. The improved HPLC purification method of the long-chain DNA primer is characterized in that a vibration processor is adopted to carry out a vibration treatment purification process on a mixture of the long-chain DNA primer and an acetic acid aqueous solution, the vibration processor comprises a reaction frame (1) and vibration devices (2) arranged on the reaction frame (1), and the two vibration devices (2) are symmetrically arranged on two sides of the reaction frame (1);
the two end side surfaces of the reaction frame (1) are provided with mounting cylinders (101);
the vibration device (2) comprises a supporting bottom plate (201), a sliding groove (202) is formed in the surface of the supporting bottom plate (201), a sliding block (203) is installed in the sliding groove (202), and the sliding block (203) is in sliding fit with the sliding groove (202); a second mounting through hole is formed in the surface of the sliding block (203);
a support vertical plate (204) is vertically fixed at the edge position of the surface of the support base plate (201), a first installation through hole is formed in the surface of the support vertical plate (204), a first shaft lever (205) is installed in the first installation through hole in a penetrating manner through a bearing, a first disc (206) is fixed on the first shaft lever (205), a second disc (207) and a third disc (208) which are sequentially connected with each other are arranged on the first disc (206), and the first disc (206) and the second disc (207) as well as the second disc (207) and the third disc (208) are connected through connecting rods (209);
a second shaft lever (210) is fixed at the center of the surface of the third disc (208), and the second shaft lever (210) is movably arranged in a second circular through hole on the surface of the sliding block (203);
the side surface of the sliding block (203) is vertically fixed with a connecting rod (211), the connecting rod (211) is matched with the installation barrel (101), and the reaction frame (1) is installed on the connecting rod (211) through the installation barrel (101).
2. The method for improving HPLC purification of a long-chain DNA primer according to claim 1, wherein said support riser (204) is disposed parallel to the chute (202).
3. The improved HPLC purification method for long-chain DNA primers of claim 1, wherein the surfaces of the first disc (206), the second disc (207) and the third disc (208) are all provided with fixed shafts along the circumferential direction, and the fixed shafts are movably connected with the connecting rod (209).
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Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1155887A (en) * | 1994-07-05 | 1997-07-30 | 海布里顿公司 | Purficiation of oligodeoxynucleotide phosphorothioates using DEAE-5PW anion ion-exchange chromatography and hydrophobic interaction chromatography |
| CN1514842A (en) * | 2001-06-07 | 2004-07-21 | ISISҩ�﹫˾ | Processes of purifying oligonucleotides |
| CN1678618A (en) * | 2002-08-28 | 2005-10-05 | Quia技术股份有限公司 | Process for separating and deprotecting oligonucleotides |
| CN106632558A (en) * | 2016-08-25 | 2017-05-10 | 淮阴师范学院 | Method for purifying oligonucleotide and application |
| CN109641928A (en) * | 2016-06-14 | 2019-04-16 | 比奥根Ma公司 | Hydrophobic interaction chromatography for oligonucleotides purifying |
-
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Patent Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1155887A (en) * | 1994-07-05 | 1997-07-30 | 海布里顿公司 | Purficiation of oligodeoxynucleotide phosphorothioates using DEAE-5PW anion ion-exchange chromatography and hydrophobic interaction chromatography |
| CN1514842A (en) * | 2001-06-07 | 2004-07-21 | ISISҩ�﹫˾ | Processes of purifying oligonucleotides |
| CN1678618A (en) * | 2002-08-28 | 2005-10-05 | Quia技术股份有限公司 | Process for separating and deprotecting oligonucleotides |
| CN109641928A (en) * | 2016-06-14 | 2019-04-16 | 比奥根Ma公司 | Hydrophobic interaction chromatography for oligonucleotides purifying |
| CN106632558A (en) * | 2016-08-25 | 2017-05-10 | 淮阴师范学院 | Method for purifying oligonucleotide and application |
Non-Patent Citations (1)
| Title |
|---|
| Analysis and Purification of Synthetic Oligonucleotides by Reversed-Phase High-Performance Liquid Chromatography with Photodiode Array and Mass Spectrometry Detection;Martin Gilar;《Analytical Biochemistry》;20011231;全文 * |
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