CN1056124A - High efficient ultrosonic wave genetic conduction method - Google Patents
High efficient ultrosonic wave genetic conduction method Download PDFInfo
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- CN1056124A CN1056124A CN 91103622 CN91103622A CN1056124A CN 1056124 A CN1056124 A CN 1056124A CN 91103622 CN91103622 CN 91103622 CN 91103622 A CN91103622 A CN 91103622A CN 1056124 A CN1056124 A CN 1056124A
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Abstract
The present invention relates to new method of gene introduction, i.e. high efficient ultrosonic wave genetic conduction method in a kind of animal-plant gene engineering.Present method is that ultrasonic wave is acted on the biomaterial in the dna solution, because ultrasonic wave makes cytolemma produce local and temporary transient structural modification, the dna molecular in the solution can diffuse into cell by this.Ultrasonic wave induced gene transfer method is not limited by host range, and foreign gene directly can be imported the vegetable cell of being with wall, particularly in the tissue block of plant materials, cultivate much easier by their regeneration plants ratio from protoplastis, it is simple that the supersonic method quiding gene also has a device, advantage such as easy to operate has wide practical use in the animal-plant gene engineering.
Description
The present invention relates to the method for gene introduction in the genetically engineered, belong to the animals and plants genetic engineering.
One of gordian technique of plant genetic engineering is how efficiently with exogenous gene transfered plant cell, and the transfer-gen plant of regenerating, and is used for improvement of crop cultivar, to cultivate high yield, high-quality, disease-resistant new variety.At present, the transgenosis of vegetable cell has biological method, chemical process and physical method.Relatively Chang Yong physical method is that electricity swashs gene transfer method.Operating process is to fill cell and DNA(thymus nucleic acid) the special small vessels of liquid places between the positive and negative electrode of electric impulser, applies electricimpulse then.Under the effect of high electric field pulse, cytolemma forms small hole because of provisional breaking taken place, and before it was repaired automatically, the dna molecular membership in the solution helped concentration difference to be diffused in the cell by hole, and finally enter nucleus inside, be incorporated in the karyomit(e) of recipient cell.The advantage of this method is easy and simple to handle, and gene transfering efficiency is higher and do not have host range restriction etc., and therefore, electrization is a gene transfer method commonly used in the animal-plant gene engineering.Shortcoming is that acceptor generally is a protoplastis when carrying out the plant gene transfer, and protoplastis is cultivated the relatively difficulty of plant of regenerating, special all the more so for the cereal crop that economic implications is arranged, therefore the sharp gene transfer method of electricity is restricted in plant gene shifts.
The objective of the invention is to propose a kind of new physical method metastatic gene, promptly using ultrasound ripple induced gene shifts, and this method is applicable to the transgenosis of animals and plants and microorganism simultaneously.
Content of the present invention is biomaterial (plant tissue piece, plant suspension cell, zooblast, microorganism cells etc.) to be mixed with dna solution put into ultrasonic cell; Again ultrasonic wave is introduced ultrasonic cell, acted on the biomaterial in the dna solution.Ultrasonic wave is continuous or pulse mode, and the ultrasonic wave sound intensity that biomaterial is experienced is less than 10W/cm
2, frequency is less than 5MHZ, be greater than 0.01 second action time.
The principle that the ultrasonic wave induced gene shifts is when ultrasonic wave is propagated in liquid, can produce cavatition.So-called cavatition is exactly some micro-bubbles that exist in the liquid, resonance takes place under action of ultrasonic waves and supervene a series of physical phenomenons.When taking place as cavatition, can produce partial High Temperature High Pressure at the cavitation center, make cell walls produce partial perforation, also make cytolemma produce partial and temporary transient structural modification, promptly also cause a series of perforation effect, the dna molecular in the solution can take this to diffuse into cell.In many tissues of higher plant, there are some micro-bubbles in the intercellular passage, these bubbles will become the cavitation center under ultrasonication, thereby cell is on every side had an effect.Because this bubble is ubiquitous, this just makes a large amount of cells can both experience cavitation effect of ultrasonic waves.
Ultrasonic wave induced gene transfer method is not limited by host range, and foreign gene directly can be imported in the tissue block of the vegetable cell, particularly plant materials of being with wall, cultivates much easier by their regeneration plants ratio from protoplastis.Advantages such as the ultrasonic wave gene transfer method is equally applicable to transgenosis of animal and the transgenosis of microorganism, and simultaneously also to have a device simple for the supersonic method quiding gene, easy to operate have wide practical use in the animal-plant gene engineering.
Five embodiment of the present invention are:
1. be 0.5W/cm with the sound intensity
2And 1.0W/cm
2, frequency is the pulse ultrasonic wave of 800KHZ, be 30 minutes action time, realizes exogenous genes introduced into wheat children fringe evoked callus is obtained transient expression.
2. be 0.5W/cm with the sound intensity
2, frequency is the pulse ultrasonic wave of 800KHZ, realizes foreign gene is imported the tobacco leaf piece 30 minutes action time, obtains stably express.
3. be 2.0W/cm with the sound intensity
2, frequency is the continuous ultrasound ripple of 800KHZ, be 30 seconds action time, and external source fluorescent substance calcium flavin is imported in the chlamydomonas cell.
4. be 0.5W/cm with the sound intensity
2, frequency is the pulse ultrasonic wave of 800KHZ, be 15 seconds action time, and external source fluorescent substance calcium flavin is imported in the red corpuscle.
5. be 2.0W/cm with the sound intensity
2, frequency is the pulse ultrasonic wave of 800KHZ, be 10 minutes action time, and external source fluorescent substance calcium flavin is imported in the yeast cell.
Claims (6)
1, a kind of high efficient ultrosonic wave genetic conduction method is characterized in that the step that gene imports is:
(1) (plant tissue piece, plant suspension cell, zooblast or microorganism cells etc. mix with dna solution with biomaterial;
(2) introduce ultrasonic wave, act on the biomaterial in the dna solution.
2, method of gene introduction as claimed in claim 1 is characterized in that the ultrasonic wave introduced for continuously or pulse ultrasonic wave, acts on the ultrasonic wave sound intensity on the biomaterial less than 10W/cm
2, frequency is less than 5MHZ, and action time was greater than 0.01 second.
3, method of gene introduction as claimed in claim 1 is characterized in that described biomaterial is the wheat children tassel evoked callus, and the ultrasonic wave of introducing is a pulse ultrasonic wave, and the ultrasonic wave sound intensity that acts on the chlamydomonas cell is 1.0W/cm
2, frequency is 800KHZ, be 30 minutes action time.
4, method of gene introduction as claimed in claim 1 is characterized in that described biomaterial is the chlamydomonas cell, and the ultrasonic wave of introducing is the continuous ultrasound ripple, and the ultrasonic wave sound intensity that acts on the chlamydomonas cell is 2.0W/cm
2, frequency is 800KHZ, be 30 seconds action time.
5, method of gene introduction as claimed in claim 1 is characterized in that described biomaterial is a red corpuscle, and the ultrasonic wave of introducing is a pulse ultrasonic wave, and the ultrasonic wave sound intensity that acts on the red corpuscle is 0.5W/cm
2, frequency is 800KHZ, be 15 seconds action time.
6, method of gene introduction as claimed in claim 1 is characterized in that described biomaterial is a yeast cell, and the ultrasonic wave of introducing is a pulse ultrasonic wave, and the ultrasonic wave sound intensity that acts on the yeast cell is 2.0W/cm
2, frequency is 800KHZ, be 10 minutes action time.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN 91103622 CN1056124A (en) | 1991-06-05 | 1991-06-05 | High efficient ultrosonic wave genetic conduction method |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN 91103622 CN1056124A (en) | 1991-06-05 | 1991-06-05 | High efficient ultrosonic wave genetic conduction method |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN1056124A true CN1056124A (en) | 1991-11-13 |
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Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN 91103622 Pending CN1056124A (en) | 1991-06-05 | 1991-06-05 | High efficient ultrosonic wave genetic conduction method |
Country Status (1)
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| CN (1) | CN1056124A (en) |
Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US7211248B2 (en) | 2001-07-10 | 2007-05-01 | Sonogene, L.L.C. | Enhancement of transfection of DNA into the liver |
| CN102169108A (en) * | 2011-01-12 | 2011-08-31 | 四川大学 | Partially adjustable supersonic wave biological effect experiment loading device and supersonic wave loading method |
| US9101745B2 (en) | 2013-03-14 | 2015-08-11 | Sonogene Llc | Sonochemical induction of ABCA1 expression and compositions therefor |
| CN106868049A (en) * | 2016-12-29 | 2017-06-20 | 天津大学 | A kind of gatherer and introduction method |
-
1991
- 1991-06-05 CN CN 91103622 patent/CN1056124A/en active Pending
Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US7211248B2 (en) | 2001-07-10 | 2007-05-01 | Sonogene, L.L.C. | Enhancement of transfection of DNA into the liver |
| CN102169108A (en) * | 2011-01-12 | 2011-08-31 | 四川大学 | Partially adjustable supersonic wave biological effect experiment loading device and supersonic wave loading method |
| US9101745B2 (en) | 2013-03-14 | 2015-08-11 | Sonogene Llc | Sonochemical induction of ABCA1 expression and compositions therefor |
| US9393328B2 (en) | 2013-03-14 | 2016-07-19 | Sonogene Llc | Sonochemical induction of ABCA1 expression and compositions therefor |
| CN106868049A (en) * | 2016-12-29 | 2017-06-20 | 天津大学 | A kind of gatherer and introduction method |
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