CN1313027A - Method for breeding wolfberry autotetraploid - Google Patents
Method for breeding wolfberry autotetraploid Download PDFInfo
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- CN1313027A CN1313027A CN 01111148 CN01111148A CN1313027A CN 1313027 A CN1313027 A CN 1313027A CN 01111148 CN01111148 CN 01111148 CN 01111148 A CN01111148 A CN 01111148A CN 1313027 A CN1313027 A CN 1313027A
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- matrimony vine
- autotetraploid
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Abstract
A method for breeding wolfberry autotetraploid includes such steps as a, cutting the disinfected excellent asexual wolfberry leaf into blocks and culturing it in the MS culture medium containing 6-penzylaminopurine and indole-3-acetic acid: b, reduplicating the cultured callus with colchicin as chromosome; c, grafting the processed callus in said MS culture medium to grow its plant and reproducing it after it is proved to be tetraploid plant by microscopic examination. The culturing conditions for steps a and c are 26-30 deg.C, 1800-2200 luxes for 10-12 hr per day, and 25-30 days for culturing it. Its advantages are high mutagenic ratio and optimized rate, and high reproduction speed.
Description
The present invention relates to a kind of breeding method of matrimony vine polyploid, especially relate to a kind of breeding method of matrimony vine autotetraploid.
In the prior art, the matrimony vine autotetraploid normally utilizes mutagen to handle matrimony vine seed or matrimony vine seedling growing point or matrimony vine stem apex as colchicine and makes to cultivate after its chromosome doubling and obtain, the shortcoming of this method is: (1) mutagenic frequency is low, as handling the matrimony vine seed with mutagen, its radicle somatic double, but its Cheng Miaohou stem apex does not have the cell of doubling through microscopy, and handling matrimony vine seedling growing point mutagenic frequency only is 1/130-200; (2) breeding cycle is long, is example with the method for handling matrimony vine seedling growing point mutagenesis polyploid, needs the time more than 2 years from sowing, mutagenesis, observation, microscopy to being defined as the matrimony vine autopolyploid; (3) preferred rate is low, owing to do not know the genetic character of mutant materials itself in advance, thereby preferred rate is low; (4), thereby be difficult to obtain pure autotetraploid strain system owing to utilize said method to induce the matrimony vine autotetraploid chimeric frequency height to occur.
The objective of the invention is to overcome the defective of above-mentioned prior art, a kind of mutagenic frequency height is provided, breeding cycle is short, preferred rate height, good practical effect and can obtain the breeding method of pure matrimony vine autotetraploid.
Technical scheme of the present invention is:
A kind of breeding method of matrimony vine autotetraploid is characterized in that, comprises following process:
A. inducing of good matrimony vine clone callus: after the sterilization of choiceness matrimony vine blade, be cut into piece, be seeded in the MS medium that contains 6-benzylaminopurine (6-BA) and indole-3-acetic acid (IAA) and cultivate;
B. inducing of matrimony vine autotetraploid: above-mentioned cultured callus is carried out chromosome doubling with colchicine handle;
C. matrimony vine autotetraploid plant is cultivated: the callus after the above-mentioned processing is seeded in the medium that contains 6-benzylaminopurine (6-BA) and indole-3-acetic acid (IAA) cultivates, Cheng Miaohou, microscopy, confirming as tetraploid plant expands numerous, the condition of culture of above-mentioned a process and c process is: cultivation temperature 26-30 ℃, under the illumination 1800-2200 lux (LX), shine 10-12h every day, incubation time is 25-30 days.
Disinfectant program is in the above-mentioned a process, earlier with 70% alcohol-pickled 0.5-1 minute, uses 0.1% mercury chloride (mercuric chloride Hgcl again
2) soaked 8-10 minute, use aseptic water washing at last.
The program of inducing of above-mentioned b process is: the callus that will handle cuts into fritter, puts into culture dish, and the absorbent cotton that soaks into 0.2% colchicine solution is all laid in the upper and lower of this fritter, handles 48h then under aseptic, dark condition.
6-benzylaminopurine (6-BA) consumption is 0.5-1.5mg/l in above-mentioned a and the c process, and the consumption of indole-3-acetic acid (IAA) is 0.5-2.0mg/l.
6-benzylaminopurine (6-BA) consumption is 1mg/l in the above-mentioned a process, and the consumption of indole-3-acetic acid (IAA) is 0.5mg/l.
6-benzylaminopurine (6-BA) consumption is 0.5mg/l in the above-mentioned c process, and the consumption of indole-3-acetic acid (IAA) is 1mg/l.
Above-mentioned MS medium is used for general tissue culture.
The breeding method of matrimony vine autotetraploid of the present invention, because before carrying out chromosome mutagenesis and doubling, at first carry out the cultivation of high-quality callus, thereby improved induced mutation rate, evidence callus type difference, there is utmost point significant difference in its mutagenic frequency, only be 2.3% as white loose type callus mutagenic frequency, and the callus induced mutation rate of light green color, hard, the surperficial redgreen bud point of quality is up to 97.5%; The mutagenic condition of the autotetraploid that the present technique means are provided, make callus extent of damage after colchicine is handled reduce to minimum, evidence, the callus fritter is in same concentration, under the same processing time condition, if be immersed in the colchicine soup, it is few to handle back differentiation bud, the monoblock that has does not have differentiation capability, and adopts this technology can make mutagenesis nursery stock differentiation rate improve 63.7%; Simultaneously, because microscopy and the clonal foundation of autotetraploid are carried out synchronously, thereby can provide purebred autotetraploid, shortening the breeding cycle, as in the seed selection of no seed matrimony vine new varieties 99-3, autotetraploid 98-2 as one of parent is defined as pure autotetraploid in February, 98, and March, rooting and transplant was promptly bloomed, the result then mid-July.
Embodiment:
Adopt the clonal blade of good matrimony vine to wash 3-4h, sterilization: 70% alcohol-pickled 1min → 0.1% mercuric chloride (Hgcl with running water
2) soak 10min → usefulness aseptic water washing 3-5 time, blade is vertically cut, cross-section again one-tenth fritter is seeded in MS+6-BAlmgL
-1+ IAA0.5mgL
-1In the medium, be placed in the culturing room and cultivate.Condition of culture is: 28 ℃ of temperature, under the illumination 2000LX, illumination every day 12h cultivated 30 days, can form a large amount of callus.
The MS medium is used for general tissue culture, fills a prescription to be: unit is mg/l
| NH 4NO 3(ammonium nitrate) | ?????1650 | KNO 3(potassium nitrate) | ?????1900 | KH 2PO 4(potassium dihydrogen phosphate) | 170 |
| MgSO 4.7H 2O (magnesium sulfate) | ??????370 | CaCL 2.2H 2O (calcium chloride) | ??????44O | FeSO 4.7H 2O (ferrous sulfate) | 27.85 |
| Na 2-EDTA (disodium ethylene diamine tetraacetate) | ????37.25 | MnSO 4.4H 2O (manganese sulphate) | ?????22.3 | ZnSO 4.7H 2O (zinc sulphate) | 8.6 |
| H 3BO 3(boric acid) | ??????6.2 | KI (potassium iodide) | ?????0.83 | Na 2MoO 4.2H 2O (molybdic acid is received) | 0.25 |
| ?CuSO 4.5H 2O (copper sulphate) | ????0.025 | ??CoCL 2.6H 2O (cobalt chloride) | ????0.025 | Glycine | ?2 |
| Thiamine hydrochloride | ??????0.4 | Puridoxine hydrochloride | ??????0.5 | Nicotinic acid | 0.5 |
| Myo-inositol | ??????100 | Sucrose | ????30000 | Agar | 10000 |
Select light green, quality is closely hard, the callus of surface redgreen bud point, be divided into the fritter of 0.5cm * 0.5cm, put into the culture dish of crossing through autoclave sterilization, culture dish bottom shop one deck is put the callus fritter thereon with the absorbent cotton that 0.2% colchicine soaks into, and the absorbent cotton that soaks into 0.2% colchicine covers on it then.Aseptic, dark condition is used aseptic water washing 3-4 time after handling 48h down, is seeded in MS+6-BA0.5mgL
-1+ IAA0.5mgL
-1Medium in cultivate, condition of culture such as callus culture condition can induce seedling in 25-30 days.
When treating height of seedling 1.5-2cm, seedling is taken out from base portion, do chromosome examination with spire (5-6 sheet around the growing point) and growing point, the remainder individual plant is seeded in one bottle of medium, and it is numerous that tetraploid expansion is confirmed as in inspection.
Expand numerous employing MS+6-BA0.5mg/l+IAAI.5mg/l medium, take root and adopt 1/2MS+IBA (indole-3-butyric acid) 0.6mg/l medium.The culture of rootage inoculation was taken root after 7 days, began to practice transplantation of seedlings when root grows to 0.5-1cm.Transplanting is carried out in nutritive cube, and nutritive cube bottom 2/3 is nutrition soil, the vermiculite of upper end 1/3 for sterilizing with 0.2% carbendazim.Transplanting the back notes preserving moisture, sheltering from heat or light.Remove plastics bow canopy after 25 days and practice seedling, carry out the green transplantation of seedlings in land for growing field crops about 10cm when height of seedling grows to.
Claims (6)
1. the breeding method of a matrimony vine autotetraploid is characterized in that, comprises following process:
A. inducing of good matrimony vine clone callus: after the sterilization of choiceness matrimony vine blade, be cut into piece, be seeded in the MS medium that contains 6-benzylaminopurine and indole-3-acetic acid and cultivate;
B. inducing of matrimony vine autotetraploid: above-mentioned cultured callus is carried out chromosome doubling with colchicine handle;
C. matrimony vine autotetraploid plant is cultivated: the callus after the above-mentioned processing is seeded in the MS medium that contains 6-benzylaminopurine and indole-3-acetic acid cultivates, Cheng Miaohou, microscopy, confirming as tetraploid plant expands numerous, the condition of culture of above-mentioned a process and c process is: cultivation temperature 26-30 ℃, every day is shone 10-12h in illumination 1800-2200 lux, and incubation time is 25-30 days.
2. by the breeding method of the described matrimony vine autotetraploid of claim 1, it is characterized in that: disinfectant program is in the above-mentioned a process, earlier with 70% alcohol-pickled 0.5-1 minute, soaks 8-10 minute with 0.1% mercury chloride again, uses aseptic water washing at last.
3. press the breeding method of the described matrimony vine autotetraploid of claim 1, it is characterized in that: the program of inducing of above-mentioned b process is: the callus that will handle cuts into fritter, put into culture dish, the absorbent cotton that soaks into 0.2% colchicine solution is all laid in the upper and lower of this fritter, handles 48h then under aseptic, dark condition.
4. by the breeding method of claim 1,2 or 3 described matrimony vine autotetraploids, it is characterized in that: 6-benzylaminopurine consumption is 0.5-1.5mg/l in above-mentioned a and the c process, and the consumption of indole-3-acetic acid is 0.5-2.0mg/l.
5. by the breeding method of the described matrimony vine autotetraploid of claim 4, it is characterized in that: 6-benzylaminopurine consumption is 1mg/l in the above-mentioned a process, and the consumption of indole-3-acetic acid is 0.5mg/l.
6, by the breeding method of the described matrimony vine autotetraploid of claim 4, it is characterized in that: 6-benzylaminopurine consumption is 0.5mg/l in the above-mentioned c process, and the consumption of indole-3-acetic acid is 1mg/l.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN 01111148 CN1313027A (en) | 2001-04-06 | 2001-04-06 | Method for breeding wolfberry autotetraploid |
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| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN 01111148 CN1313027A (en) | 2001-04-06 | 2001-04-06 | Method for breeding wolfberry autotetraploid |
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| CN1313027A true CN1313027A (en) | 2001-09-19 |
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| CN 01111148 Pending CN1313027A (en) | 2001-04-06 | 2001-04-06 | Method for breeding wolfberry autotetraploid |
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Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103181324A (en) * | 2013-03-23 | 2013-07-03 | 甘肃农业大学 | Method for rapidly propagating high-quality seedlings of Lycium ruthenicum Murr. |
| CN103749289A (en) * | 2014-01-08 | 2014-04-30 | 河北科技师范学院 | Breeding method and preparation method of new variety of health-care boxthorn leaf tea |
| CN104094841A (en) * | 2014-03-24 | 2014-10-15 | 甘肃农业大学 | Tissue culture and rapid propagation method of solanaceae lycium brevipes |
| CN106561448A (en) * | 2016-10-24 | 2017-04-19 | 天津农学院 | Method for induction of lycium ruthenicum murr tetraploid through tissue culture |
-
2001
- 2001-04-06 CN CN 01111148 patent/CN1313027A/en active Pending
Cited By (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103181324A (en) * | 2013-03-23 | 2013-07-03 | 甘肃农业大学 | Method for rapidly propagating high-quality seedlings of Lycium ruthenicum Murr. |
| CN103181324B (en) * | 2013-03-23 | 2014-06-18 | 甘肃农业大学 | Method for rapidly propagating high-quality seedlings of Lycium ruthenicum Murr. |
| CN103749289A (en) * | 2014-01-08 | 2014-04-30 | 河北科技师范学院 | Breeding method and preparation method of new variety of health-care boxthorn leaf tea |
| CN104094841A (en) * | 2014-03-24 | 2014-10-15 | 甘肃农业大学 | Tissue culture and rapid propagation method of solanaceae lycium brevipes |
| CN104094841B (en) * | 2014-03-24 | 2016-05-11 | 甘肃农业大学 | Solanaceae Lycium short handle matrimony vine tissue is cultivated and method for quickly breeding |
| CN106561448A (en) * | 2016-10-24 | 2017-04-19 | 天津农学院 | Method for induction of lycium ruthenicum murr tetraploid through tissue culture |
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