CN106035059B - A kind of method of artificial synthesized anti-clubroot brassica napus - Google Patents
A kind of method of artificial synthesized anti-clubroot brassica napus Download PDFInfo
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- CN106035059B CN106035059B CN201610363713.3A CN201610363713A CN106035059B CN 106035059 B CN106035059 B CN 106035059B CN 201610363713 A CN201610363713 A CN 201610363713A CN 106035059 B CN106035059 B CN 106035059B
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- clubroot
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- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01H—NEW PLANTS OR NON-TRANSGENIC PROCESSES FOR OBTAINING THEM; PLANT REPRODUCTION BY TISSUE CULTURE TECHNIQUES
- A01H1/00—Processes for modifying genotypes ; Plants characterised by associated natural traits
- A01H1/02—Methods or apparatus for hybridisation; Artificial pollination ; Fertility
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- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01G—HORTICULTURE; CULTIVATION OF VEGETABLES, FLOWERS, RICE, FRUIT, VINES, HOPS OR SEAWEED; FORESTRY; WATERING
- A01G7/00—Botany in general
- A01G7/06—Treatment of growing trees or plants, e.g. for preventing decay of wood, for tingeing flowers or wood, for prolonging the life of plants
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- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01H—NEW PLANTS OR NON-TRANSGENIC PROCESSES FOR OBTAINING THEM; PLANT REPRODUCTION BY TISSUE CULTURE TECHNIQUES
- A01H1/00—Processes for modifying genotypes ; Plants characterised by associated natural traits
- A01H1/06—Processes for producing mutations, e.g. treatment with chemicals or with radiation
- A01H1/08—Methods for producing changes in chromosome number
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- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01H—NEW PLANTS OR NON-TRANSGENIC PROCESSES FOR OBTAINING THEM; PLANT REPRODUCTION BY TISSUE CULTURE TECHNIQUES
- A01H4/00—Plant reproduction by tissue culture techniques ; Tissue culture techniques therefor
- A01H4/008—Methods for regeneration to complete plants
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Abstract
The invention discloses a kind of method of artificial synthesized anti-clubroot cabbage type rape new material, hybridize using the wild cabbage of anti-clubroot to be maternal with the pakchoi of anti-clubroot, after F1 generation is pollinated 15 days, select seed and carry out rescue culture, utilize the immature seed of B5 fluid nutrient medium cultures containing 2% sucrose, when growing cotyledon after rataria appearance, embryo is moved on the B5 solid mediums containing 2% sucrose, doubled when small using 0.1% colchicine processing 2 during 34 leaf phase, moved to after rinsed clean in soil, make the plant selfing of synthesis using the method for cabage-type rape or Spraying in the flowering stage brine；The seed of selfing is inoculated with using clubroot P4 biological strains in germination period, while is inoculated with the same time as compareing using susceptible material, after 35 days, is taken out the root of seedling, is cleaned and has been seen whether lump, the plant that no lump illustrates to synthesize has clubroot resistance.The present invention provides the strain effective way of the anti-clubroot cabbage type rape of a selection and breeding, clubroot inoculation efficiency is improved.
Description
Technical field
The present invention relates to the artificial synthesized technical field of cabbage type rape, is specifically a kind of artificial synthesized anti-clubroot Wild cabbage type
The method of rape material.
Background technology
Clubroot is a kind of fungal disease as caused by Brassica genus plasmodiophora brassicae (Plasmodiophora brassicae), existing
The whole world is distributed in, Temperate Region in China incidence is particularly acute.Nineteen ninety-five, China report rapeseed clubroot first, then should
Disease national large area spread, at present major part provinces, municipalities and autonomous regions be distributed, but be mainly distributed on Yunnan, Sichuan, Guizhou,
The ground such as Hubei, Anhui, Southern Shaanxi.Almost all of crucifer is infected and endangered to the germ, and particularly Brassica genus is planted
Thing, such as Chinese cabbage, wild cabbage (Piao etc., 2009).Rapeseed clubroot takes place from seedling stage, mainly infects root, causes plant
Growth retardation, blade turn yellow, and plant is downgraded, and shows water shortage, and seedling is withered when serious.The susceptible tumour of Adult plant mostly occurs in side
The lower part of root and main root, advanced stage tumours cracking and also it is coarse, finally rot, seriously affected the normal absorption of moisture and nutrient,
So as to cause rape Severe Reduction or total crop failure.Rapeseed clubroot seedling stage incidence is up to 17% according to research reports, Adult plant field
Average attack rate is 15%, typically results in underproduction 10%-20%, the serious field underproduction more than 50% (Tao Xuejun, 2015).At me
State, ten thousand hectares of the long-term hazard area of clubroot about 320-400, accounts for more than the 1/3 of crop in cruciferae cultivated area, is very popular year
Part occurs with hazard area up to 9,000,000 hectares (Wang Jing etc., 2008).
Cruciferae clubroot is there are multiple biological strains, and in China, at least there are 2 for cabbage type rape clubroot cause of disease
Number, 3 biological strain types such as No. 4 and No. 13, wherein widest in area general existing for No. 4 microspecies.At abroad, such as Canada, tool
There are at least five biological strains such as 2,3,5,6,8, wherein predominantly No. 3 biological strains.Due to corresponding to different biological strains
Disease-causing gene is not quite similar, after the kind of a certain anti-clubroot in area is planted to other region, usually due to biological strain
Change and cause original anti-clubroot kind to be changed into feeling the kind of clubroot, this brings greatly to the prevention of clubroot
It is difficult.Further, since some existing Cruciferae material resistances are relatively simple, its resistance is gradually degenerated after planting for many years
(Leboldus etc., 2012), therefore, is badly in need of cultivating the kind for polymerizeing multiple resistant genes.
Since clubroot infectiousness is strong, tool low concentration causes a disease, threshold value (Hatakeyama etc., 2013), spread speed are fast, pass
Broadcast that approach is more, and understanding deficiency of our and pathogenic disastrous mechanisms etc. so that clubroot becomes generation. the history of life to the germ
Crushing soilborne fungal pathogens in the range of boundary.Can not fundamentally it be solved using traditional chemical method to prevent clubroot
Rapeseed clubroot is propagated, while also brings the pollution of environment, is unfavorable for the sustainable development of agricultural.Current many experts are formed
Common recognition, the effective ways for solving rapeseed clubroot are the new rape varieties for cultivating anti-clubroot, are phased out the product of no resistance
Kind.But the rapeseed germplasm of cabbage type rape moderate resistance clubroot extremely lacks at present, it is difficult to be effectively applied in rapeseed breeding,
And there is the material of many anti-clubroots in the nearly edge species Chinese cabbage of cabbage type rape, wild cabbage etc..Therefore, we are badly in need of studying
The heredity of clubroot and the mechanism of action, formulate the rape material of anti-clubroot, cultivate the new rape variety of anti-clubroot, and are subject to
Promote, alleviate the harm of China's rapeseed clubroot.
At present, the inoculation method of the clubroot of most places is more extensive, or even many units are only reflected with Field inoculation
Fixed result also mixes native inoculation using bacterium solution mostly as the whether anti-clubroot of plant is judged, even if some in laboratory inoculation
Method, individual often occurs without successful phenomenon is inoculated with, the thus anti-clubroot identification to germ plasm resource is brought very big
Error, usually the prior year be accredited as anti-clubroot material and next year show as sense clubroot.Clubroot exists in recent years
China is spread rapidly rape main producing region, has seriously affected the production of China's rape, therefore is badly in need of cultivating the oil with anti-clubroot
Dish new varieties.But since the resource of China's cabbage type rape moderate resistance clubroot is extremely rare, and its nearly edge species Chinese cabbage, wild cabbage,
There are the material of anti-clubroot in black mustard, therefore the present invention intends the method using distant hybridization, utilizes resisting in Chinese cabbage and wild cabbage
The germ plasm resource of clubroot, the brassica napus of artificial synthesized anti-clubroot, gropes an anti-clubroot Wild cabbage type of synthesis
The method of napus lines, makes up the shortage of China's Brassica Napus clubroot resource.Carry out clubroot inoculation technique at the same time
Research, improves the inoculation efficiency of clubroot, and good technology guarantee is provided for the Resistance Identification of clubroot.Therefore, it is badly in need of to original
Some clubroot inoculation methods are improved, and are improved clubroot and are inoculated with successfully efficiency.
The content of the invention
It is an object of the invention to provide a kind of method of artificial synthesized anti-clubroot brassica napus, with solution
State the problem of being proposed in background technology.
To achieve the above object, the present invention provides following technical solution：
A kind of method of artificial synthesized anti-clubroot brassica napus, using the wild cabbage of anti-clubroot as maternal and anti-
The pakchoi hybridization for the disease that swells, after F1 generation is pollinated 15 days, is selected seed and carries out rescue culture, trained using the B5 liquid containing 2% sucrose
The immature seed of base culture is supported, when growing cotyledon after rataria appearance, embryo is moved into the B5 solid mediums containing 2% sucrose
On, doubled when small using 0.1% colchicine processing 2 during the 3-4 leaf phases, moved to after rinsed clean in soil, utilize stripping
Flower bud pollinates or the method for Spraying in the flowering stage brine makes the plant selfing of synthesis, and the seed of selfing utilizes clubroot P4 in germination period
Biological strain is inoculated with, while using susceptible material as control, is inoculated with the same time；After 35 days, the root of seedling is taken out, clean observation is
No to have lump, no lump illustrates that the plant of synthesis has clubroot resistance.
As the further scheme of the present invention：The wild cabbage be cabbage variety century green offspring, the little Bai
Dish is the offspring of varieties of Brassica campestris four seasons green vegetables.
As further scheme of the invention：Peduncle-growing period for rapeseed choose tender leaf using Ploidy Analyzer to plant into
Row Ploidy Identification, the fertility of plant is observed during the florescence, final to judge double whether to succeed.
As further scheme of the invention：It is not affine that the stripping flower bud selfing or Spraying in the flowering stage brine remove selfing.
As further scheme of the invention：Specific inoculation method is as follows：By ripe seed vernalization on filter paper
Afterwards, root is soaked in the bacterium solution of the P4 biological strains of 107 spores/milliliter 2 it is small when, then transplanting is to containing weakly acidic summary
In wet matrix, while every young shoot is inoculated with 5 milliliters of 107 spores/milliliter bacterium solution into matrix, and transplanting young shoot is moved to greenhouse
In.
As further scheme of the invention：The Greenhouse day temperature is 22 DEG C, and night temperatures are 18 DEG C.
Compared with prior art, the beneficial effects of the invention are as follows：Provide the anti-clubroot cabbage type rape of a selection and breeding
Strain effective way, improves clubroot inoculation efficiency.
Brief description of the drawings
Fig. 1 is the anti-clubroot cabbage type rape strain in the method for artificial synthesized anti-clubroot brassica napus
Synthetic line figure.
Fig. 2 is clubroot Disease Resistance Identification schematic diagram in the method for artificial synthesized anti-clubroot brassica napus.
Embodiment
Below in conjunction with the attached drawing in the embodiment of the present invention, the technical solution in the embodiment of the present invention is carried out clear, complete
Site preparation describes, it is clear that described embodiment is only part of the embodiment of the present invention, instead of all the embodiments.It is based on
Embodiment in the present invention, those of ordinary skill in the art are obtained every other without making creative work
Embodiment, belongs to the scope of protection of the invention.
Please refer to Fig.1~2, in the embodiment of the present invention, a kind of method of artificial synthesized anti-clubroot brassica napus,
It is the maternal and pakchoi of anti-clubroot with the wild cabbage (offspring for coming from cabbage variety-" century green ") of anti-clubroot
(offspring for coming from varieties of Brassica campestris " four seasons green vegetables ") hybridizes, F1After generation pollination 15 days, select seed and carry out rescue culture, utilize
The immature seed of B5 fluid nutrient medium cultures containing 2% sucrose, when growing cotyledon after rataria appearance, embryo is moved to and is contained
On the B5 solid mediums of 2% sucrose, doubled when small using 0.1% colchicine processing 2 during the 3-4 leaf phases, rinsing is dry
Moved to after net in soil, tender leaf is chosen using Ploidy Analyzer to plant progress Ploidy Identification in peduncle-growing period for rapeseed, when florescence is seen
The fertility of plant is examined, it is final to judge double whether to succeed.Method using cabage-type rape or Spraying in the flowering stage brine makes synthesis
Plant selfing (remove selfing not affine), the seed of selfing are inoculated with using clubroot P4 biological strains in germination period, while with sense
The material of disease is inoculated with as control, after 35 days, takes out the root of seedling, clean and seen whether lump, illustrate without lump at the same time
The plant of synthesis has clubroot resistance.
Specific inoculation method is as follows：By ripe seed on filter paper after vernalization, by root 107Spore/milliliter
When immersion 2 is small in the bacterium solution of P4 biological strains, then transplanting extremely contains in weakly acidic matrix (slightly wet), while is inoculated with per young shoot
The 10 of 5 milliliters7Spore/milliliter bacterium solution moves to transplanting young shoot in greenhouse (22/18 DEG C, day/night), first 3 days into matrix
Try not to water, if occurring water shortage after 3 days, just spray a small amount of moisture in right amount with watering can, only keep the moistening of matrix, should not
There is bright water.
Using Chinese cabbage and the anti-clubroot resource in wild cabbage, the cabbage type rape of artificial synthesized anti-clubroot, is adopted the present invention
The method identified with indoor inoculation, adjusts original indoor inoculation parameter, in Germination period inoculation seedling root, uses at the same time
Stringent management measure, raising are inoculated with successfully efficiency.
It is obvious to a person skilled in the art that the invention is not restricted to the details of above-mentioned one exemplary embodiment, Er Qie
In the case of without departing substantially from spirit or essential attributes of the invention, the present invention can be realized in other specific forms.Therefore, no matter
From the point of view of which point, the present embodiments are to be considered as illustrative and not restrictive, and the scope of the present invention is by appended power
Profit requires rather than described above limits, it is intended that all in the implication and scope of the equivalency of claim by falling
Change is included in the present invention.Any reference numeral in claim should not be considered as to the involved claim of limitation.This
Outside, it will be appreciated that although the present specification is described in terms of embodiments, not each embodiment only includes an independence
Technical solution, this narrating mode of specification is only that those skilled in the art should make specification for clarity
For an entirety, the technical solutions in the various embodiments may also be suitably combined, formed it will be appreciated by those skilled in the art that
Other embodiment.
Claims (5)
- A kind of 1. method of artificial synthesized anti-clubroot brassica napus, it is characterised in that using the wild cabbage of anti-clubroot as It is maternal to hybridize with the pakchoi of anti-clubroot, F1After generation pollination 15 days, select seed and carry out rescue culture, using containing 2% sucrose The immature seed of B5 fluid nutrient medium cultures, when rataria appearance after grow cotyledon when, embryo is moved into the B5 containing 2% sucrose and is consolidated On body culture medium, doubled when small using 0.1% colchicine processing 2 during the 3-4 leaf phases, soil is moved to after rinsed clean In, the plant selfing of synthesis is made using the method for cabage-type rape or Spraying in the flowering stage brine, the seed of selfing is in germination period, utilization Clubroot P4 biological strains are inoculated with, while using susceptible material as control, are inoculated with the same time, the specific method of the inoculation is such as Under：By ripe seed on filter paper after vernalization, by root 107Immersion 2 is small in the bacterium solution of the P4 biological strains of spore/milliliter When, then transplanting extremely contains in weakly acidic slightly wet matrix, while the 10 of every 5 milliliters of young shoot inoculation7Spore/milliliter bacterium solution arrives In matrix, transplanting young shoot is moved in greenhouse；After 35 days, the root of seedling is taken out, cleans and has seen whether lump, said without lump The plant of bright synthesis has clubroot resistance.
- 2. the method for artificial synthesized anti-clubroot brassica napus according to claim 1, it is characterised in that described Wild cabbage is the green offspring in cabbage variety century, and the pakchoi is the offspring of varieties of Brassica campestris four seasons green vegetables.
- 3. the method for artificial synthesized anti-clubroot brassica napus according to claim 1, it is characterised in that taking out A kind of sedge phase chooses tender leaf and carries out Ploidy Identification to plant using Ploidy Analyzer, and the fertility of observation plant, finally sentences during the florescence It is disconnected to double whether to succeed.
- 4. the method for artificial synthesized anti-clubroot brassica napus according to claim 1, it is characterised in that described Shell flower bud selfing or the removal selfing of Spraying in the flowering stage brine is not affine.
- 5. the method for artificial synthesized anti-clubroot brassica napus according to claim 1, it is characterised in that described Greenhouse day temperature be 22 DEG C, night temperatures be 18 DEG C.
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| CN108617494A (en) * | 2017-03-17 | 2018-10-09 | 四川省农业科学院作物研究所 | A kind of selection of Brassica Napus clubroot maintainer |
| CN109042291A (en) * | 2018-07-28 | 2018-12-21 | 湖南科技大学 | A method of novel vegetable is created using Chinese cabbage and wild cabbage distant hybridization |
| CN110999779B (en) * | 2019-12-13 | 2023-01-20 | 西北农林科技大学 | Method for synthesizing clubroot-resistant brassica napus by using polymerization breeding and biotechnology |
| CN114303516B (en) * | 2022-01-04 | 2022-12-13 | 沈阳农业大学 | Method for inoculating plasmodiophora brassicae and application of plasmodiophora brassicae in screening of plasmodiophora brassicae resistant breeding materials |
| CN115918517A (en) * | 2022-10-31 | 2023-04-07 | 云南省农业科学院园艺作物研究所 | Rapid breeding method of anti-clubroot bolting vegetable breeding material |
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| CN101785425B (en) * | 2009-12-29 | 2013-04-17 | 西南大学 | Method of improved artificially synthesized Brassica napa L. for heterosis utilization |
| CN103210839B (en) * | 2013-05-06 | 2014-09-24 | 西南大学 | Method for improving sclerotiniose resistance of oilseed rapes |
| CN104404124A (en) * | 2014-11-20 | 2015-03-11 | 上海市农业科学院 | Identification method for clubroot resistance of non-heading cabbages |
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