CN106148394A - A kind of nonobstructive azoospermia method utilizing open country chrysanthemum foot bud to be outer implant - Google Patents
A kind of nonobstructive azoospermia method utilizing open country chrysanthemum foot bud to be outer implant Download PDFInfo
- Publication number
- CN106148394A CN106148394A CN201510252909.0A CN201510252909A CN106148394A CN 106148394 A CN106148394 A CN 106148394A CN 201510252909 A CN201510252909 A CN 201510252909A CN 106148394 A CN106148394 A CN 106148394A
- Authority
- CN
- China
- Prior art keywords
- foot
- open country
- bud
- bacterium solution
- plant
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Pending
Links
Landscapes
- Breeding Of Plants And Reproduction By Means Of Culturing (AREA)
Abstract
The non-group of training genetic transforming method utilizing open country chrysanthemum foot bud to be outer implant, belongs to field of plant genetic.The present invention utilizes the method for the non-tissue culture of plant, it is that outer implant carries out genetic transformation by vacuum infiltration with plant foot bud, not only speed is fast, simple to operate, structural integrity, maternal merit can be kept, compensate for traditional plant gene genetic method for transformation operation complexity, the weak point of transformation period length.
Description
Technical field
The invention belongs to field of plant genetic, relate to non-group of training genetic transforming method of a kind of open country chrysanthemum ' flame '.
Background technology
The core of plant genetic engineering research is Genetic Transformation in Higher Plants, i.e. use certain transgenic method such as agrobacterium-mediated transformation, particle bombardment etc. to be proceeded in recipient plant cell by exogenous gene (including genes of interest fragment, plasmid DNA and plant STb gene), and stably entail offspring.Research worker constantly researches and develops new plant genetic transformation method to improve transformation efficiency and to simplify Transformation Program.At present, the genetic transforming method being applied in agricultural transgenic mainly divides two big classes: genetic transforming method based on tissue culture and nonobstructive azoospermia method.Genetic transforming method based on tissue culture has agrobacterium-mediated transformation, particle bombardment, electric shocking method, PEG method etc., but easily occurs the problems such as pollution, brown stain, vitrification in tissue culture, adds tissue culture's difficulty, thus affects genetic transformation efficiency.Therefore the research work to plant nonobstructive azoospermia method is becoming a focus of plant genetic engineering field, studying it would be possible to plant transformation efficiency is greatly improved further of nonobstructive azoospermia method, the most significant to the research work of plant molecular breeding and late gene transcriptional expression.
Open country chrysanthemum (Chrysanthemum) is a class novel Dendranthema morifolium Varieties group, and for Compositae Chrysanthemum, herbaceos perennial, its plant type is short strong, have cold-resistant, drought resisting, anti-barren, Resistant, the characteristic such as the flouring and extensive management of blooming, afforestation occupies critical role.The existing Dendranthema morifolium Varieties of China up to more than 3000.Flos Chrysanthemi is mainly used in park, garden, flower bed and indoor appreciation, and along with expanding economy, the raising of living standards of the people, Flos Chrysanthemi has become a consumption hot spot of domestic flower market.
Summary of the invention
The invention aims to set up a kind of with non-group of training genetic transforming method that open country Radix Chrysanthemi portion foot bud is outer implant, the genetic transformation of plant is completed by utilizing the fluid medium of Agrobacterium bacterium solution that external source genetic recipient is carried out vacuum infiltration, this nonobstructive azoospermia method not only speed is fast, transformation efficiency is high, without carrying out tissue culture's work of complexity, the more important thing is and be not likely to produce variation that virus-free pollution can keep maternal merit.
In current open country chrysanthemum transgenic protocol, tissue culture's job demand spends substantial amounts of man power and material, occupies the biggest proportion, and in operation, virus inevitably infects plant in experimentation cost, and this is also the reason making transformation benefit decline.Simplify open country chrysanthemum transgenic protocol by the present invention, shorten the transformation period, the most easy and simple to handle, and in reproductive process, occur that the danger of variation is few, and virus-free pollution.
Owing to open country chrysanthemum genetic background is sufficiently complex, genotype height heterozygosis, traditional gene genetic method for transformation suffers from the polyploidy of chromosome and the impact of aneuploidy, brings the problems such as gene silencing, transformation efficiency are low.Additionally, open country chrysanthemum conventional genetic method for transformation also exists transformation period too long of problem.In terms of the genetic transformation particularly resistance cultivation of current open country chrysanthemum new varieties, utilize non-group of training transgenic technology can overcome above all unfavorable conditions.But successfully first gene transformation depends on the foundation of good receptor system, the present invention set up with open country chrysanthemum foot bud be outer implant be preferable gene transformation receptor system.
Accompanying drawing explanation
Fig. 1 is open country chrysanthemum ' flame ' root foot bud；Fig. 2 is the root foot bud after open country chrysanthemum ' flame ' bacterium solution vacuum infiltration conversion processing；Fig. 3 is PCR detection electrophoretogram (special primer: PMI-F/R M:D2000+: positive control wt: negative control) of open country chrysanthemum ' flame ' transformed plant；Fig. 4 is GUS colored graph (the A. WT lines of open country chrysanthemum ' flame ' transformed plant；B and C. bacterium solution vacuum infiltration plant；Bar=5mm).
Detailed description of the invention
Detailed description of the invention, as a example by open country chrysanthemum ' flame ' nonobstructive azoospermia method, sequentially includes the following steps:
(1) the choosing of open country chrysanthemum ' flame ' root foot bud
Taking the healthy and strong foot bud that open country chrysanthemum ' flame ' base portion of nursery lot plantation is sprouted, remove the blade grown, flowing water rinses 30min.
(2) nonobstructive azoospermia method processes test foot bud
The LB fluid medium utilizing Agrobacterium bacterium solution carries out non-group of training method conversion processing to foot bud, the method processed is as follows: Agrobacterium bacterium solution grows into OD value when being 0.4, foot bud is put into (amount putting into foot bud is 50 foot buds with bacterium solution ratio: 50mL bacterium solution) in bacterium solution, 2 times (liquid boiling is that standard has calculated a sub-percolation to vacuum infiltration, a length of 3min a when time, twice duration is about 7min).
(3) the foot bud cottage propagation after nonobstructive azoospermia method being processed
Test foot bud after being processed by vacuum infiltration is cottage propagation in husky basin, after dark surrounds cultivates 1d at normal lighting conditions under grow.
(4) plant after converting is carried out molecular biology identification
Transplanting after long root in soil, after 1-2 to be grown month, take CTAB method to extract the STb gene of open country chrysanthemum ' flame ' blade newly grown, then DNA product carries out the Preliminary screening of the PCR detection of specific gene, testing result display transformation efficiency can reach 10%-15%；After growth 3-4 month, again carrying out the PCR detection of specific gene, testing result display transformation efficiency can reach 2%-3%.The positive plant of PCR detection is carried out GUS dyeing, and result shows that the gene converted has significantly expression in open country chrysanthemum flame body.
Claims (2)
1. choosing open country chrysanthemum foot bud and carry out non-group of training method genetic transformation after flowing water rinses 30min, culture medium prescription: the LB fluid medium of Agrobacterium bacterium solution, OD value is 0.4.
2. it is outer implant with open country Radix Chrysanthemi portion foot bud, with the LB fluid medium of Agrobacterium bacterium solution, it is processed, the method processed is: put into by foot bud (amount putting into foot bud is 50 foot buds with bacterium solution ratio: 50mL bacterium solution) in bacterium solution, 2 times (liquid boiling is that standard has calculated a sub-percolation to vacuum infiltration, a length of 3min a when time, twice duration is about 7min).After process, cuttage is in sandy soil, and cultivates 1d in dark surrounds；Transplant after long root in soil, laggard performing PCR detection in 1-2 month.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201510252909.0A CN106148394A (en) | 2015-05-18 | 2015-05-18 | A kind of nonobstructive azoospermia method utilizing open country chrysanthemum foot bud to be outer implant |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201510252909.0A CN106148394A (en) | 2015-05-18 | 2015-05-18 | A kind of nonobstructive azoospermia method utilizing open country chrysanthemum foot bud to be outer implant |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN106148394A true CN106148394A (en) | 2016-11-23 |
Family
ID=57348162
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201510252909.0A Pending CN106148394A (en) | 2015-05-18 | 2015-05-18 | A kind of nonobstructive azoospermia method utilizing open country chrysanthemum foot bud to be outer implant |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN106148394A (en) |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN116042694A (en) * | 2022-11-24 | 2023-05-02 | 中国科学院南京土壤研究所 | Non-tissue culture genetic transformation method for pennisetum plants of Gramineae |
Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104450779A (en) * | 2014-12-15 | 2015-03-25 | 贵州大学 | Fast and efficient agrobacterium-mediated rice stem tip genetic transformation method |
-
2015
- 2015-05-18 CN CN201510252909.0A patent/CN106148394A/en active Pending
Patent Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104450779A (en) * | 2014-12-15 | 2015-03-25 | 贵州大学 | Fast and efficient agrobacterium-mediated rice stem tip genetic transformation method |
Non-Patent Citations (5)
| Title |
|---|
| 佟玲: "露地菊分化体系的构建及DFR基因遗传转化的研究", 《中国优秀硕士学位论文全文数据库农业科技辑》 * |
| 张燕红等: "农杆菌真空渗透法转化棉花花粉的初步研究", 《棉花学报》 * |
| 李晓等: "农杆菌真空渗透法转化棉花花粉获得棉花转acsB基因植株", 《作物学报》 * |
| 王羽晗等: "露地菊脚芽农杆菌真空渗透转化技术体系的建立", 《南方农业学报》 * |
| 银春华: "《花卉栽培》", 1 April 2015 * |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN116042694A (en) * | 2022-11-24 | 2023-05-02 | 中国科学院南京土壤研究所 | Non-tissue culture genetic transformation method for pennisetum plants of Gramineae |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN101869591B (en) | Method for producing alkannin by utilizing Arnebia euchroma(Royle)Johnst hairy root | |
| CN104206270B (en) | The tissue cultures of a kind of flourishing torch root tuber of aromatic turmeric and rhizome expand method | |
| WO2022135246A1 (en) | R gene for controlling matching of soybean-rhizobium, protein and use thereof | |
| CN102119655A (en) | Natural light rapid breeding method for dendrobium officinale | |
| CN105432464B (en) | Cultivation method for inducing autotetraploid of paulownia catalpa through colchicine | |
| CN104041414A (en) | Method for obtaining haploid regenerated plant by chilli anther culturing | |
| CN108300735A (en) | A kind of Lilium tenuifolium efficient genetic trasformation system based on somatic embryo occur | |
| CN102286526B (en) | Method for quickly obtaining capsicum transgenic plant | |
| CN103060369B (en) | Hybrid crop transgenic safety control method and gene deletion system for implementing same | |
| CN104012416A (en) | Method for efficiently regenerating Sapium sebiferum leaf disc | |
| CN106854659A (en) | A kind of method of sinocalamus latiflorus genetic transformation and plant regeneration | |
| CN105039396A (en) | Silvergrass seed callus transforming method mediated by agrobacterium and application of method | |
| CN101775409A (en) | Method for obtaining transgenic corns by infecting young ears with agrobacterium | |
| CN106148394A (en) | A kind of nonobstructive azoospermia method utilizing open country chrysanthemum foot bud to be outer implant | |
| CN115044606B (en) | Method for establishing agrobacterium rhizogenes-mediated jujube genetic transformation system | |
| CN103026966B (en) | Method for identifying tomato yellow leaf curl virus resistance by utilizing detached leaf | |
| CN104285788B (en) | A kind of method setting up green bamboo regenerating system by body embryogenesis path | |
| CN103120127A (en) | Method for quickly inducing and breeding cotton embryogenic callus | |
| CN103740751B (en) | A kind of agriculture bacillus mediated sugarcane genetically modified method | |
| CN110305894A (en) | A kind of Chinese catalpa genetic transforming method rapidly and efficiently | |
| CN106613970B (en) | The quick breeding by group culture method of sealwort leaf elegant jessamine | |
| CN103215307B (en) | Genetic transformation method of agrobacterium-induced plum blossom mature cotyledon regeneration system | |
| CN103053421B (en) | Chinese pistache rapid propagation method | |
| CN103238517A (en) | Induction method of nitraria loose embryonic callus | |
| Lei et al. | Agrobacterium-mediated transformation of cotton shoot apex with SNC1 gene and resistance to cotton Fusarium wilt in T1 generation |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| C06 | Publication | ||
| PB01 | Publication | ||
| C10 | Entry into substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| RJ01 | Rejection of invention patent application after publication |
Application publication date: 20161123 |
|
| RJ01 | Rejection of invention patent application after publication |