CN106258984B - A kind of Chinese cabbage microspore differential medium formula - Google Patents
A kind of Chinese cabbage microspore differential medium formula Download PDFInfo
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- CN106258984B CN106258984B CN201610873779.7A CN201610873779A CN106258984B CN 106258984 B CN106258984 B CN 106258984B CN 201610873779 A CN201610873779 A CN 201610873779A CN 106258984 B CN106258984 B CN 106258984B
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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
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Abstract
The present invention provides a kind of Chinese cabbage microspore differential medium, its formula contains KNO3, phenylacetic acid potassium, MgSO4∙7H2O、CaCl2∙2H2O、(NH4)2SO4、NaH2PO4∙H2O、Na2‑EDTA、FeSO4∙7H2O、MnSO4∙4H2O、AgNO3、H3BO3、ZnSO4∙7H2O、KI、Na2MoO4∙2H2O、CuSO4∙5H2O、CoCl2∙6H2O, glutamine, vitamin B1, vitamin B6,2 morpholino b acids, nicotinic acid, folic acid, biotin, inositol, sorbierite, glycine, proline, sucrose, plant gel, NAA, Prohexadione calcium, colchicine, protein hydrolysate and activated carbon.The culture medium can significantly improve the green seedling regeneration efficiency of Chinese cabbage microspore callus, have important application value.
Description
Technical field
The present invention relates to a kind of Chinese cabbage microspore differential medium formula, and in particular to one kind can significantly improve great Bai
The green seedling regeneration efficiency of dish microspore callus, the Chinese cabbage microspore differential medium formula with significant application value, belong to
Vegetable cultivation science and technology field.
Background technology
Chinese cabbage（Brassica campestris ssp. pekinensisLour.）Belong to Cruciferae Brassica genus rue
A kind of sedge kind Chinese cabbage subspecies, are a kind of important vegetables for originating in China.Chinese cabbage ecotype is various, and distribution is wide, yield
Height, storage tolerance, supply phase length, nutrition is comprehensive, and eating method is various, and it is low to plant simple, saving of labor, cost, in China's vegetable basket
It is middle to occupy irreplaceable critical role.The cultivated area and yield of China Chinese cabbage are Chinese cabbage first of various vegetable crops
Production occupies leading position in vegetable production.Due to history, ecology, production and consumption custom difference, northern China
Area is the main producing region of Chinese cabbage.
With the improvement of people's life quality, it is also proposed the requirement of higher to Chinese cabbage quality, just there is an urgent need to educate for this
Kind worker selects more high-quality, disease-resistant, high yield improved seeds.Chinese cabbage is cross-pollinatd plant, is had significant
Hybrid vigour, mainly carries out New Chinese Cabbage Variety cultivation using cross breeding method at present.Conventional hybridization breeding method is bred as
A series of Chinese cabbage cultivars, the contribution of brilliance is made that for vegetables production.However, there are the cycle for Chinese cabbage crossing breeding method
The long, outstanding problem such as efficiency is low, is bred as a stables Elite inbred and generally requires 6-8, or even for more time, selection and breeding are selfed
System is very bothersome, takes a lot of work.Recently as the increase of cost of labor, the deficiency of conventional breeding is increasingly obvious in real work.
Therefore, scholars begin look for the new way and new method of breeding.
Microspore-isolated culture, refers to that free, fresh microspore colony is directly obtained from bud or flower medicine to carry out
Culture, via the induction of embryoid or callus, regenerates complete haplobiont, then passes through artificial or Natural double,
As the technology of normal fertile, homozygous liploid plant.Microspore-isolated culture has pure haploidy, unicellular property and culture
The features such as cell quantity is big, therefore increasingly favored be subject to breeder.The technology, which is used for breeding, to be obtained in 1-2
Elite inbred and the not affine pure lines of selfing, therefore the breeding time limit is substantially shorter, so as to improve breeding efficiency.Isolated microspore
Culture technique, more successful research field is grass family and Solanaceae at present, and a variety of grass family and solanaceous crops are proved to be successful
Go out efficient induction and differentiated system.The isolated microspore culture technique of brassicaceous vegetable has also carried out a series of researchs, right
The embryogenetic mechanism of Chinese cabbage microspore, influence factor have done substantial amounts of research and probe, and achieve many important researchs
Progress.
The Isolated Microspore in Chinese Cabbage culture studies in China start from the late 1980s.1988, agriculture section of Henan Province
Institute Horticultural Research Institute Li Genyi etc. at home first induces Isolated Microspore in Chinese Cabbage into embryo and embryo reproductive success.Then, river
The technology is applied to the culture of DH pure lines and parent material by Nan Sheng academies of agricultural sciences, has successfully been bred as Henan Chinese cabbage No. 7, has been become domestic and international
The new varieties that first Isolated Microspore in Chinese Cabbage cultural method is cultivated.Afterwards, and 14 successively are selected by above the provincial level
Examine（Mirror）Fixed different type New Chinese Cabbage Variety, the wherein popularizing area between 1994-1997 of Henan Chinese cabbage No. 7 reach 3.01 ten thousand
hm2.It is special that the Exocarpium Citri Rubrum heart kind that the Vegetable Research center of Beijing in 2002 is bred as using microspore culture obtains country's invention
Profit.Although Isolated Microspore in Chinese Cabbage culture breeding technique achieves a series of research progress, Henan Chinese cabbage No. 7, Henan are also cultivated
New No. 3, Yuyuan Garden 5, the improved seeds such as the Exocarpium Citri Rubrum heart, but Isolated Microspore Culture in Chinese cabbage and imperfection are mainly big
The problems such as Chinese cabbage Isolated microspore inductivity is not high, embryoid planting percent is low are never well solved, and directly limit
Application of the Isolated Microspore Culture in Chinese cabbage in breeding.
Culture medium is the material base of microspore-isolated culture, is directly related to the growth and differentiation of microspore.Microspore
Embryo callus subculture induces and callus breaks up two committed steps that two stages are Chinese cabbage microspores cultures.We are by for many years
Experiment is groped, and the improvement of creativeness has been carried out to Chinese cabbage microspore inducing culture, has substantially increased Chinese cabbage microspore embryoid
Property callus inductivity, established solid foundation for the foundation of the efficient microspores culture system of Chinese cabbage.But at present
Chinese cabbage microspore differential medium is not high to the green seedling regeneration efficiency of pollen callus, and becoming us, to establish Chinese cabbage efficiently small
Spore cultivation system must solve the problems, such as.
The content of the invention
The green seedling regeneration efficiency of Chinese cabbage microspore callus can be significantly improved the object of the present invention is to provide one kind, there is weight
The Chinese cabbage microspore differential medium formula for the application value wanted.
The purpose of the present invention is what is solved by the following technical programs：
A kind of Chinese cabbage microspore differential medium formula, it is characterised in that：The formula of the culture medium is as follows：
KNO32000-2400mg/L, phenylacetic acid K43 0-470mg/L, MgSO4∙7H2O 280-340mg/L, CaCl2∙
2H2O 270-320mg/L, (NH4)2SO490-110mg/L, NaH2PO4∙H2O 120-140mg/L, Na2-EDTA 35.6-
39mg/L, FeSO4∙7H2O 26-29.6mg/L, MnSO4∙4H2O 15-17mg/L, AgNO34.2-4.8mg/L, H3BO3 4.2-
4.8mg/L, ZnSO4∙7H2O 9.5-10.5mg/L, KI 0.7-0.8mg/L, Na2MoO4∙2H2O 0.2-0.3mg/L, CuSO4∙
5H2O 0.02-0.03mg/L, CoCl2∙6H2O 0.02-0.03mg/L, glutamine 750-850mg/L, vitamin B1 0.5-
0.6mg/L, vitamin B6 0.7-0.8mg/L, 2-morpholine ethane sulfonic acid 220-260mg/L, nicotinic acid 0.7-0.8mg/L, folic acid
0.15-0.25mg/L, biotin 0.06-0.08mg/L, inositol 90-110mg/L, 18~22g/L of sorbierite, glycine
1.8-2.2mg/L, proline 85-95mg/L, sucrose 27-33g/L, plant gel 5.3-5.7g/L, NAA 0.11-
0.15mg/L, Prohexadione calcium 0.08-0.12mg/L, colchicine 0.4-0.5mg/L, protein hydrolysate 1.3-1.5g/L, activity
Charcoal 0.025-0.035g/L, pH 5.5-5.7.
Chinese cabbage microspore differential medium provided by the present invention, is existing studies have found that further being created on basis
The achievement of the property made improvement, is the optimum combination screened by a large amount of single-factors, multifactor experiment, We conducted substantial amounts of reality
Test research and also show that the component proportion of the present invention is optimal.Great Bai can be significantly improved using culture medium provided by the present invention
The green seedling regeneration efficiency of dish microspore callus, have important application value.
Embodiment
With reference to embodiment, the present invention is further illustrated.
A kind of Chinese cabbage microspore differential medium formula, it is characterised in that：The formula of the culture medium is as follows：
KNO32000-2400mg/L, phenylacetic acid K43 0-470mg/L, MgSO4∙7H2O 280-340mg/L, CaCl2∙
2H2O 270-320mg/L, (NH4)2SO490-110mg/L, NaH2PO4∙H2O 120-140mg/L, Na2-EDTA 35.6-
39mg/L, FeSO4∙7H2O 26-29.6mg/L, MnSO4∙4H2O 15-17mg/L, AgNO34.2-4.8mg/L, H3BO3 4.2-
4.8mg/L, ZnSO4∙7H2O 9.5-10.5mg/L, KI 0.7-0.8mg/L, Na2MoO4∙2H2O 0.2-0.3mg/L, CuSO4∙
5H2O 0.02-0.03mg/L, CoCl2∙6H2O 0.02-0.03mg/L, glutamine 750-850mg/L, vitamin B1 0.5-
0.6mg/L, vitamin B6 0.7-0.8mg/L, 2-morpholine ethane sulfonic acid 220-260mg/L, nicotinic acid 0.7-0.8mg/L, folic acid
0.15-0.25mg/L, biotin 0.06-0.08mg/L, inositol 90-110mg/L, 18~22g/L of sorbierite, glycine
1.8-2.2mg/L, proline 85-95mg/L, sucrose 27-33g/L, plant gel 5.3-5.7g/L, NAA 0.11-
0.15mg/L, Prohexadione calcium 0.08-0.12mg/L, colchicine 0.4-0.5mg/L, protein hydrolysate 1.3-1.5g/L, activity
Charcoal 0.025-0.035g/L, pH 5.5-5.7.
In white No. 17 of Chinese cabbage Shandong and all-victorious initial bloom stage, take the bud of 2.2mm-3.0mm to take back laboratory, be placed in 4
It is inoculated with DEG C refrigerator after Cold pretreatment 1-2d, Chinese cabbage microspore is extracted under aseptic condition, is inoculated into what we researched and developed early period
In Isolated Microspore in Chinese Cabbage inducing culture, 20-22 DEG C of light culture, embryo callus starts to occur within 3 weeks or so, works as induction
When the Chinese cabbage microspore callus of culture grows to 0.3-0.6cm sizes, Chinese cabbage microspore provided by the present invention is gone to
Green seedling is induced in differential medium, condition of culture control is：22-24 DEG C of temperature, humidity 70%-85%, illumination 2500-3500lx,
Photoperiod light l4h/ dark 10h.After Shoot regeneration produces, green seedling regeneration rate is calculated.
Green seedling regeneration rate（%）Callus block number × 100% of=green seedling number/transfer.
It is culture medium that Chinese cabbage microspore differential medium more provided by the present invention and forefathers use to Chinese cabbage
The difference of the green seedling palingenesis of Isolated microspore embryo callus, in addition we have selected 2 kinds of forefathers and adopt used Chinese cabbage
Isolated microspore differential medium formula, for test variety also using Shandong white No. 17 and complete victory, tissue culture operating method, cultural method are equal
Identical, Shoot regeneration counts the green seedling regeneration of white No. 17 of Shandong and all-victorious microspore callus in 2 kinds of culture mediums respectively after producing
Rate.
Other 2 kinds of Chinese cabbages microspore differential medium formulas are：
NLN-13 culture medium+0.1g/L activated carbons, pH 5.8, a step cultivation（Documents come from Sun Dan《Great Bai
The small robe culture regeneration strain of dish and its otherness preliminary analysis》）；
B5+0.2mg/L 6-BA+0.02mg/L NAA, pH 5.8（Documents come from that to pay text graceful etc.《Chinese cabbage is swum
From microspore embryoid induction and plant regeneration》）.
Culture medium contrast experiment
No. 17 white to Shandong of the Chinese cabbage microspore differential medium provided by the present invention obtained and all-victorious small spore will be counted
The green seedling regeneration rate of son it is white to Shandong with culture medium used in other two kinds of forefathers in comparative example No. 17 and complete victory microspore it is green
Seedling regeneration rate is compared, and comparative result is as shown in table 1 below.
By table 1 it can be found that Chinese cabbage microspore differential medium provided by the present invention white to Chinese cabbage cultivar Shandong 17
Number and the all-victorious green seedling regeneration rate average value of microspore callus reached 90%, and other two kinds of culture mediums pair used by forefathers
White No. 17 of Chinese cabbage cultivar Shandong and the all-victorious green seedling regeneration rate average value of microspore callus are only 34.7% and 54.5% respectively, can be with
It was found that Chinese cabbage microspore differential medium provided by the present invention the green seedling regeneration rate of Chinese cabbage microspore callus is made that it is aobvious
The raising of work.
Chinese cabbage microspore differential medium provided by the present invention, is existing studies have found that further being created on basis
The achievement of the property made improvement, is the optimum combination screened by a large amount of single-factors, multifactor experiment, We conducted substantial amounts of reality
Test research and also show that the component proportion of the present invention is optimal.Great Bai can be significantly improved using culture medium provided by the present invention
The green seedling regeneration efficiency of dish microspore callus, have important application value.
Above example is merely illustrative of the invention's technical idea, it is impossible to protection scope of the present invention is limited with this, it is every
According to technological thought proposed by the present invention, any change done on the basis of technical solution, each falls within the scope of the present invention
Within；The technology that the present invention is not directed to can be realized by the prior art.
Claims (1)
- A kind of 1. Chinese cabbage microspore differential medium formula, it is characterised in that：The formula of the culture medium is as follows：KNO3 2000- 2400mg/L, phenylacetic acid K43 0-470mg/L, MgSO4 7H2O 280-340mg/L, CaCl2 2H2O 270-320mg/L, (NH4) 2SO4 90-110mg/L, NaH2PO4 H2O 120-140mg/L, Na2-EDTA 35.6-39mg/L, FeSO4 7H2O 26-29.6mg/L, MnSO4 4H2O 15-17mg/L, AgNO3 4.2-4.8mg/L, H3BO3 4.2-4.8mg/L, ZnSO4 7H2O 9.5-10.5mg/L, KI 0.7-0.8mg/L, Na2MoO4 2H2O 0.2-0.3mg/L, CuSO4 5H2O 0.02- 0.03mg/L, CoCl2 6H2O 0.02-0.03mg/L, glutamine 750-850mg/L, vitamin B1 0.5-0.6mg/L, Vitamin B6 0.7-0.8mg/L, 2-morpholine ethane sulfonic acid 220-260mg/L, nicotinic acid 0.7-0.8mg/L, folic acid 0.15- 0.25mg/L, biotin 0.06-0.08mg/L, inositol 90-110mg/L, 18~22g/L of sorbierite, glycine 1.8- 2.2mg/L, proline 85-95mg/L, sucrose 27-33g/L, plant gel 5.3-5.7g/L, NAA 0.11-0.15mg/L, Prohexadione calcium 0.08-0.12mg/L, colchicine 0.4-0.5mg/L, protein hydrolysate 1.3-1.5g/L, activated carbon 0.025- 0.035g/L, pH 5.5-5.7.
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| CN107173235B (en) * | 2017-07-19 | 2018-12-07 | 徐州嘉农农业发展有限公司 | A kind of pumpkin microspore differential medium |
| CN107211895B (en) * | 2017-07-19 | 2018-12-07 | 徐州嘉农农业发展有限公司 | A kind of muskmelon Anther culture breeding differential medium |
| CN108184677A (en) * | 2018-04-02 | 2018-06-22 | 张瑞明 | A kind of pakchoi Isolated microspore Fiber differentiation based formulas |
| CN108849521A (en) * | 2018-08-07 | 2018-11-23 | 江苏高航农业科技有限公司 | A kind of differential medium and its preparation flow improving woaded blue Efficiency |
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