CN108094194A - A kind of selection of richness potassium tobacco bred - Google Patents
A kind of selection of richness potassium tobacco bred Download PDFInfo
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- CN108094194A CN108094194A CN201711144959.2A CN201711144959A CN108094194A CN 108094194 A CN108094194 A CN 108094194A CN 201711144959 A CN201711144959 A CN 201711144959A CN 108094194 A CN108094194 A CN 108094194A
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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
- A01H1/04—Processes of selection involving genotypic or phenotypic markers; Methods of using phenotypic markers for selection
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Abstract
The present invention provides a kind of selections of rich potassium tobacco bred, it selects and handles including seed, low potassium culture medium is prepared and sowing culture, and low-kalium resistant tobacco seedlings are not screened, crop field verification, isogeneity, seed sows culture after processing catalysis shows money or valuables one carries unintentionally in low potassium culture medium in the above process, filters out not low-kalium resistant cigarette strain according to root long situation after cultivating 30 days, and cigarette strain is divided into three groups, the cigarette strain after grouping is transplanted to crop field again and is verified, and carries out isogeneity.The method of selection and breeding richness potassium tobacco bred of the present invention is simply direct, and the later stage further verifies result with reference to field experiment, and experimental result is more accurate, stablizes.
Description
Technical field
The invention belongs to tobacco bred selection and breeding fields, and in particular to one kind is using tobacco mutant body as raw material, low potassium culture item
The method that rich potassium tobacco bred is screened under part.
Background technology
Potassium is one of most important Quality Element of tobacco, is pacified with Maturity of Tobacco Leaf, flammability, flavor and taste and cigarette products
Full property is closely related.Compared with external potassium content of tobacco leaf (general ＞ 3%), most of China area especially northern area, cigarette
The leaf potassium content that is averaged rarely exceeds 1.5%, and relatively low potassium content of tobacco leaf is one of key index for restricting Tobacco increased quality.
Production sound tobacco must need 2~3 times of potash fertilizer of potassium amount using optimum yeild is formed, and can just significantly improve potassium content in tobacco leaf.
But China's Potassic fertilizer resources are deficient, have certain limit again by improving agronomic conditions to improve potassium content, therefore screening or selection and breeding are baked
Cigarette potassium high efficiency gene type kind, make full use of soil neutralize application potassium, to improve potassium content of tobacco leaf, reduce production cost,
It economizes on resources significant.
At present, there are mainly two types of the methods for cultivating rich potassium volcanic rocks tobacco bred：First, by different potassium absorption efficiencies
Tobacco parent hybridized, measure the potassium content of parent and filial generation, hybrid vigour carried out to parent and cross combination, is matched somebody with somebody
With joint efforts and Hereditary Capacity, so as to obtain potassium high efficiency gene type plant；Second is that some potassium are efficiently planted using technique for gene engineering
The potassium high efficiency gene of strain is transferred in other plant and induces its expression, can quickly and efficiently obtain potassium high-efficiency plant.It is right
The screening of flue-cured tobacco potassium high efficiency gene type mainly carries out in terms of following two：
1. pass through hydroponics, potting and the more existing tobacco germplasm potassium nutrition property difference of field trial, choosing
Go out potassium high efficiency gene type；
2nd, measure and absorbed with potassium and transport expression of the relevant gene in blade, wherein with the relevant pass of potassium content
Key gene be TORK1, NtTPK1 and CIPK23, selection and breeding and identification the efficient flue-cured tobacco cultivars of potassium.
The hereditary capacity of K+ efficiency is had a quantitative inheritance feature by controlled by multiple genes, TORK1 as stated above,
NtTPK1 and CIPK23, it is influenced by environmental conditions there are apparent gene interaction.It is absorbed if merely measured with potassium
With the relevant gene of transhipment, obtained result may be inconsistent with field experiment result.If crop field only is carried out to germ plasm resource
Verification, effect is very direct, but experimental work amount is very big, and the high kind of selected next K+ efficiency can under other ecological environments
It can there are inconsistent results.
The content of the invention
To solve hybrid screening approach time length, heavy workload, Primary structure method is by environmental influence, experiment
As a result may with field experiment result it is inconsistent the problem of, the present invention provide it is a kind of by seedling screening richness potassium tobacco mutant body,
The method of selection and breeding richness potassium tobacco bred.
The present invention is established on the basis of tablet culture technique, utilizes low potassium Screening of Media rich potassium volcanic rocks tobacco mutant
Body, the time is short, and tobacco seedling growth environment is consistent, determines the mainly genotype of its phenotype, and the later stage combines field experiment to result
Further verification, experimental result is more accurate, stablizes.According to existing result of study, low potassium tolerant mutant is not than low-kalium resistant mutation
Body potassium content of tobacco leaf significantly improves, and, strong operability time saving and energy saving compared with conventional method improves screening efficiency.
To achieve the above object, the technical solution adopted by the present invention is：
A kind of selection of richness potassium tobacco bred, comprises the following steps：
(1) seed selection and processing
Full consistent tobacco seed is selected first, then tobacco seed is impregnated into 3h in 50 DEG C of warm water；Through quality point
Number is 75% CH3CH2OH aqueous solutions handle 30s, are rinsed repeatedly with sterile water 5 times；Again with the hypochlorous acid that mass fraction is 10%
Aqueous solution handles 30min, is showed money or valuables one carries unintentionally with vernalization is carried out after aseptic water washing 5 times；
(2) preparation of low potassium culture medium and sowing culture
By step (1), treated that seed bud is seeded on low potassium culture medium, and often two row of disk, is put into phjytotron and trains
It supports, erects, daylight is 350 μm of ol photonsm-2·s-1, night illumination be 0, light dark period 14/10h, round the clock
Temperature is 28/25 DEG C, and relative air humidity is 60%~70%；
(3) the not screening of low-kalium resistant tobacco seedlings
After culture 30 days, the single plant of not low-kalium resistant is filtered out according to root long, and is classified as three grades, root long ＜
0.5cm is the first kind, and root long is the second class in 0.5~1.0cm, and root long ＞ 1.0cm are three classes, are transplanted into respectively in seed plate,
Normal potassium culture solution is supplied, crop field is transplanted to when growing to 6~7 true leaves；
(4) crop field is verified
30d, 40d, 50d impose potash fertilizer after transplanting, and every plant of 5g is configured to solution and spreads manuer in holes at stem 20cm, divides
80d, 110d divide single plant to choose the 12nd leaf position, the blade of the 18th leaf position, 110 DEG C of water-removings, 80 DEG C of drying, mill of weighing not after transplanting
Broken 60 mesh sieve of mistake, HCl extract-flame spectrophotometric determination potassium content, the high single plant of mark potassium content；
(5) isogeneity
The high single plant of step (4) mark potassium content is harvested into seed, is repeated the above steps, rear next year passes through identical method
Plantation is selected again, until genotype is homozygous, while measures the potassium content of cured tobacco leaf.
Preferably, treated that seed bud is seeded on normal potassium culture medium by step (1) for the step (2), is put into artificial gas
It waits and is cultivated in room, daylight is 350 μm of ol photonsm-2·s-1, night illumination is 0, light dark period 14/10h, daytime
Night temperature is 28/25 DEG C, and relative air humidity is 60%~70%, after cultivating 10 days, is transplanted on no potassium culture medium, often disk two
Row, root stand upside down placement upwards；
The screening of the step (3) not low-kalium resistant tobacco seedlings after culture 20 days, observes curved situation, according to curved root long and plant
Strain size filters out the single plant of not low-kalium resistant, and is classified as three grades, and it is first group not yet to occur curved, curved ＜
1cm is second group, and curved root long ＞ 1.0cm are the 3rd group, are transplanted into respectively in seed plate, supply normal potassium culture solution, grow to 6~
Crop field is transplanted to during 7 true leaves.
Preferably, the constituent of the low potassium culture medium includes 945mg/L Ca (NO3)2·4H2O, 425mg/L
NaNO3, 80mg/L NH4NO3, 20.4mg/L KH2PO4, 132.6mg/L NaH2PO4·2H2O, 493mg/L MgSO4·7H2O,
2.5ml/L EDTA-Fe solution, 5mg/L trace element solutions, agar powder 8g/L.
Preferably, the constituent of the normal potassium culture medium includes 945mg/L Ca (NO3)2·4H2O, 506mg/L
KNO3, 80mg/L NH4NO3, 136mg/L KH2PO4, 493mg/L MgSO4·7H2O, 2.5ml/L
EDTA-Fe solution, 5mg/L trace element solutions, agar powder 8g/L.
Preferably, the constituent of the no potassium culture medium includes 945mg/L Ca (NO3)2·4H2O, 425mg/L
NaNO3, 80mg/L NH4NO3, 156mg/L NaH2PO4, 493mg/L MgSO4·7H2O, 2.5ml/L
EDTA-Fe solution, 5mg/L trace element solutions, agar powder 8g/L.
Compared with prior art, beneficial effects of the present invention are：
(1) selection of the rich potassium tobacco bred of the present invention is established on the basis of tablet culture technique, is trained using low potassium
Foster base can be carried out at the same time screening to tens thousand of a variation types, and screening amount is big, obtain the time of rich potassium volcanic rocks tobacco mutant body
It is short, plantation verification is carried out to the seedling screened, it is as a result more accurate, stable.According to existing result of study, low-kalium resistant is not dashed forward
Variant is significantly improved than low potassium tolerant mutant potassium content of tobacco leaf, and, strong operability time saving and energy saving compared with conventional method, screening efficiency is normal
Thousands of times of rule method.
(2) selection of the present invention screens rich potassium mutation using the characteristic of rich potassium volcanic rocks not low-kalium resistant since seedling stage
Body single plant, and later stage progress crop field verification, considering comprehensively influences of both genotypic and environment, selected single plant tobacco leaf potassium
Content is high and stablizes.
(3) selection of the invention is easy to operate, is particularly suitable for the screening of high-volume Variants and germ plasm resource.
Description of the drawings
Fig. 1 is the flow chart of selection of the present invention.
Specific embodiment
With reference to embodiment, the present invention is described in further detail.
Embodiment 1
It is a kind of with low potassium Screening of Media not low-kalium resistant the present embodiment provides a kind of selection of rich potassium tobacco bred
Cigarette strain, and then the method for obtaining rich potassium tobacco bred, specifically include following steps：
The first step, seed selection and processing
Full consistent tobacco seed is selected first, then tobacco seed is impregnated into 3h in 50 DEG C of warm water；Through quality point
Number is 75% CH3CH2OH aqueous solutions handle 30s, are rinsed repeatedly with sterile water 5 times；Again with the hypochlorous acid that mass fraction is 10%
Aqueous solution handles 30min, and carrying out vernalization after aseptic water washing 5 times shows money or valuables one carries unintentionally, which may refer to He Wei, Yang Zhongyi,
Different flue-cured tobacco cultivars root growths and difference [J] Chinese tobacco journals of Gen Mao forms under the low potassium stress such as invention, 2010,
03:43-48。
In the present embodiment, the tobacco seed of immersion is 75%CH through mass fraction3CH2OH aqueous solutions handle the specific behaviour of 30s
It is as method：Tobacco seed is fitted into the centrifuge tube that volume is 2ml, adds in the CH that mass fraction is 75% thereto3CH2OH
Aqueous solution 1ml, shakes up, and impregnates 30s.
In the present embodiment, the concrete operation method for the hypochloric acid water solution treatment 30min for being 10% with mass fraction is：It will
Tobacco seed after aseptic water washing is fitted into the centrifuge tube that volume is 2ml, adds in the secondary chlorine that mass fraction is 10% thereto
Aqueous acid 1ml, shakes up, and impregnates 30min, since soaking time is longer, need to shake up for several times during immersion treatment.
Second step, low potassium culture medium configuration and sowing culture
In the present embodiment, the constituent of low potassium culture medium is as shown in table 1.
The constituent table of 1 low potassium culture medium of table
The constituent of normal potassium culture medium is as shown in table 2.
The constituent table of 2 normal potassium culture medium of table
Constituent without potassium culture medium is as shown in table 3.
Constituent table of the table 3 without potassium culture medium
In the present embodiment, EDTA-Fe solution and trace element solution are prepared to use and prepare both solution in the prior art
Method prepared, the EDTA-Fe in low potassium culture medium and normal potassium culture medium and trace element amount be respectively 2.5ml/L and
5ml/L meets the nutritional condition needed for plant production, when content is below or above the numerical value, can cause tobacco seedlings to other battalion
It supports the inhibition of element absorption or cannot fully meet the needs of Plant To Nutrient.
Sowing the detailed process cultivated is：
By the first step, treated that seed bud is seeded on low potassium culture medium, and often two row of disk, is put into phjytotron and cultivates,
It erects, daylight is 350 μm of ol photonsm-2·s-1, night illumination is 0, light dark period 14/10h, diurnal temperature
For 28/25 DEG C, relative air humidity is 60%~70%.
The screening of 3rd step, not low-kalium resistant tobacco seedlings
After culture 30 days, the single plant of not low-kalium resistant is filtered out according to root long, and is classified as three grades, root long ＜
0.5cm is the first kind, and root long is the second class in 0.5~1.0cm, and root long ＞ 1.0cm are three classes, are transplanted into respectively in seed plate,
Normal potassium culture solution is supplied, crop field is transplanted to when growing to 6~7 true leaves.Wherein, normal potassium culture solution is not add in normal potassium culture medium
Add agar powder, remaining ingredient is identical.
The single plant of not low-kalium resistant is screened in the present embodiment according to root long, root long can be measured accurately, and result of the test is more accurate
Really.
4th step, crop field verification
30d, 40d, 50d impose potash fertilizer after transplanting, and every plant of 5g is configured to solution and spreads manuer in holes at stem 20cm, divides
80d, 110d divide single plant to choose the 12nd leaf position, the blade of the 18th leaf position, 110 DEG C of water-removings, 80 DEG C of drying, mill of weighing not after transplanting
Broken 60 mesh sieve of mistake, HCl extract-flame spectrophotometric determination potassium content, the high single plant of mark potassium content.
5th step, isogeneity
The high single plant harvest seed of 4th step mark potassium content, repeats the above steps, rear next year passes through identical method kind
Plant is selected again, until genotype is homozygous, while measures the potassium content of cured tobacco leaf.
Potassium content distribution situation, strain number and this shared group are total in the blade of the 12nd leaf position of single plant in every group in the present embodiment
Several percentages are shown in Table 4.
The percentage of potassium content distribution situation, strain number and this shared group sum in the blade of the 12nd leaf position of 4 single plant of table
The consistent representative cigarette of four plants of growing ways is selected per class, root system is completely dug out, takes Rhizosphere Soil, survey root morphology
Parameter, root system potassium absorb index of correlation and rhizosphere root system secretion Accumulation of Organic Acids amount.
Wherein, the difference letter in each column after number represents the significant difference in 0.05 level.
Embodiment 2
The present embodiment is substantially the same manner as Example 1, except that
Second step, the detailed process for sowing culture are：
By the first step, treated that seed bud is seeded on normal potassium culture medium, is put into phjytotron and cultivates, daylight
For 350 μm of ol photonsm-2·s-1, night illumination is 0, and light dark period 14/10h, diurnal temperature is 28/25 DEG C, empty
Gas relative humidity is 60%~70%, after culture 10 days, is transplanted on no potassium culture medium, often two row of disk, root stands upside down placement upwards.
The screening of 3rd step, not low-kalium resistant tobacco seedlings
After culture 20 days, curved situation is observed, the single plant of not low-kalium resistant is filtered out according to curved root long and plant size, and will
It is divided into three grades, and it is first group not yet to occur curved, and curved ＜ 1cm is second group, and curved root long ＞ 1.0cm are the 3rd
Group is transplanted into seed plate respectively, supplies normal potassium culture solution, crop field is transplanted to when growing to 6~7 true leaves.
The principle of richness potassium tobacco bred selection of the invention is that rich potassium kind absorption potassium ability is strong, is had larger
Vmax values, but not low-kalium resistant, Cmin values are higher, after rich potassium tobacco mutant body is grown on low potassium culture medium, are in soon
Potassium deficiency state, influences root growth and development, potassium deficiency symptom occurs, and gained potassium deficiency single plant is verified using crop field, you can obtains richness
Potassium tobacco mutant body.
It although an embodiment of the present invention has been shown and described, for the ordinary skill in the art, can be with
Understanding without departing from the principles and spirit of the present invention can carry out these embodiments a variety of variations, modification, replace
And modification, the scope of the present invention is defined by the appended.
Claims (5)
1. a kind of selection of richness potassium tobacco bred, which is characterized in that comprise the following steps：
（1）Seed selects and processing
Full consistent tobacco seed is selected first, then tobacco seed is impregnated into 3h in 50 DEG C of warm water；It is through mass fraction
75% CH3CH2OH aqueous solutions handle 30s, are rinsed repeatedly with sterile water 5 times；Again with the hypochloric acid water solution that mass fraction is 10%
30min is handled, is showed money or valuables one carries unintentionally with vernalization is carried out after aseptic water washing 5 times；
（2）The preparation of low potassium culture medium and sowing culture
By step（1）Seed bud that treated is seeded on low potassium culture medium, and often two row of disk, is put into phjytotron and cultivates, and is erected
Vertical, daylight is 350 μm of ol photonsm-2·s-1, night illumination is 0, light dark period 14/10h, and diurnal temperature is
28/25 DEG C, relative air humidity is 60% ~ 70%；
（3）The not screening of low-kalium resistant tobacco seedlings
After culture 30 days, the single plant of not low-kalium resistant is filtered out according to root long, and is classified as three grades, root long ＜ 0.5cm are
The first kind, root long are the second class in 0.5 ~ 1.0cm, and root long ＞ 1.0cm are three classes, are transplanted into respectively in seed plate, supply normal potassium
Culture solution is transplanted to crop field when growing to 6 ~ 7 true leaves；
（4）It verifies in crop field
30d, 40d, 50d impose potash fertilizer after transplanting, every plant of 5g, are configured to solution and spread manuer in holes at stem 20cm, respectively at
80d, 110d divide single plant to choose the 12nd leaf position, the blade of the 18th leaf position, 110 DEG C of water-removings after transplanting, and 80 DEG C of drying are weighed milled
60 mesh sieves, HCl extract-flame spectrophotometric determination potassium content, the high single plant of mark potassium content；
（5）Isogeneity
By step（4）The single plant harvest seed that potassium content is high is marked, is repeated the above steps, rear next year is planted by identical method
It selects again, until genotype is homozygous, while measures the potassium content of cured tobacco leaf.
A kind of 2. selection of rich potassium tobacco bred according to claim 1, which is characterized in that the step（2）It will
Step（1）Seed bud that treated is seeded on normal potassium culture medium, is put into phjytotron and is cultivated, and daylight is 350 μm of ol
photons·m-2·s-1, night illumination is 0, and light dark period 14/10h, diurnal temperature is 28/25 DEG C, relative air humidity
It for 60% ~ 70%, after culture 10 days, is transplanted on no potassium culture medium, often two row of disk, root stands upside down placement upwards；
The step（3）The not screening of low-kalium resistant tobacco seedlings after cultivating 20 days, observes curved situation, big according to curved root long and plant
Small to filter out the single plant of not low-kalium resistant, and be classified as three grades, it is first group not yet to occur curved, curved ＜ 1cm's
For second group, curved root long ＞ 1.0cm are the 3rd group, are transplanted into respectively in seed plate, supply normal potassium culture solution, grow to 6 ~ 7 very
Crop field is transplanted to during leaf.
A kind of 3. selection of rich potassium tobacco bred according to claim 1 or 2, which is characterized in that the low potassium training
Supporting the constituent of base includes 945mg/L Ca (NO3)2·4H2O, 425mg/L NaNO3, 80mg/L NH4NO3, 20.4mg/L
KH2PO4, 132.6mg/L NaH2PO4·2H2O, 493mg/L MgSO4·7H2O, 2.5ml/L EDTA-Fe solution, 5mg/L are micro-
Secondary element solution, agar powder 8g/L.
A kind of 4. selection of rich potassium tobacco bred according to claim 1 or 2, which is characterized in that the normal potassium training
Supporting the constituent of base includes 945mg/L Ca (NO3)2·4H2O, 506mg/L KNO3, 80mg/L NH4NO3, 136mg/L
KH2PO4, 493mg/L MgSO4·7H2O, 2.5ml/L EDTA-Fe solution, 5mg/L trace element solutions, agar powder 8g/L.
A kind of 5. selection of rich potassium tobacco bred according to claim 1 or 2, which is characterized in that the no potassium training
Supporting the constituent of base includes 945mg/L Ca (NO3)2·4H2O, 425mg/L NaNO3, 80mg/L NH4NO3, 156mg/L
NaH2PO4, 493mg/L MgSO4·7H2O, 2.5ml/L EDTA-Fe solution, 5mg/L trace element solutions, agar powder 8g/
L。
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN111138519A (en) * | 2020-01-06 | 2020-05-12 | 河南农业大学 | Over-expression gene capable of improving potassium content of tobacco, and coding product and application thereof |
| CN116210506A (en) * | 2022-11-14 | 2023-06-06 | 广西南亚热带农业科学研究所 | Macadimia nut germplasm resource screening method |
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| CN106613924A (en) * | 2016-12-26 | 2017-05-10 | 上海烟草集团有限责任公司 | Method for screening and identifying potassium-rich tobacco |
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Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN111138519A (en) * | 2020-01-06 | 2020-05-12 | 河南农业大学 | Over-expression gene capable of improving potassium content of tobacco, and coding product and application thereof |
| CN111138519B (en) * | 2020-01-06 | 2022-03-18 | 河南农业大学 | Over-expression gene capable of improving potassium content of tobacco, and coding product and application thereof |
| CN116210506A (en) * | 2022-11-14 | 2023-06-06 | 广西南亚热带农业科学研究所 | Macadimia nut germplasm resource screening method |
| CN116210506B (en) * | 2022-11-14 | 2024-04-26 | 广西南亚热带农业科学研究所 | Macadimia nut germplasm resource screening method |
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Application publication date: 20180601 |