CN102577798A - Method for identifying salt tolerance of corn - Google Patents

Abstract

The invention relates to a method for identifying salt tolerance of corn, which comprise the following steps of: carrying out a multilayer filter paper method germination test on a variety to be authenticated; carrying out salt stress treatment by using 100-120 MMNaCl solution after germination; preliminarily judging the salt tolerance according to treated growing states; carrying out seedling state identification (a sand culture method), i.e. carrying out salt stress treatment when seedling is grown to a quatrefoil stage, wherein the salt stress is obvious when leaf blades are curled remarkably; sampling and measuring soluble sugar content in leaf blades of each variety; and comprehensively judging the salt tolerance of each variety. The method has the characteristics of accurate measuring result, high reliability, strong operability, short period, simplicity and convenience in operation, high efficiency and the like.

Description

A kind of corn salt resistance authentication method

Technical field

The present invention relates to agricultural technology field, be specifically related to a kind of corn salt resistance authentication method.

Background technology

The soil salinization is one of factor that has a strong impact on agricultural production and ecotope.The arable land in the whole world about 20% and half the nearly irrigateing land are influenced by salination.China has saline land 2,700 ten thousand hm now ², wherein about 7,000,000 hm ²Be the farmland, its area is still constantly enlarging now.According to estimates, China reaches tens billion of units because of the agriculture direct losses that cause in the saline land every year.Along with the high speed development of industry, the excessive exploitation in soil, the excessive use of chemical fertilizer, the expansion of irrational irrigation and facility cultivation area, secondary salinized soil area also continuing expansion, causes heavy losses to agricultural production.

Corn is the second largest crop of China, can be used as grain, feed and the raw material of industry etc., is the pillar of China's livestock breeding industry.Yet corn is to the salinity medium sensitivity, and salt resistance is relatively poor relatively, and this has greatly limited the expansion of its cultivated area.Report about the corn salt resistance is identified is less relatively; Recent Progress in Study on Salt Tolerance to corn also focuses mostly on aspect the Physiology and biochemistry mechanism; Relevant information people to corn germplasm and salt resistance understand seldom; Owing to lack research, seriously limited the genetic research of corn salt resistance and the seed selection of haloduric corn new varieties to corn salt tolerant germ plasm resource.And screening and cultivate the drought resistance and salt tolerance corn variety, for making full use of the salination soil, preserve the ecological environment, the conserve water resource, increasing farmers' income, ensure that national food safety is most important.

The crop salt tolerant alkali authentication method that adopts is at present concluded and can be divided into 4 types, and promptly evaluation is asked in laboratory, greenhouse, artificial field and natural field.Traditional salt tolerant alkalescence identifies that majority carries out under natural salt alkaline earth and the artificial field salt alkaline stress environment that is provided with; This method can be held the salt tolerant alkalescence of a large amount of germ plasm resources and identify; But, influence accuracy of experimental results because the saline and alkaline stress conditions in field is difficult for reaching consistent.Greenhouse pot culture is identified and the salinization of soil nutrient solution is identified, can be used for the crop salt tolerant alkalescence evaluation in the time of infertility, but often identifies limited amount, has certain limitation.

Summary of the invention

The technical problem that the present invention will solve provide a kind of result accurately, strong operability, be easy to the corn salt resistance authentication method that carries out in batches.

For solving the problems of the technologies described above, the technical scheme that the present invention adopts is:

A kind of corn salt resistance authentication method may further comprise the steps:

(1) germination test: in clean culture dish, the three metafiltration paper that tile, it is moistening to pour distilled water into; Water is submergence filter paper 0.3cm approximately, then corn seed hilum court is placed down on the filter paper 5～10 seeds of every ware; Put in the artificial culture case and cultivate down in 28 ℃, the salt processing was carried out with 100～120MMNaCl solution in the back of germinateing in three days, established blank simultaneously; Measure the germination length that each is handled after six days, its length and its contrast differ more little, can judge tentatively that then its salt resistance ability is strong more;

(2) identify seedling stage: user's opaque container, the fine sand of packing into accounts for container 2/3; Broadcast into the same corn seed of step (1), cover fine sand then, watering makes sandy soil drenched fully; 6 seeds of sowing in each container are carried out at temperature and cultivate down for 30 ℃ in the greenhouse, treating to grow to four leaves by seedling, to get blade appearance during the phase subsequent use; Carry out salt stress simultaneously and handle (and establish and do not carry out the blank that salt is handled), after seven days, treat that obviously curling appears in blade is that salt stress is when obvious; Get blade appearance, measure its soluble sugar content, the processing salt resistance ability that soluble sugar content is low in the blade is high.

Said soluble sugar assay method is an anthrone method.

In said step (2), salt stress used salt solution is 100～120MM NaCl solution.

In said step (2), salt stress was handled after seven days, took out whole plant; Get fine sand and dust express developed with deionized water, blot the deionized water of plant remained on surface, then plant is put into baking oven with 105 ℃ of 10 min that complete with blotting paper; Dry to constant weight for 80 ℃, weigh then, by formula: dry weight rate of change (%)=[(contrast-processing)/contrast] * 100%; Calculate the dry weight rate of change, and confirm the salt tolerant grade of each material with criterion:

Rank	Salt resistance	Dry weight rate of change (%)
			1	High anti-	0～20.0
2	Salt tolerant	20.1～40.0
			3	In anti-	40.1～60.0
4	Responsive	60.1～80.0
			5	High sense	80.1～100

According to the growth of cereal crop seedlings corn Seedling Salt-tolerance is fallen into 5 types:

1 grade is high anti-kind: it is normal basically to grow, and does not have the symptom of obviously being injured, and 3～4 greenery are arranged; 2 grades is the salt tolerant kind: it is normal basically to grow, and 3 greenery are arranged, and the blade flavescence or the blade tip green grass or young crops of the 1st true leaf are withered; 3 grades be in anti-kind: the growth suppressed, 2 greenery are arranged, the blade flavescence or the blade tip green grass or young crops of the 1st, 2 true leaf are withered; 4 grades are sensitive varieties: seriously be injured, 1 greenery or only lobus cardiacus survival are only arranged; 5 grades is high sense kind: whole plant tangerine is dead or approaching dead.

The present invention has actively useful effect:

Have that test result is accurate, reliability is high, strong operability, short, easy and simple to handle, efficient advantages of higher of cycle.

Embodiment

Further set forth the present invention below in conjunction with specific embodiment.Assay method among the following embodiment or inspection method are conventional method if no special instructions, and material therefor and reagent are commercially available if no special instructions.

1 one kinds of corn salt resistances of embodiment authentication method may further comprise the steps:

(1) germination test (multilayer filter paper method): in clean culture dish, the three metafiltration paper that tile, it is moistening to pour distilled water into, and water is submergence filter paper 0.3cm approximately, then experiment material is placed on (hilum of seed is down) on the filter paper, 5～10 seeds of every ware.

Material to be identified (seed) comprises salt tolerant strain and check clone (non-salt tolerant), and is as shown in table 1, each material 2 ware, and 10 in every ware is established salt and is handled and blank.Experimental situation is artificial incubator, temperature 28.After germinateing in three days, carry out salt stress and handle (using 100MMNaCl solution), and need to change filter paper and salting liquid every day, and establish blank.Add up germination after six days, it is long to measure the bud seedling, referring to table 2.

Table 1 test material

Table 2 different disposal bud seedling growing state

From table 2, can find out: salt is handled and blank seed growth situation significant difference; 2011ZW39,2011ZW31,2011ZW30 salt resistance ability reduce successively; Basically identical as a result between two repetitions.

(2) identify (Sha Peifa) seedling stage: utilize the seed of germination test to identify or carry out Sha Peifa separately.Sha Peifa user opaque container, the fine sand of packing into accounts for container 2/3, broadcasts into seed, covers fine sand then, pours into running water and makes sandy soil drenched fully, 6 seeds of sowing in each container, two repetitions of each material.Put in the greenhouse in 30 ℃ and cultivate down, treat that seedling grows to four leaves and take a sample during the phase and carry out salt stress (100MM Nacl solution) simultaneously, after seven days, obviously curling appears in blade is that salt stress is obvious, and laboratory detection leaf soluble is taken back in sampling rapidly.

Soluble sugar detects: gets blade 0.1g, grinds fully, add 5～10ml distilled water and go to 20ml in vitro, and boiling water bath 20min, cooled and filtered is gone in the 25ml volumetric flask, washes test tube repeatedly and residue is settled to scale.Get the 1ml extract in the 20ml test tube, add 1ml distilled water, 0.5ml anthrone ethyl acetate slowly adds the 5ml concentrated sulfuric acid again, shakes up boiling water bath 10min, 620nm place colorimetric record OD.

Soluble sugar calibration curve equation: Y=0.0119X-0.0036, R ²=0.9988

Sugar content (%)=C * V/A * n/W * 10 ⁶

In the formula: C be the calibration curve amount of finding ( μ g);

A is for drawing sample liquid volume (ml);

V is for extracting liquid measure (ml);

N is an extension rate;

W is tissue weight (g)

Result of calculation is as shown in table 3.

Before handling, table 3 salt stress handles back maize leaf soluble sugar content with salt stress

From table 3, can find out: soluble sugar content no significant difference before salt stress is handled in 2011ZW39,2011ZW31, the 2011ZW30 experiment material blade; Salt stress is handled back salt tolerant strain soluble sugar content and is reduced successively, and promptly the salt resistance ability of 2011ZW39,2011ZW31,2011ZW30 reduces successively.This result and germination test result coincide, confirmation each other, and reliability greatly improves.

Though, the present invention has been done detailed description in the preceding text with general explanation and specific embodiments, on basis of the present invention, can to some modifications of do or improvement, this will be apparent to those skilled in the art.Therefore, these modifications or improvements all belong to the scope of protection of present invention without departing from theon the basis of the spirit of the present invention.

Claims (4)

1. corn salt resistance authentication method may further comprise the steps:

(1) germination test: in clean culture dish; Three metafiltration paper tile; It is moistening to pour distilled water into, and water logging there is not filter paper 0.3cm, then corn seed hilum court is placed down on the filter paper; Put in the ARTIFICIAL CULTURE case and cultivate down in 28 ℃; Carry out salt stress with 100～120MMNaCl solution after germinateing in three days and handle, establish blank simultaneously, measure germination length that each is handled after six days; Its length and its contrast differ more little, can judge tentatively that then its salt resistance ability is strong more;

(2) identify seedling stage: the fine sand of in culture vessel, packing into, broadcast into the same corn seed of step (1), cover fine sand then; Watering makes sandy soil drenched fully, and 6 seeds of sowing in each container are carried out at temperature and cultivate down for 30 ℃ in the greenhouse; Treating to grow to four leaves by seedling, to get blade appearance during the phase subsequent use, carries out salt stress simultaneously, after seven days; Treat that obviously curling appears in blade is salt stress when obvious, get blade appearance, measure its soluble sugar content; The processing salt resistance ability that soluble sugar content is low in the blade is high, in conjunction with the result of the test of said step (1), accurately judges the salt resistance of each kind.

2. corn salt resistance authentication method according to claim 1 is characterized in that said soluble sugar assay method is an anthrone method.

3. corn salt resistance authentication method according to claim 1 is characterized in that, in said step (2), salt stress used salt solution is 100～120MM NaCl solution.

4. corn salt resistance authentication method according to claim 1 is characterized in that, in said step (2); Salt stress was handled after seven days, took out whole plant, got fine sand and dust express developed with deionized water; Blot the deionized water of plant remained on surface with blotting paper, then plant is put into baking oven with 105 ℃ of 10 min that complete, dry to constant weight for 80 ℃; Weigh then; By formula: dry weight rate of change (%)=[(contrast-processing)/contrast] * 100%, calculate the dry weight rate of change, and confirm the salt resistance grade of each kind or strain with criterion:

Rank Salt resistance Dry weight rate of change (%) 1 High anti- 0～20.0 2 Salt tolerant 20.1～40.0 3 In anti- 40.1～60.0 4 Responsive 60.1～80.0 5 High sense 80.1～100

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