CN101897253A - Method for identifying seedling stage stress resistance of tobacco variety - Google Patents
Method for identifying seedling stage stress resistance of tobacco variety Download PDFInfo
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Abstract
The invention discloses a method for identifying the seedling stage stress resistance of a tobacco variety. The method comprises the following steps of: disinfecting seeds of the tobacco variety to be tested, germinating the disinfected seeds under adverse condition and normal condition for 16 days and calculating a germination percentage and a germination index; germinating the seeds under normal condition for 7 days, allowing the buds to grow under the adverse condition for 5 days, allowing the buds to grow under normal condition for 4 days, allowing the buds to grow under normal condition for 16 days and testing the heights and dry weights of seedlings; and taking relative values of the germination percentage, the germination index, the seedling heights and the dry weights of the seedlings as indexes for evaluating the seedling stage stress resistance of the tobacco variety, setting a property relative value to be xL/xN, performing cluster analysis on the property relative value by a group average method and determining the seedling stage stress resistance of the tobacco variety to be tested according to the average value of property relative values. The method has the advantages of simple and convenient operation, low cost, rapid identification, easy operation, stable and accordant result, capability of eliminating the difference between the genetic backgrounds of different varieties due to the adoption of the property relative value as the index for evaluating the seedling stage stress resistance of the variety and good application prospect.
Description
Technical field
The invention belongs to tobacco bred resistance authenticate technology field, be specifically related to a kind of authentication method of tobacco bred seedling stage stress resistance.
Background technology
Tobacco originates in the subtropics, be the warm crop of happiness, but big cigarette district temperature during floating seedlings of China is lower, and often meet bad weathers such as sleet is freezing, cause temperature of fry nursing pool lower, even can cause larval rearing water to freeze, formed low temperature stress tobacco seed and seedling.Simultaneously, because the supply of moisture and nutrient is to transport from the bottom up by capillarity, the evaporation of moisture makes a large amount of accumulation of salinity in stromal surface, thereby causes the medium of seedling bed surface saltingization, has formed the salt stress to tobacco seed and seedling.In addition, mountain area, China part hills is still using the tobacco usual manner to grow seedlings at present, and these regional soil are prone to water deficit, have formed the drought stress to tobacco seed and seedling.
Tobacco in seedling stage low temperature stress, salt stress and drought stress have caused the emergence rate of tobacco seed-coating pelleted seed to reduce, it is irregular to emerge, growth of seedling is slow, a little less than the growth potential, even do not emerge or seedling yellow, death, have a strong impact on normal leaf tobacco production, brought the tremendous economic loss for country and tobacco grower, become the outstanding problem in the present leaf tobacco production.Seed selection multiresistance kind is to reduce the effective way of low temperature to the leaf tobacco production influence, but breeding of new variety, to promote the time cycle longer, can not solve the existing problems in the present leaf tobacco production fast.Resistance (cold resistance, salt-resistance and drought resistance) field appraisal cost is higher, is subjected to natural environment influence bigger, and repeatability is relatively poor, easily causes the result inaccurate.Therefore, how tobacco bred is carried out cold-resistant, anti-salt in indoor seedling stage, quick, the accurately evaluation of drought resistance, just become the technical problem that needs to be resolved hurrily in the present leaf tobacco production.
Summary of the invention
The objective of the invention is to the problem that exists in the current leaf tobacco production, a kind of method that can fast, accurately identify the tobacco bred seedling stage stress resistance is provided.
Purpose of the present invention is achieved by the following technical programs.
Except as otherwise noted, percentage of the present invention is percetage by weight.
A kind of authentication method of tobacco bred seedling stage stress resistance may further comprise the steps:
1, seed disinfection
The seed of tobacco bred to be measured with 1.0% copper-bath soaking disinfection 15min, is taken out seed and cleans with clear water, and at room temperature Hui Gan is to water content 4.0～7.0%, and is standby;
2, seed sprouting
Tobacco seed through sterilization is placed under the adverse environmental factor illumination every day 12h, 3 repetitions, 100 seeds of every repetition; Simultaneously, 3 repetitions are set, place under the normal condition, illumination every day 12h, 100 seeds of every repetition.Write down the chitting piece number every day, 16d statistics germination rate and germination index, germination rate (%)=(subnumber is planted experimentally in normal seeds germinated number/confession in 16 days) * 100%, germination index=∑ (Gt/Dt), Gt is the germinative number at different time in the formula, and Dt is the germination number of days;
3, growth of seedling
Tobacco seed after the sterilization is placed the 7d that germinates under the normal condition, change growth 5d under the adverse environmental factor then over to, change regrowth 4d under the normal condition afterwards again over to, illumination every day 12h, 3 repetitions, 100 seeds of every repetition; Simultaneously, 3 repetitions are set, place the 16d that grows under the normal condition, illumination every day 12h, 100 seeds of every repetition.Picked at random 50 strain seedling are measured seedling height; Seedling is dried 24h down at 80 ℃, measure the seedling dry weight;
4, the calculating of tobacco bred resistance evaluation index
Adopt relative germination rate, relative germination index, relative height of seedling, relative seedling dry weight as the index of estimating the kind seedling stage stress resistance, proterties relative value=x
L/ x
N, in the formula, x
LBe each character value that records under the adverse environmental factor, x
NBe each character value that records under the normal condition;
5, cluster analysis
Adopt Euclidean distance, the proterties relative value is carried out cluster analysis with the class method of average,, determine the seedling stage stress resistance of tobacco bred to be measured again according to the size of the mean value of each proterties relative value.
Described normal condition is: seed or seedling evenly place the germination box that is lined with wet filter paper, place incubator, germinate under 25 ℃ of conditions, grow.
Described adverse environmental factor is a kind of in low temperature stress, salt stress and the drought stress.
Described low temperature stress condition is: seed or seedling evenly place the germination box that is lined with wet filter paper, place incubator, in 11 ℃ of occurs at low temperatures buds, growth.
Described condition of salt stress is: seed or seedling evenly place the germination box that is lined with filter paper, and adding concentration is the sodium chloride solution of 10g/L, places incubator, germinate under 25 ℃ of conditions, grow.
Described drought stress condition is: seed or seedling evenly place the germination box that is lined with filter paper, and adding concentration is the polyglycol solution of 150g/L, places incubator, germinate under 25 ℃ of conditions, grow.
The present invention is easy and simple to handle, and is with low cost, identify fast.The indoor identification easy operating, temperature condition can artificially be controlled, as a result stable and consistent; Adopt the proterties relative value as the index of estimating the kind seedling stage stress resistance, can eliminate the difference of different cultivars genetic background.Have a good application prospect.
Description of drawings
Fig. 1 is the cluster analysis dendrogram of 5 tobacco breds in the Application Example.
Embodiment
By specific embodiment given below, can further clearly understand the present invention.But they are not limitation of the invention.
Embodiment 1
The seed of tobacco bred to be measured with 1.0% copper-bath soaking disinfection 15min, is taken out seed and cleans with clear water, and at room temperature Hui Gan is to water content 4.0～7.0%, and is standby.Tobacco seed through sterilization is placed the germination box that is lined with wet filter paper uniformly, place 11 ℃ of incubators, illumination every day 12h, 3 repetitions, 100 seeds of every repetition; Simultaneously, 3 repetitions are set, place 25 ℃ of incubators, illumination every day 12h, 100 seeds of every repetition.Write down the chitting piece number every day, 16d statistics germination rate and germination index, germination rate (%)=(subnumber is planted experimentally in normal seeds germinated number/confession in 16 days) * 100%, germination index=∑ (Gt/Dt), Gt is the germinative number at different time in the formula, and Dt is the germination number of days.
Tobacco seed after the sterilization is placed the germination box that is lined with wet filter paper uniformly, place 25 ℃ of incubators 7d that germinates, change 11 ℃ of growths of low temperature 5d then over to, again change 25 ℃ of following regrowth 4d afterwards over to, illumination every day 12h, 3 repetitions, 100 seeds of every repetition; Simultaneously, 3 repetitions are set, place 25 ℃ of incubators 16d that grows, illumination every day 12h, 100 seeds of every repetition.Picked at random 50 strain seedling are measured seedling height; Seedling is dried 24h down at 80 ℃, measure the seedling dry weight.
Adopt relative germination rate, relative germination index, relative height of seedling, relative seedling dry weight as estimating kind cold resistant index in seedling stage, proterties relative value=x
L/ x
N, in the formula, x
LBe each character value that records under the low temperature, x
NIt is each character value that records under 25 ℃.Adopt Euclidean distance, the proterties relative value is carried out cluster analysis with the class method of average,, determine the cold resistance in seedling stage of tobacco bred to be measured again according to the size of the mean value of each proterties relative value.
Embodiment 2
Repeat embodiment 1, following difference is arranged: the adverse environmental factor of employing is that seed or seedling evenly place the germination box that is lined with filter paper, and adding concentration is the sodium chloride solution of 10g/L, places incubator, germinates under 25 ℃ of conditions, grows.
Embodiment 3
Repeat embodiment 1, following difference is arranged: the adverse environmental factor of employing is that seed or seedling evenly place the germination box that is lined with filter paper, and adding concentration is the polyglycol solution of 150g/L, places incubator, germinates under 25 ℃ of conditions, grows.
Application Example
Select MS cloud and mist 85, MS cloud and mist 87, MSk326, cloud and mist 203, the main tobacco bred of planting in 5 whole nations of the big gold dollar of safflower for use, each kind skewer is at random got the 1g seed, with 1.0% copper-bath soaking disinfection 15min, taking out seed cleans with clear water, at room temperature Hui Gan is to water content 4.0～7.0%, and is standby.
Tobacco seed through sterilization is placed the germination box that is lined with two-layer wet filter paper uniformly, place 11 ℃ of incubators, illumination every day 12h, 3 repetitions, 100 seeds of every repetition.Simultaneously, 3 repetitions are set, 100 seeds of every repetition place 25 ℃ of incubators, illumination every day 12h.Write down the chitting piece number every day, 7d calculates germination vigor, 16d adds up germination rate, average germinating time and germination index, germination vigor (%)=(subnumber is planted experimentally in the normal seeds germinated number of 7d/confession) * 100%, germination rate (%)=(subnumber is planted experimentally in the normal seeds germinated number of 16d/confession) * 100%, and average germinating time (h)=∑ (Gt * Dt)/germination rate, germination index=∑ (Gt/Dt), Gt is the germinative number at different time in the formula, and Dt is the germination number of days.
Tobacco seed after the sterilization is placed the germination box that is lined with two-layer wet filter paper uniformly, place 25 ℃ of incubators 7d that germinates, change 11 ℃ of growths of low temperature 5d then over to, change 25 ℃ of following regrowth 4d afterwards again over to, illumination every day 12h.Simultaneously, 3 repetitions are set, 100 seeds of every repetition place 25 ℃ of incubators 16d that grows, illumination every day 12h.Picked at random 50 strain seedling, it is long to measure seedling total length, seedling height (overground part height) and root.Seedling is dried 24h down at 80 ℃, measure the seedling gross dry weight.
Adopt the proterties relative value as estimating the cold resistant index of kind.Proterties relative value=x
L/ x
N, in the formula, x
LBe each character value that records under the low temperature, x
NBe each character value that records under the normal temperature.Then, according to Euclidean distance, the proterties relative value is carried out cluster analysis with the class method of average.
The result shows that under normal temperature (25 ℃), the germination rate of 5 tobacco breds all reaches more than 90% (table 1).Except the big gold dollar of safflower, low temperature (11 ℃) adverse circumstance has all significantly reduced the germination rate of all the other kinds.Low temperature has prolonged the average germinating time of all kinds, has reduced germination index.From relative germination rate, the big gold dollar cold resistance of safflower is stronger, and the MSk326 cold resistance is the most weak.Just relatively average germinating time sees that the big gold dollar time of safflower is the shortest, but only significantly is lower than MS cloud and mist 85, and with its excess-three kind there was no significant difference.With regard to relative germination index, the big gold dollar of safflower is significantly higher than all the other 4 kinds, and MSk326 is minimum.
The variation of tobacco seed germination characteristic under the table 1 low temperature adverse circumstance
Annotate:
*Lowercase represent between different cultivars significant difference (α=0.05, LSD).
Under low temperature stress, the height of seedling of each kind, the long seedling (table 2) that all is lower than normal growth of root.Just height of seedling sees that the big gold dollar of MS cloud and mist 87 and safflower is significantly higher than its excess-three kind relatively; But just root length sees not have significant difference between each kind relatively.Under the low temperature, the performance and the height of seedling of seedling total length are similar; Except that the big gold dollar of safflower, all the other kind seedling dry weights all reduce (table 3), and wherein the MSk326 dry weight is 60% of a normal seedling dry weight.With regard to relative dry weight, the cold resistance of the big gold dollar of safflower is stronger, and the cold resistance of MSk326 is the most weak.
Tobacco height of seedling, the long variation of root under the table 2 low temperature adverse circumstance
Annotate:
*Lowercase represent between different cultivars significant difference (α=0.05, LSD).
The variation of tobacco seedling total length, dry weight under the table 3 low temperature adverse circumstance
Annotate:
*Lowercase represent between different cultivars significant difference (α=0.05, LSD).
Correlation analysis shows that two pairwise correlations between relative germination rate, relative germination index, relative height of seedling, relative 4 indexs of seedling dry weight reach significantly or utmost point significance level, and the strong and weak basically identical (table 4) as a result of the cold resistance that is obtained by these 4 indexs is described.The result who obtains according to these four indexs shows that the big gold dollar of safflower is cold-resistant kind, and MSk326 is a low-temperature sensitive type kind, and the cold hardiness of its excess-three kind falls between.
Correlation coefficient between each proterties relative value of table 4 tobacco bred
Annotate:
*With
*Be illustrated respectively on P＜0.05 and P＜0.01 level remarkable.
With germination rate, average germinating time, germination index, height of seedling, root length, seedling total length, the seedling dry weight of each kind under low temperature and contrast, and the relative value of these 7 indexs totally 21 indexs, carry out cluster analysis.The result shows, when the genetic distance threshold value gets 15, can be divided into 3 classes (table 5).Class I is the big gold dollar of cold resistant variety safflower, germination rate height under the low temperature adverse circumstance, and average germinating time is short, and the seedling growing way is better, and its gross dry weight is not subjected to low temperature effect (table 6) substantially.Class II is medium cold resistant variety, comprises MS cloud and mist 85, MS cloud and mist 87 and cloud and mist 203.In such, occurs at low temperatures bud rate reduces, and the seedling effect length is less, and dry weight suitably reduces.Class III is MSk326, and occurs at low temperatures bud rate, germination index reduce significantly, and height of seedling reduces bigger, and dry weight significantly reduces, and dry-matter accumulation is received very big inhibition.This result and correlation analysis be basically identical as a result.
The average relative value of each proterties of the different cold hardiness tobacco bred of 3 classes after table 5 cluster
Claims (6)
1. the authentication method of a tobacco bred seedling stage stress resistance may further comprise the steps:
(1) seed disinfection
The seed of tobacco bred to be measured with 1.0% copper-bath soaking disinfection 15min, is taken out seed and cleans with clear water, and at room temperature Hui Gan is to water content 4.0～7.0%, and is standby;
(2) seed sprouting
Tobacco seed through sterilization is placed under the adverse environmental factor illumination every day 12h, 3 repetitions, 100 seeds of every repetition; Simultaneously, 3 repetitions are set, place under the normal condition, illumination every day 12h, 100 seeds of every repetition.Write down the chitting piece number every day, 16d statistics germination rate and germination index, germination rate (%)=(subnumber is planted experimentally in normal seeds germinated number/confession in 16 days) * 100%, germination index=∑ (Gt/Dt), Gt is the germinative number at different time in the formula, and Dt is the germination number of days;
(3) growth of seedling
Tobacco seed after the sterilization is placed the 7d that germinates under the normal condition, change growth 5d under the adverse environmental factor then over to, change regrowth 4d under the normal condition afterwards again over to, illumination every day 12h, 3 repetitions, 100 seeds of every repetition; Simultaneously, 3 repetitions are set, place the 16d that grows under the normal condition, illumination every day 12h, 100 seeds of every repetition.Picked at random 50 strain seedling are measured seedling height; Seedling is dried 24h down at 80 ℃, measure the seedling dry weight;
(4) calculating of tobacco bred resistance evaluation index
Adopt relative germination rate, relative germination index, relative height of seedling, relative seedling dry weight as the index of estimating the kind seedling stage stress resistance, proterties relative value=x
L/ x
N, in the formula, x
LBe each character value that records under the adverse environmental factor, x
NBe each character value that records under the normal condition;
(5) cluster analysis
Adopt Euclidean distance, the proterties relative value is carried out cluster analysis with the class method of average,, determine the seedling stage stress resistance of tobacco bred to be measured again according to the size of the mean value of each proterties relative value.
2. the authentication method of a kind of tobacco bred seedling stage stress resistance according to claim 1, it is characterized in that: described normal condition is: seed or seedling evenly place the germination box that is lined with wet filter paper, place incubator, under 25 ℃ of conditions, germinate, grow.
3. the authentication method of a kind of tobacco bred seedling stage stress resistance according to claim 1 is characterized in that: described adverse environmental factor is a kind of in low temperature stress, salt stress and the drought stress.
4. the authentication method of a kind of tobacco bred seedling stage stress resistance according to claim 3, it is characterized in that: described low temperature stress condition is: seed or seedling evenly place the germination box that is lined with wet filter paper, place incubator, in 11 ℃ of occurs at low temperatures buds, growth.
5. the authentication method of a kind of tobacco bred seedling stage stress resistance according to claim 3, it is characterized in that: described condition of salt stress is: seed or seedling evenly place the germination box that is lined with filter paper, adding concentration is the sodium chloride solution of 10g/L, place incubator, under 25 ℃ of conditions, germinate, grow.
6. the authentication method of a kind of tobacco bred seedling stage stress resistance according to claim 3, it is characterized in that: described arid temperature stress conditions is: seed or seedling evenly place the germination box that is lined with filter paper, adding concentration is the polyglycol solution of 150g/L, place incubator, under 25 ℃ of conditions, germinate, grow.
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| CN109287192A (en) * | 2018-12-01 | 2019-02-01 | 中国农业科学院棉花研究所 | A kind of method of cotton seeds Identification of Drought |
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Open date: 20101201 |