CN107290395A - A kind of cold-resistant evaluation of lily and screening technique and the application of cold-resistant variety - Google Patents
A kind of cold-resistant evaluation of lily and screening technique and the application of cold-resistant variety Download PDFInfo
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- CN107290395A CN107290395A CN201710427350.XA CN201710427350A CN107290395A CN 107290395 A CN107290395 A CN 107290395A CN 201710427350 A CN201710427350 A CN 201710427350A CN 107290395 A CN107290395 A CN 107290395A
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Abstract
The invention discloses a kind of cold-resistant evaluation of lily and method and the application of cold-resistant variety screening, belong to agricultural high-effiency production technology.Climax leaves, as material, are cut into a certain size leaf block, and leaf block is pre-processed by this method using second first climax leaves of wheel below lily plant strain growth point.Afterwards, leaf block is handled using low-temperature circulating instrument, then determines relative conductivity.Finally, the semilethal low temperature (LT of each lily cultivar is calculated using statistical software50), lily cultivar cold resistance is evaluated according to semilethal low temperature, and filter out cold resistance lily cultivar.With it is traditional using lily ball or tissue-cultured seedling compared with the cold-resistant evaluation method of material, this method can more react lily cold resistance evaluation result the actual cold resistance in lily field, more representative than conventional method.
Description
Technical field
The invention mainly relates to lily physiology field, and in particular to a kind of cold-resistant evaluation of lily and the screening of cold-resistant variety
Method and application, this method mainly carry out low-temperature treatment by the blade to lily vegetative growth phase, by measuring blade conductance
Rate obtains relative conductivity Fitting Logistic regression equation, and then obtains the semilethal low temperature (LT of each kind of lily50),
Lily cultivar cold resistance is evaluated, and filters out cold resistance lily cultivar.
Technical background
Lily is the perennial flowering bulb plant of Liliaceae (Liliaceae).Usually said lily refers to lilium
(Lilium spp.) plant is various and general name of kind.Lily flower is very large, pattern is gorgeous, spend appearance different expressions, fragrance pleasant, makees
Cut-flower, potted flower can be done for important ornamental plant, can be applied again in Garden Greenland, while some lily cultivar edibles and medicine
With depth is liked by vast to consumer.At present, lily turns into one of big Fresh Cutting flower in the world five, and in the world, Market of Fresh Cut Flower is accounted for
There is highly important status.In recent years, the new varieties that lily was cultivated are more and more, and the area of China plantation lily is also year by year
It is rapid to expand.But lily is harsher to temperature requirement in growth course, it can then stop higher than 30 degrees Celsius or less than 5 degrees Celsius
Only grow.In Chinese Middle-north Area, winter-spring season temperature is low, although most of produce uses heliogreenhouse, but due to
Overall base is weak, its illumination, temperature and controllability is poor therefore winter-spring season low temperature has turned into northern area lily facility and planted
The restraining factors of training.Therefore, the cold-resistant lily of seed selection is carried out cultivating particularly necessary in production, and the cold-resistant lily cultivar premise of seed selection is needed
Set up the technical system of a set of evaluation lily cultivar cold resistance.At present, in the screening of lily cultivar cold resistance, mainly pass through
Low-temperature treatment is carried out to lily ball and tissue-cultured derived plant lily seedling, by determining electrical conductivity under low temperature stress, MDA, POD, protection
Enzymatic activity etc. screens lily cold-resistant variety, but these methods have some in drawback, and such as bulb low-temperature treatment screening is in screening
During the processing of each kind will consume substantial amounts of bulb, when kind is more, even more waste time and energy；Tissue-cultured seedling low-temperature treatment
High is required to the growing state of seedling, and tissue-cultured seedling growth cycle is long, if being even more to waste time and energy since tissue culture；This
Outside, often with lily plant field cold resistance there is larger difference in the cold-resistant evaluation result to lily ball and tissue-cultured seedling.Therefore,
The screening system for setting up a lily cultivar that is simple and convenient and meeting needs of production again is urgent problem to be solved.
The content of the invention
It is an object of the invention to provide a screening side that is simple and meeting the lily cold-resistant variety needed for actual production again
Method.Second first leaf of wheel is won below lily plant strain growth point first, cleans up rear Transverse Shear into after fragment and pretreatment
Processing in low-temperature circulating instrument is put into, is extracted every two days, then measures before boiling and boils rear electrical conductivity and (freeze electrical conductivity and boil
Boil electrical conductivity), relative conductivity is obtained, semilethal low temperature (LT is calculated50).It is cold-resistant to lily different cultivars according to semilethal low temperature
Property height be ranked up, filter out resistance and the weak kind of resistance.
The purpose of the present invention is achieved through the following technical solutions
A kind of screening technique of lily cold-resistant variety, comprises the following steps：
(1) second first climax leaves of wheel below lily plant strain growth point are won and are cleaned up；
(2) climax leaves after cleaning are cut into leaf block, and leaf block is pre-processed；
(3) pretreated leaf block is handled using low-temperature circulating instrument；
(4) sample after processing terminates in low-temperature circulating instrument is taken out into defrosting, the sample extraction after defrosting is stayed overnight；
(5) survey leaching liquor frost electrical conductivity, boiled after having surveyed and cool down again measure electrical conductivity obtain boiling electrical conductivity,
According to formula relative conductivity ,=frost electrical conductivity/boils electrical conductivity × 100%, obtains relative conductivity；
(6) it is with Fitting Logistic regression equation：Y=K/ (1+ae-bx) is analyzed relative conductivity, wherein Y
Cell damage rate is represented, x represents treatment temperature, and K is the saturated capacity of cell damage rate, and a, b is equation parameter；In order to determine a,
B value, linearization process is carried out by equation, and ln [(K-y)/y]=lna-bx makes y=ln [(k-y)/y], is then converted into cell
The linear equation of injury rate (Y) and treatment temperature (x)；A, b values and coefficient R are tried to achieve by the method for linear regression2, calculate
The semilethal low temperature LT of lily cultivar50=In [(1/a)]/b；
(7) lily cultivar cold resistance is evaluated according to semilethal low temperature, and filters out cold resistance lily cultivar.
The mistake pre-processed described in step (2) is referred to as described leaf block uniformly wrapping up 1 layer with wet gauze, is put into glass examination
Guan Zhong, is placed in 4 DEG C of refrigerator overnights.
It is using the process of low-temperature circulating instrument processing leaf block in step (3)：, will when low-temperature circulating instrument temperature drops to 4 DEG C
Pretreated sample is put into processing in low-temperature circulating instrument；The low-temperature circulating instrument treatment temperature be 5 gradients 0, -3, -6, -
9、-12℃；Each gradient processing 1h, temperature fall time is 30min between gradient and gradient.
The sample taking-up after processing terminates in low-temperature circulating instrument is put into defrosting in 4 DEG C of refrigerators in step (4), will be thawed
Sample afterwards takes out extraction and stayed overnight.
The time boiled described in step (5) is 15min.
Lily plant strain growth stalwartness, no disease and pests harm infection described in step (1).Same lily cultivar second takes turns first
Climax leaves are in the same size.
Climax leaves described in step (2) are laterally cut into a width of 1cm leaflet block with scissors.
Extraction described in step (4) is extracted to add 30ml distilled water in each sample.
Application of the above-mentioned method in the screening of lily cold-resistant variety.
The detailed process of the screening technique of lily cold-resistant variety of the present invention is comprised the steps of：
(1) second first leaf of wheel below lily plant strain growth point is won, each kind obtains 10 leaves of the same size
Piece, is respectively cleaned three times, then blotted the water on blade with blotting paper with running water and distilled water.
(2) blade in step (1) is laterally cut into a width of 1cm leaflet block with scissors, then every 2 leaves are portion,
One layer is uniformly wrapped up with wet gauze, is put into 25mm × 150mm teat glass, 4 DEG C of nights of refrigerator one are put into, second day stand-by.
(3) sample in step (2) is taken out, parallel is tightened on rack for test tube and with rubber band, then waits low-temperature circulating
When instrument temperature drops to 4 DEG C, processing in low-temperature circulating instrument is put it into.Treatment temperature be 50 DEG C of gradients, -3 DEG C, -6 DEG C, -9
℃、-12℃.Each gradient processing 1h, temperature fall time is 30min between gradient and gradient.Then 4 DEG C of ice are put into after sample being taken out
Thawed in case, further took out to use by second day.
(4) step 3) thaw after sample take out and start extraction, the sample in each test tube divides equally three parts and is put into test tube respectively
In, each middle addition 30ml distilled water, extraction is stayed overnight.
(5) step 4) sample after extracted starts to survey electrical conductivity.15min is boiled after having surveyed, waits and is measured again after cooling
Electrical conductivity.Then its property is obtained to electrical conductivity according to formula I：
Relative conductivity (REC, %)=frost electrical conductivity/boils electrical conductivity × 100% (I)
(6) resist cold in research, relative conductivity Fitting Logistic regression equation is：Y=K/ (1+ae-bx).Wherein y generations
Table cell damage rate, x represents treatment temperature, and K is the saturated capacity of cell damage rate, and a, b are equation parameter.In order to determine a, b
Value, equation is subjected to linearization process, ln [(K-y)/y]=lna-bx makes y1=In [(K-y)/y], is then converted into thin
The linear equation of born of the same parents' injury rate (y1) and treatment temperature (x).A, b values and coefficient R are tried to achieve by the method for linear regression2
(degree of fitting), semilethal low temperature is obtained with equation II.
LT50=In [(1/a)]/b (II)
(7) lily different cultivars cold resistance height is ranked up according to semilethal low temperature, filters out resistance and resistance
Weak kind.
Beneficial effects of the present invention：
The present invention provides an effectively convenient method, cold-resistant variety after being for the screening of lily cold resistance kind
Seed selection and research establish certain basis.Also there is following key character 1 simultaneously) it is right during lily nutrient growth
Lily blade low-temperature treatment, closer to actual production.2) a variety of screening techniques are combined, progressive screening, not only science but also convenient.
Brief description of the drawings
Fig. 1 is ' Te Lasuo oranges ' relative conductivity degree of fitting curve；
Fig. 2 is ' timber ' relative conductivity degree of fitting curve；
Fig. 3 is ' shady deal ' relative conductivity degree of fitting curve.
Embodiment
With reference to instantiation, the present invention is described in further detail.
The present invention carries out experiment screening (table 1) with blade low-temperature treatment to 30 cutting flower varieties.
The strain and Tuber size of the experiment material of table 1
Experiment material is planted in the glasshouse of Agricultural University Of Nanjing's chrysanthemum base, starts to carry out in fact in by the end of October, 2016 in
Test, specific implementation example is as follows：
1.1 win below lily plant strain growth point second first climax leaves of wheel, each kind take 10 it is of the same size
Blade, is respectively cleaned three times with running water and distilled water, then is blotted the water on blade with blotting paper, then with scissors that blade is horizontal
To the leaflet block for being cut into a width of 1cm, then every 2 leaves are portion, be divided into 5 parts, one layer is uniformly wrapped up with wet gauze, 25mm is put into
In × 150mm teat glass, 4 DEG C of nights of refrigerator one being put into, sample being taken out in second day, parallel is on rack for test tube and uses rubber band
Tighten, when then grade low-temperature circulating instrument temperature drops to 4 DEG C, put it into processing in low-temperature circulating instrument, instrument is Polyscience
The PP28-R30 low-temperature circulating instrument of company.Treatment temperature be 50 DEG C of gradients, -3 DEG C, -6 DEG C, -9 DEG C, -12 DEG C,.Each ladder
Degree processing 1h, temperature fall time is 30min between gradient and gradient.Each sample was put into defrosting in 4 DEG C of refrigerators after taking out, by second day
Further take out to use.Sample after defrosting, which takes out, starts extraction, and the sample in each test tube is divided equally 3 parts (3 repetitions) and is put into respectively
In test tube, each middle addition 30ml distilled water, extraction is stayed overnight.Sample after extracted starts to survey electrical conductivity, is boiled after having surveyed
15min, waits and measures electrical conductivity after cooling again.Relative conductivity (REC, %)=frost electrical conductivity/boil electrical conductivity ×
100%, so as to obtain its relative conductivity.
1.2 data processing
Relative conductivity Fitting Logistic regression equation is：
Y=K/ (1+ae-bx)
Semilethal low temperature (LT50)
LT50=In [(1/a)]/b
Y represents cell damage rate, and x represents treatment temperature, and K is the saturated capacity of cell damage rate, and a, b are equation parameter.
1.3 interpretation of result
1.3.1 cell membrane and cold resistance
Electrolyte leakage measurement result shows：30 parts of material electrolyte leakage degree increase with the decline for the treatment of temperature
Greatly, therefore relative conductivity rises also with temperature drop, and relative conductivity curve substantially S-type (Fig. 1~3), in figure
The corresponding temperature of 5 points be 0 respectively, -3, -6, -9, -12 DEG C.As we can clearly see from the figure ' Te Lasuo orange ' and ' shady deal '
Between 0 DEG C Dao -9 DEG C, relative conductivity is in obvious ascendant trend, no longer rises and becomes substantially between being spent at -9 DEG C to -12 DEG C
In stable tendency；' Te Lasuo orange ' relative conductivity suddenly rises between -3 DEG C and -7 DEG C, ' shady deal ' -4 DEG C to -8 DEG C it
Between relative conductivity suddenly rise, illustrate that this 2 kinds injure maximum in this corresponding temperature.S type flex points are exactly that they divide
Not corresponding semilethal low temperature.Curve corresponding to ' timber ' is substantially in rising trend at 0 DEG C Dao -12 DEG C, and -5 are arrived at 0 DEG C
Slowly rise between DEG C, be in rapid increase trend between -6 DEG C to -10 DEG C, decelerating trend is just begun with after -10 DEG C, it
S types flex point then between -6 DEG C and -10 DEG C.
1.3.2 semilethal low temperature (LT50) comparative analysis
30 lily cultivars according to the fitting regression equation obtained by relative conductivity, are intended under different low-temperature treatments
Right and semilethal low temperature is shown in Table 3, is arranged according to semilethal low temperature ascending order.30 lily cultivar turbine blades in electrochemical matter extravasation methods resist cold
Property result of the test shows that the electrolyte osmotic rate of experiment material is raised with the reduction of temperature, and so kind it is relative
The matched curve of electrical conductivity is all into obvious S types, and degree of fitting is preferably, and the thermograde of this explanation experimental design is rational.
LT50It is used as plant cold-resistant evaluation index, LT50Lower cold resistance is stronger, conversely, then weaker.From table 3 it can be seen that 30 parts of realities
The semilethal low temperature of material is tested between -5 DEG C and -9 DEG C, this illustrates that these lily cultivar cold resistance are relatively weak.Wherein resistance
Weaker kind has ' Te Lasuo oranges ', ' Ka Sitaibai ', ' Ye Luolin ', ' small gross guest ', but also has several kinds to resist cold relatively
Property it is relatively strong such as ' timber ', ' cardinal ' (table 2, table 3).
2 30 lily cultivar Logistic equations of table and R2
3 30 lily cultivar LT of table50
By above example it can be seen that：30 lily cultivar vanes are crossed after low-temperature treatment, and electrical conductivity is also with a temperature of
Drop and rise, and relative conductivity curve is substantially S-type, degree of fitting is all higher, and the semilethal low temperature of each kind holds very much
Easily obtain, and then judge that its cold resistance is strong and weak.In this screen lily cold-resistant variety method compared with other screening techniques more
It is simple more easy to operate, when especially being screened to multiple kinds, it can more embody the superiority of this method.Meanwhile, this side
Method is more fitted reality than other method, is screened in the lily plant strain growth phase, otherwise and other method is to lily bulb
Or tissue-cultured seedling handles to screen, then what is come out with actual CROP PING also differs farther out.Therefore, this method is commented lily cold resistance
Valency result can more react the actual cold resistance in lily field, more representative than conventional method.Only it is the excellent of the present invention described in upper
Select embodiment, it should be pointed out that：For those skilled in the art, the premise of the principle of the invention is not being departed from
Under, some improvements and modifications can also be made, these improvements and modifications also should be regarded as protection scope of the present invention.
Claims (10)
1. a kind of screening technique of lily cold-resistant variety, it is characterised in that：Comprise the following steps：
(1) second first climax leaves of wheel below lily plant strain growth point are won and are cleaned up；
(2) climax leaves after cleaning are cut into leaf block, and leaf block is pre-processed；
(3) pretreated leaf block is handled using low-temperature circulating instrument；
(4) sample after processing terminates in low-temperature circulating instrument is taken out into defrosting, the sample extraction after defrosting is stayed overnight；
(5) survey leaching liquor frost electrical conductivity, boiled after having surveyed and cool down again measure electrical conductivity obtain boiling electrical conductivity, according to
Formula relative conductivity=frost electrical conductivity/boils electrical conductivity × 100%, obtains relative conductivity；
(6) it is with Fitting Logistic regression equation：Y=K/ (1+ae-bx) is analyzed relative conductivity, and wherein Y is represented
Cell damage rate, x represents treatment temperature, and K is the saturated capacity of cell damage rate, and a, b is equation parameter；In order to determine a, b's
Value, carries out linearization process, ln [(K-y)/y]=lna-bx makes y=ln [(k-y)/y], is then converted into cell damage by equation
The linear equation of rate (Y) and treatment temperature (x)；A, b values and coefficient R are tried to achieve by the method for linear regression2, calculate lily
The semilethal low temperature LT of kind50=In [(1/a)]/b；
(7) lily cultivar cold resistance is evaluated according to semilethal low temperature, and filters out cold resistance lily cultivar.
2. according to the method described in claim 1, it is characterised in that：The mistake pre-processed described in step (2) is referred to as will be described
Leaf block uniformly wraps up 1 layer with wet gauze, is put into teat glass, is placed in 4 DEG C of refrigerator overnights.
3. according to the method described in claim 1, it is characterised in that：The mistake of leaf block is handled in step (3) using low-temperature circulating instrument
Cheng Wei：When low-temperature circulating instrument temperature drops to 4 DEG C, pretreated sample is put into low-temperature circulating instrument and handled；The low temperature
Circulating instrument treatment temperature be 5 gradients 0, -3, -6, -9, -12 DEG C；Each gradient processing 1h, temperature fall time between gradient and gradient
For 30min.
4. according to the method described in claim 1, it is characterised in that：Will be after processing terminates in low-temperature circulating instrument in step (4)
Sample take out and be put into defrosting in 4 DEG C of refrigerators, the sample after defrosting is taken out into extraction and stayed overnight.
5. according to the method described in claim 1, it is characterised in that：The time boiled described in step (5) is 15min.
6. according to the method described in claim 1, it is characterised in that：Lily plant strain growth described in step (1) is healthy and strong, disease-free
Insect pest is infected.
7. according to the method described in claim 1, it is characterised in that：In step (1) same lily cultivar second take turns first into
Ripe leaf is in the same size.
8. according to the method described in claim 1, it is characterised in that：Climax leaves described in step (2) are laterally cut into scissors
A width of 1cm leaflet block.
9. according to the method described in claim 1, it is characterised in that：Extraction described in step (4) is addition in each sample
30ml distilled water is extracted.
10. application of any described method in the screening of lily cold-resistant variety in claim 1~9.
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN110268979A (en) * | 2019-06-03 | 2019-09-24 | 南京农业大学 | A kind of method and application of the foundation without pollen lily high-efficiency regeneration system |
| CN110308247A (en) * | 2019-07-02 | 2019-10-08 | 海南大学 | A kind of plant cold resistance efficient living body identification method |
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| JPS62239993A (en) * | 1986-04-14 | 1987-10-20 | Oji Paper Co Ltd | Method for transformation of plant cell using spheroplast of escherichia coli |
| JPH06113669A (en) * | 1992-09-29 | 1994-04-26 | Toppan Printing Co Ltd | Seedling bed using pulp foam and its production |
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN110268979A (en) * | 2019-06-03 | 2019-09-24 | 南京农业大学 | A kind of method and application of the foundation without pollen lily high-efficiency regeneration system |
| CN110308247A (en) * | 2019-07-02 | 2019-10-08 | 海南大学 | A kind of plant cold resistance efficient living body identification method |
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Application publication date: 20171024 |