CN103571789B - Jasmine pollen in-vitro germination liquid culture medium and method for measuring activity of jasmine pollen - Google Patents
Jasmine pollen in-vitro germination liquid culture medium and method for measuring activity of jasmine pollen Download PDFInfo
- Publication number
- CN103571789B CN103571789B CN201310565165.9A CN201310565165A CN103571789B CN 103571789 B CN103571789 B CN 103571789B CN 201310565165 A CN201310565165 A CN 201310565165A CN 103571789 B CN103571789 B CN 103571789B
- Authority
- CN
- China
- Prior art keywords
- pollen
- jasmine
- germination
- vitro
- activity
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Active
Links
Landscapes
- Breeding Of Plants And Reproduction By Means Of Culturing (AREA)
Abstract
The invention discloses a jasmine pollen in-vitro germination liquid culture medium and a method for measuring the activity of jasmine pollens. The jasmine pollen in-vitro germination liquid culture medium takes distilled water as a solvent and contains the components including100 mg/L H3BO3, 100 mg/L KNO2, 350 mg/L Ca(NO3)2.4H2O, 150 mg/L MgSO4.7H2O, 80 g/L saccharose and 120 g/L PEG4000, and the pH value is 6.0. The activity of the jasmine pollens can be measured through the jasmine pollen in-vitro germination liquid culture medium and a pollen tube micro observation technology, and a measurement result is stable and reliable; the pollen germination rate reflects the proportion of viable pollens in the measured pollens; the micro observation and measurement of the growth of a pollen tube can reflect the growth state and the physiological status of pollen particles; an effective and reliable method is provided for measuring the activity of the jasmine pollens through jasmine pollen in-vitro germination and the pollen tube micro observation technology; an important application value and a great practical significance for development of researches on jasmine genetic breeding can be achieved.
Description
one, technical field
The invention belongs to biological technical field, relate to a kind of method of jasmine in-vitro pollen germination liquid nutrient medium and mensuration jasmine Pollen Activity.
two, background technology
Jasmine (
jasminum sambacaiton) Flower of Arabian Jasmine, bluish dogbane is had another name called, for Oleaceae gelsemium is evergreen upright or climb up by holding on to shape shrub.The Han dynasty of jasmine before more than 1700 years just imports China into from the west and south, Asia, and milpa is in the coastland in Hainan and Guangdong and Guangxi Provinces and Fujian the earliest, then gradually goes up north until the Yangtze valley.Because its plant type is exquisite, with luxuriant foliage and spreading branches in leafy profusion, Ye Se is as emerald, and flower like beautiful bell, and spends many phases long, fragrance delicate fragrance and lasting, strong and not turbid, is described as treasure, " the first under heaven perfume " (Dong Lijuan, 2001) of Hua Shuzhong by common people.Jasmine has been widely used, have higher to view and admire, tea with and pharmaceutical use, Flower of Arabian Jasmine is smoked except jasmine tea except being directly used in, also can extract medicinal extract and essential oil for perfume industry, be one of main raw material of high-grade daily-use cosmetic essence and high quality toilet soap essence, especially prepare the important source material of highclass perfumes essence.Flower of Arabian Jasmine can potted plant or do flower hedge view and admire, also often wear or corbeil as boutonniere, always very popular.At present, domestic commerial growing jasmine except Fujian, Guangdong, Guangxi, also have the provinces such as Zhejiang, Taiwan, Jiangsu, Anhui, Jiangxi, Hunan, Sichuan, be mainly used for basement Jasmine tea and potted plantly view and admire, there is huge economic benefit, China is made to become the maximum country (Dong Lijuan, 2001) of Flower of Arabian Jasmine cultivated area and output in the world.Because Flower of Arabian Jasmine under natural condition is usually shaky, long-term vegetative propagation makes jasmine seeds of flowering plants sexual involution, and yield poorly, resistance is weak.But during various places produce, improved seeds comparatively lack, and have had a strong impact on the development that China jasmine produces.Therefore, the seed selection carrying out improved seeds is an important process of jasmine investigator.Carry out the research of jasmine pollen vitality test, to the seed selection etc. of Germ-plasma resources protection and innovation, improved seeds, there is important theory and practice meaning.
Jasmine dysgenesia is serious, and under natural condition, setting percentage is extremely low or not solid.Rely show one's high ideals (1995) to find that the germination rate of jasmine pollen is 3.68% ~ 9.68%, report the result of its flower bud development being observed by paraffin method subsequently, think that its syngenesis is degenerated (rely and show one's high ideals, 1996); Dong Lijuan and Zhang Shuguang (2001) think that its pollen germination percentage is about 3%; Zeng Zhen etc. (2002) have also carried out anatomical research by paraffin method to its growth course, finding that the normal pollen granule of form accounts for 23.5%(normal is 100:324 with abnormal ratio), think that jasmine Devflopment Ofmle Gametophyte is scarcely normal, male fertile is lower is cause its acarpous one of the main reasons.In addition, jasmine is hermaphrodite flower, and Post flowering pollen maturation also sheds in pollen sac, even if the pollen granule of maturation has certain vigor, but its vigor still with physiological status, environment and the passing of time etc., certain change can occur.Therefore, understand the vigor condition of jasmine Post flowering pollen, to the research of the foundation of jasmine hybridization system and jasmine germplasm innovation all tool be of great significance.
At present, Pollen Activity method for measuring mainly contains morphologic observation, staining, germination in vitro method etc.Morphologic observation judges vigor according to pollen profile, pollen sterile in process of growth owing to being subject to the impact of some factor, ateliosis or bad and normal in lopsided, and the normal well-regulated profile of pollen.Although can differentiate Pollen Activity according to form, normal pollen granule vigor in storage may be lost, and considerable change does not occur form; Although also there is partial pollen profile normal, physiological function is poor, thus makes pollen morphology observation have certain error when differentiating Pollen Activity.Staining is by using staining agent to dye to pollen granule, judging the height of Pollen Activity according to colour-change.Conventional staining reagent has IKI (I
2-KI) solution and triphenyltetrazolium chloride (TTC) solution etc.Because the colour generation of jasmine pollen own is comparatively dark, experiment finds that the effect of staining is unsatisfactory, and experimental repeatability is poor, can cause very large experimental error, can not the actual vigor height of Accurate Determining pollen.Paraffin method has advantage very intuitively on Study of Pollens developmental process and characteristic, but also can only judge Pollen Activity according to form, and often also there is a certain distance between actual vigor; And as mentioned above, between different research reports, gap is very large.
The result of in-vitro pollen germination and pollen tube microscopic observation is comparatively stable, more can react the development condition of pollen and actual vigor, be considered to effect best, closest to the method (Zhao Hongbo etc., 2007) of baseline results.But in-vitro pollen germination needs suitable substratum, otherwise is also difficult to obtain desirable effect.The kinds of culture medium that different Plant Pollen Germination needs is different with concentration, often will screen minimum medium or improve, and also needs if desired to add some element promoting pollen germination or materials on this basis, as Ca
2+, K
2+, PEG, sugar etc.In recent years, people have carried out more research to plant pollen germination in vitro and pollen tube growth.Zhang Biyu (1983) reports that the sucrose of suitable concentration can promote the sprouting of pollen, and Wang Siqing etc. (1993) and Zhao Hongbo etc. (2005) think that PEG has remarkable promoter action to chrysanthemum and germination in vitro of Mei flower growth; Niu Dongling etc. (2004) report the appropriate media of cistanche-pollen germination in vitro, and old and bright etc. (2006) have studied herba fibraureae recisae in-vitro pollen germination condition and low temperature to the impact of Pollen Activity.Liu, from just waiting (2011) to report the germination in vitro of root of large-flowered skullcap pollen, finds that BK substratum adds certain density sucrose and PEG can make pollen germination well-grown.But there is not been reported both at home and abroad about the research of jasmine in-vitro pollen germination and pollen tube growth.
Compared with other crop pollen germination in vitro liquid nutrient mediums, though jasmine in-vitro pollen germination liquid nutrient medium involved in the present invention with BK substratum for minimum medium, carried out suitable improvement and (improve Ca
2+concentration and correspondingly reduce Mg
2+concentration), and simultaneously on the pH value of the sucrose added and the concentration of PEG, substratum, cultivate time the sample time of flower pesticide, the temperature and light of the cultivation factor sprouted according to the impact such as intensity, incubation time all carried out screening and optimum combination, thus be more suitable for the germination in vitro of jasmine pollen and the normal growth of pollen tube, also advantageously in the qualification of jasmine Pollen Activity.
three, summary of the invention
technical problemthe object of this invention is to provide a kind of jasmine in-vitro pollen germination liquid nutrient medium and the method for measuring jasmine Pollen Activity thereof, first screening, improved culture medium, optimum culture condition, solve the low problem of jasmine in-vitro pollen germination rate, the germination rate of jasmine pollen is increased substantially, thus reliable, the effective vigor method measuring jasmine pollen is provided.
technical schemein order to realize above-mentioned task, the present invention is achieved by the following technical programs:
1)a kind of jasmine in-vitro pollen germination liquid nutrient medium, it is characterized in that, this obtained jasmine in-vitro pollen germination liquid nutrient medium take distilled water as solvent, and its composition contains the H of 100mg/L
3bO
3, 100mg/L KNO
3, 150mg/L MgSO
47H
2ca (the NO of O, 350mg/L
3)
24H
2the sucrose of O, 80g/L and the PEG4000 of 120g/L, pH value is 6.0;
2)above-mentioned jasmine in-vitro pollen germination liquid nutrient medium, for measuring the method for jasmine Pollen Activity, is characterized in that, comprises the following steps:
1. pollen collection
Choose growing way stalwartness, grow normal, without the jasmine plant of disease and pest, full-bloom stage in the morning 8:00 9:00 choose a bud just ready to burst flower, together adopt down together with segment length 10 ~ 20cm branch and blade, insert rapidly and fill in the triangular flask of clear water, push petal aside after taking back laboratory, take off flower pesticide with tweezers and divide on template, put 1 2h in 35 ° of C baking ovens, short pollen sac dehydration is broken loose powder, is got up by pollen collecting stand-by;
2. in-vitro pollen germination
Get above-mentioned jasmine in-vitro pollen germination liquid nutrient medium, drip 35 drop in slide glass middle part and make thinner, dip appropriate jasmine pollen with writing brush and be evenly sprinkling upon liquid culture primary surface, slide glass is placed in the culture dish that bottom is lined with 1 layer of waterlogged filter paper, and the waterlogged rayon balls moisturizing that is of moderate size respectively is placed in slide glass both sides, put into incubator illumination cultivation after covering culture dish lid, culture temperature is 25 DEG C 30 DEG C, and intensity of illumination is 35 50 μm of olm
-2s
-1; After in-vitro pollen germination cultivates 30min, partial pollen starts to start to be sprouted, and pollen germination pore expands;
3. the mensuration of pollen tube growth and Pollen Activity
After pollen tube length after sprouting exceedes pollen granule diameter, pollen tube starts quick growth; After isolated culture 4h, carry out pollen germination rate statistics and pollen tube length measurement; Pollen diameter is greater than for pollen germination standard with pollen tube length, random observation under the microscope, often processes repetition more than 3 times, often repeats random observation 35 visuals field, every visual field is observed pollen number and is no less than 40, and statistics pollen germination rate is to reflect great-hearted pollen proportion; Simultaneously, pollen tube length is measured with micro-micrometer, every visual field random measurement 10 20 pollen tubes, often process and measure 100 pollen tubes altogether, calculate pollen tube length mean value, in-vitro pollen germination pollen tube growth situation can reflect that pollen granule grows the quality of whether normal and physiological situation, to reflect Pollen Activity size.
beneficial effectthe present invention compared with prior art, has the following advantages and useful technique effect:
1, jasmine pollen is on jasmine in-vitro pollen germination liquid nutrient medium provided by the invention, temperature 25 DEG C 30 DEG C, intensity of illumination 35 50 μm of olm
-2s
-1cultivate under condition, jasmine pollen germination is best, and pollen tube also can better be grown; And jasmine pollen tube growth is comparatively straight, is convenient to observe and measures.Jasmine in-vitro pollen germination liquid nutrient medium, compared with on other crop pollen germination in vitro liquid nutrient medium compositions, though with BK substratum for minimum medium, has carried out certain improvement to it, has suitably improve Ca
2+content and correspondingly reduce Mg
2+content, and all screening and optimum combination have been carried out to the condition that the concentration of the sucrose that substratum adds and PEG, the pH value of substratum, sample time, culture temperature and intensity of illumination etc. affect pollen cultures.
2, the present invention is drawn by test: the morning 8:00 9:00 collect jasmine pollen, PH be 6.0 improvement BK liquid nutrient medium (composition is the H of 100mg/L
3bO
3, 100mg/L KNO
3, 150mg/L MgSO
47H
2ca (the NO of O, 350mg/L
3)
24H
2o) on, after adding 80 g/L sucrose and 120g/L PEG4000, in 25 ° of C, 30 ° of C temperature, 35 50 μm of olm
-2s
-1under intensity of illumination after isolated culture 3 4h, measure the mean length (0.55mm) of jasmine pollen germination rate (being 59.7% to the maximum) and pollen tube, the vigor condition of effective reflection jasmine pollen can be stablized, (specific experiment result is see Fig. 2 to demonstrate verity of the present invention and validity intuitively, wherein, the pollen of A: isolated culture 0.5h, the germ pore of partial pollen expands, start to start and sprout, now germination rate and pollen tube length are 0; B: cultivate the pollen of 2h and the pollen tube of growth, germination rate is 18.8%, and pollen tube length is 0.09mm, but partial pollen is still in sprouting, and its pollen tube length, still not as good as pollen granule diameter, does not reach sprouting standard, therefore still need to proceed to cultivate; C: cultivate the pollen granule of 4h and grow normal complete pollen tube, now germination rate reaches the highest (59.7%) and pollen tube still keeps comparatively straight (length is 0.55mm), is convenient to observe and measure; D: cultivate the pollen after 6h, now germination rate starts to decline, and much pollen tube occurs distortion because growing long (>10mm), interwind phenomenon, there is the unusual phenomenoies such as end expands, degraded in partial pollen pipe simultaneously, is all unfavorable for observation and compares).
3, with original authentication method as compared with paraffin section, morphologic observation etc., it is more reliable and more stable that the present invention utilizes this jasmine in-vitro pollen germination liquid nutrient medium and pollen tube growth micro-observation technology to measure the measurement result of Pollen Activity, pollen germination rate reaction has vigor pollen proportion, the observation of pollen tube growth react the quality that pollen granule grows whether normal and physiological situation with measuring, and the mensuration for jasmine Pollen Activity provides more effective, the reliable method of one.After screening, improvement, optimum combination and condition, the germination rate of jasmine pollen is maximum can be about 3 times of maximum value (23.5%) in original bibliographical information close to 60%, do not improve 2 times of BK substratum.
4, on basis of the present invention, the jasmine pollen that the vigor that can filter out is higher carries out artificial pollination and the hybridization of jasmine, makes the germplasm innovation being carried out jasmine by sexual route become possibility.
four, accompanying drawing explanation
Fig. 1 is jasmine pollen isolated culture vessel used and puts schematic diagram (left side is culture dish, slide glass, the arrangement method schematic diagram that is layered on 1 layer of waterlogged filter paper below slide glass, puts 2 waterlogged rayon balls in slide glass both sides, and the right is the lid of culture dish)
Fig. 2 is pollen germination and the pollen tube growth situation of cultivating different time; Wherein, the pollen of A: isolated culture 0.5h, the germ pore of partial pollen expands, and starts to start to sprout, and now germination rate and pollen tube length are 0; B: cultivate the pollen of 2h and the pollen tube of growth, germination rate is 18.8%, and pollen tube length is 0.09mm, but partial pollen is still in sprouting, and its pollen tube length, still not as good as pollen granule diameter, does not reach sprouting standard, therefore still need to proceed to cultivate; C: cultivate the pollen granule of 4h and grow normal complete pollen tube, now germination rate reaches the highest (59.7%) and pollen tube still keeps comparatively straight (length is 0.55mm), is convenient to observe and measure; D: cultivate the pollen after 6h, now germination rate starts to decline, and much pollen tube occurs distortion because growing long (>10mm), interwind phenomenon, there is the unusual phenomenoies such as end expands, degraded in partial pollen pipe simultaneously, is all unfavorable for observation and compares.
Below in conjunction with drawings and Examples, the present invention is described in further detail.
five, embodiment
According to technical scheme of the present invention, first prepare jasmine in-vitro pollen germination liquid nutrient medium, this jasmine in-vitro pollen germination liquid nutrient medium take distilled water as solvent, the H wherein containing 100mg/L
3bO
3, 100mg/L KNO
3, 350mg/L Ca (NO
3)
24H
2the MgSO of O, 150mg/L
47H
2the sucrose of O, 80g/L and the PEG4000 of 120g/L, pH value is 6.0;
On the basis determining jasmine in-vitro pollen germination liquid nutrient medium, this jasmine in-vitro pollen germination liquid nutrient medium is utilized to carry out isolated culture to jasmine pollen, measure the germination rate of pollen and the growing state of pollen tube, reliably, effectively can measure jasmine Pollen Activity.
Be below the embodiment that contriver provides, this embodiment is the present invention's preferably example, is mainly used in explaining further and understanding the present invention, the invention is not restricted to this embodiment.
1, the collection of jasmine pollen
Jasmine pollen is pick up from the perennial jasmine colony of Jiangsu Province Agriculture Science Institute jasmine Germplasm Resources in early July, 2012.Acquisition method is: choose growing way stalwartness, grow normal, without the jasmine plant of disease and pest, at full-bloom stage respectively at choosing the flower that a bud just ready to burst in the morning (8:00 9:00), noon (12:00 13:00) and afternoon (17:00 18:00), together adopt down together with segment length 10 ~ 20cm branch and blade, insert rapidly and fill in the triangular flask of clear water, petal is pushed aside after taking back laboratory, taking off flower pesticide with tweezers divides on template, put 1 2h in 35 DEG C of baking ovens, short pollen sac dehydration is broken loose powder, is collected by pollen stand-by respectively.
2, the screening of in-vitro pollen germination culture condition
2. the screening of 1 in-vitro pollen germination substratum
Kinds of culture medium has considerable influence to plant pollen germination in vitro, and the appropriate media of different Plant Pollen Germination is different.The minimum medium arranging 5 kinds of heterogeneities screens, and is respectively:
(1) ME
3(composition is MgSO to substratum
47H
2o 370mg/L, KNO
3950mg/L, KH
2pO
485mg/L, CaCl
22H
2o 880mg/L, NH
4nO
3412.5mg/L, KCl 175mg/L, H
3bO
350mg/L, Na
2eDTA 7.45mg/L, FeSO
47H
2o 0.025mg/L, KI 0.83mg/L, Na
2moO
42H
2o 0.25mg/L, CuSO
45H
2o 0.025mg/L, CoCl
26H
2o 0.025mg/L, VB
11.0mg/L, VB
61.0mg/L); (2) (composition is BK substratum: H
3bO
3100mg/L, KNO
3100mg/L, MgSO
47H
2o 200mg/L, Ca (NO
3)
24H
2o 300mg/L); (3) BK substratum is improved (for increasing Ca
2+content adjustment Ca (NO
3)
24H
2o concentration is 350mg/L, makes MgSO for corresponding reduction total ion concentration simultaneously
47H
2the content of O is adjusted to 150mg/L, and other compositions are constant with BK); The sucrose solution of (4) 100 g/L; (5) the PEG4000 solution of 150g/L; Cultivate in contrast (CK) with distilled water.
By pollen respectively to cultivate on above-mentioned 6 kinds of substratum under identical illumination, temperature condition, as analysed basis basal culture medium is on the impact of in-vitro pollen germination and pollen tube growth; The germination rate of pollen and the measurement of length method of pollen tube are:
Pollen diameter is greater than for pollen germination standard with pollen tube length, random observation under the microscope, often processes repetition more than 3 times, often repeats random observation 35 visuals field, every visual field is observed pollen number and is no less than 40, and statistics pollen germination rate is to reflect great-hearted pollen proportion; Meanwhile, measure pollen tube length with micro-micrometer, every visual field random measurement 10 20 pollen tubes, often process measurement altogether and be no less than 100 pollen tubes, calculate its mean value.
The results are shown in Table 1.As can be found from Table 1, jasmine pollen all can not normally be sprouted, at ME in the sucrose of single component, PEG, distilled water
3germination rate on substratum is lower than 10%, though can 20% be reached on BK substratum, but it is best to sprout effect with the BK substratum after improvement, germination rate the highest (30.1%), higher by 44.7% than BK substratum (20.8%), pollen tube length maximum (0.29mm), higher by 38.1% than BK substratum (0.21mm), be all significantly higher than other substratum.Meanwhile, in same medium, all behave oneself best with 8:00 pollen that 9:00 gets in the morning, therefore with this time period for the jasmine pollen samples optimal acquisition time, all with this time point institute sample thief, (composition is the H of 100mg/L to subsequent experimental with improvement BK substratum
3bO
3, 100mg/L KNO
3, 150mg/L MgSO
47H
2ca (the NO of O, 350mg/L
3)
24H
2o) premised on, condition is carried out.
Table 1 substratum and sample time are on the impact of jasmine pollen germination
Note: different lowercase represents significant difference between different treatment (P<0.05);-represent that not observing pollen normally sprouts
2. the screening of 2 in-vitro pollen germination conditions
When other conditions are identical, carry out single factors test, arrange respectively sucrose concentration (40,80,120,160,200 g/L 5 process), PEG concentration (40,80,120,160,200 g/L 5 process), pH value (5.0,5.5,6.0,6.5 4 process), culture temperature (20,25,30,35,40 DEG C 5 process), incubation time (0.5,1,2,3,4,6h totally 6 process), illumination condition (dark culturing and 35 50 μm of olm
-2s
-1illumination cultivation 2 process) carry out screening and the optimization of culture condition one by one.
Table 2 culture condition is on the impact of jasmine pollen germination and pollen tube growth
Note: different lowercase represents significant difference between different treatment (P<0.05)
Test-results in table 2, as can be seen from Table 2:
Effect difference sprouted by the sucrose adding different concns in substratum, and the lower concentration sucrose of 40 g/L does not have obvious promoter action, and 120 g/L and above high density have obvious inhibition, therefore determines that best sucrose concentration is 80g/L;
The PEG4000 of different concns is added in substratum, all can significantly improve jasmine pollen germination rate and promote pollen tube growth, in low strength range (≤120), germination rate increases with the increase of PEG concentration, but (>=160) pollen germination is suppressed equally after concentration is higher, make germination rate occur declining and occurring pollen tube fracture phenomena, therefore determine that best PEG concentration is 120g/L;
PH value change all has remarkably influenced to jasmine pollen germination and pollen tube growth, and when P Η value is in 5.0 6.0 scopes, pollen germination rate and pollen tube length increase with the rising of P Η, but when pH value is greater than more than 6.0, starts to decline rapidly, show substratum H
+excessive concentration or the too low pollen germination that all significantly suppresses grow, therefore best pH value is defined as 6.0;
Culture temperature has remarkably influenced to jasmine pollen germination and pollen tube growth, in 20 ° of C, 30 ° of C scopes, germination rate increases with the rising of temperature, but pollen tube growth is excessively slow during 20 ° of C lesser tempss, when comparatively high temps (>=35 ° of C), pollen tube growth is too fast, easily be distorted, coil phenomenon, impact is observed and is measured, 40 ° of C high temperature then Developing restraint cause growth failure, therefore optimum culturing temperature is defined as 25 ° of C, 30 ° of C;
Incubation time is different, also larger on the impact of sprouting effect and vigor observation, cultivating 0.5h only has minority pollen granule to start to start sprouting, occur that germ pore expands phenomenon, pollen granule diameter is greater than to partial pollen length of tube during 1h, reach sprouting standard, but germination rate is too low, and pollen tube length is all too short generally, be not suitable as judgement criteria, though minority pollen tube length is larger during 2h, but pollen tube length is inadequate because sprouting more late to still have partial pollen, still need to continue incubation growth, after cultivating 6h, find that germination rate starts to decline, distortion is there is in a lot of pollen tube because of oversize (>10mm), interwind phenomenon, and partial pollen pipe generation end expands, the unusual phenomenon such as to break, all be not suitable for observation, therefore incubation time is with 3 4h, and for the best, (now pollen germination rate is respectively 39.1 and 42.4, pollen tube mean length is respectively 0.32mm and 0.43mm),
Illumination on the impact of jasmine pollen germination significantly, can not normally to be sprouted under dark culturing and pollen tube growth is slow, therefore illumination condition is with 35 50 μm of olm
-2s
-1be advisable.
Comprehensive above-mentioned result of study, can draw: the morning 8:00 9:00 collect jasmine pollen, PH be 6.0 improvement BK liquid nutrient medium (composition is the H of 100mg/L
3bO
3, 100mg/L KNO
3, 150mg/L MgSO
47H
2ca (the NO of O, 350mg/L
3)
24H
2o) on, after adding 80 g/L sucrose and 120g/L PEG4000, in 25 ° of C, 30 ° of C temperature, 35 50 μm of olm
-2s
-1under intensity of illumination after isolated culture 3 4h, measure jasmine pollen germination rate maximum (59.7%, it is about 3 times of BK substratum, 2 times that improve BK substratum, also be have been reported more than 2 times) and the mean length (0.55mm) of pollen tube, the vigor condition of jasmine pollen can be reflected to greatest extent, (specific experiment result is see Fig. 2 to demonstrate verity of the present invention and validity intuitively, wherein, the pollen of A: isolated culture 0.5h, the germ pore of partial pollen expands, and starts to start to sprout, and now germination rate and pollen tube length are 0; B: cultivate the pollen of 2h and the pollen tube of growth, germination rate is 18.8%, and pollen tube length is 0.09mm, but partial pollen is still in sprouting, and its pollen tube length, still not as good as pollen granule diameter, does not reach sprouting standard, therefore still need to proceed to cultivate; C: cultivate the pollen granule of 4h and grow normal complete pollen tube, now germination rate reaches the highest (59.7%) and pollen tube still keeps comparatively straight (length is 0.55mm), is convenient to observe and measure; D: cultivate the pollen after 6h, now germination rate starts to decline, and much pollen tube occurs distortion because growing long (>10mm), interwind phenomenon, there is the unusual phenomenoies such as end expands, degraded in partial pollen pipe simultaneously, is all unfavorable for observation and compares).
Claims (2)
1. measure a method for jasmine Pollen Activity, it is characterized in that:
1) pollen collection
Oleaceae gelsemium jasmine (the Jasminum sambac Aiton) plant that growth selection is healthy and strong, grow the harm of normal, anosis worm, full-bloom stage in the morning 8:00 ~ 9:00 choose a bud just ready to burst flower, together adopt down together with segment length 10 ~ 20cm branch and blade, insert and fill in the triangular flask of clear water, push petal aside after taking back laboratory, take off flower pesticide with tweezers and divide on template, put 1 ~ 2h in 35 DEG C of baking ovens, short pollen sac dehydration is broken loose powder, is got up by pollen collecting stand-by;
2) in-vitro pollen germination
Get jasmine in-vitro pollen germination liquid nutrient medium, drip 3 ~ 5 drop in slide glass middle part and make thinner, dip jasmine pollen with writing brush and be evenly sprinkling upon liquid culture primary surface, slide glass is placed in the culture dish that bottom is lined with 1 layer of waterlogged filter paper, and a waterlogged rayon balls moisturizing is respectively placed in slide glass both sides, put into constant incubator illumination cultivation after covering culture dish lid, culture temperature is 25 DEG C ~ 30 DEG C, and intensity of illumination is 35 ~ 50 μm of olm
-2s
-1,
Described jasmine in-vitro pollen germination liquid nutrient medium, comprising: take distilled water as solvent, its composition contains H
3bO
3100mg/L, KNO
3100mg/L, MgSO
47H
2o 150mg/L, Ca (NO
3)
24H
2o350mg/L, sucrose 80g/L and PEG4000120g/L, pH value is 6.0;
3) mensuration of Pollen Activity
Maintenance said temperature, illumination condition impel pollen tube growth, after isolated culture 3 ~ 4h, are examined under a microscope by slide glass, carry out pollen germination rate statistics and pollen tube length measurement; Be greater than pollen diameter for pollen germination standard with pollen tube length, often process repetition more than 3 times, often repeat random observation 3 ~ 5 visuals field, every visual field is observed pollen number and is no less than 40, and statistics pollen germination rate is to reflect great-hearted pollen proportion.
2. method according to claim 1, is characterized in that:
Described step 3) mensuration of Pollen Activity time, measure pollen tube length, every visual field random measurement 10 ~ 20 pollen tubes with micro-micrometer simultaneously, often process measurement altogether and be no less than 100 pollen tubes, calculate pollen tube length mean value, to reflect the quality of pollen physiological situation.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201310565165.9A CN103571789B (en) | 2013-11-13 | 2013-11-13 | Jasmine pollen in-vitro germination liquid culture medium and method for measuring activity of jasmine pollen |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201310565165.9A CN103571789B (en) | 2013-11-13 | 2013-11-13 | Jasmine pollen in-vitro germination liquid culture medium and method for measuring activity of jasmine pollen |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN103571789A CN103571789A (en) | 2014-02-12 |
| CN103571789B true CN103571789B (en) | 2015-04-29 |
Family
ID=50044504
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201310565165.9A Active CN103571789B (en) | 2013-11-13 | 2013-11-13 | Jasmine pollen in-vitro germination liquid culture medium and method for measuring activity of jasmine pollen |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN103571789B (en) |
Families Citing this family (9)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103983644A (en) * | 2014-03-10 | 2014-08-13 | 云南农业大学 | Determination method for tomato pollen vitality |
| CN106568894A (en) * | 2016-10-18 | 2017-04-19 | 南京农业大学 | Systematic evaluation method of lily resource powder characteristics and application thereof |
| CN107325969B (en) * | 2017-09-04 | 2020-12-01 | 华北理工大学 | Pollen liquid germination device |
| CN108693308A (en) * | 2018-04-12 | 2018-10-23 | 湖北省农业科学院经济作物研究所 | A kind of method of Fast Evaluation strawberry cultivars petal falling speed |
| CN109142234A (en) * | 2018-08-28 | 2019-01-04 | 内蒙古农业大学 | A kind of method of Field study K-wire tension band root activity |
| CN110218761A (en) * | 2019-07-02 | 2019-09-10 | 中国科学院新疆生态与地理研究所 | Utilize the method for husky honey raisin tree in-vitro pollen germination Liquid Culture based assays sand honey raisin tree pollen activity |
| CN111771659B (en) * | 2020-08-04 | 2022-02-22 | 广西壮族自治区农业科学院 | Method for improving fruiting rate of jasmine flowers and pollination device thereof |
| CN114793882A (en) * | 2022-04-21 | 2022-07-29 | 贵州省园艺研究所(贵州省园艺工程技术研究中心) | Breeding method and application of pink strawberry |
| CN116008196B (en) * | 2023-03-20 | 2023-06-02 | 中国科学院昆明植物研究所 | Method for rapidly and minimally invasively detecting activity of young garlic fruits |
Family Cites Families (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101633902B (en) * | 2009-08-25 | 2011-01-26 | 商洛学院 | In-vitro pollen germination liquid medium of balloon flower and method using same to determine pollen viability thereof |
| CN101717747B (en) * | 2009-12-04 | 2012-05-09 | 西北农林科技大学 | Liquid medium for Radix scutellariae pollen in-vitro germination and method for testing activity of Radix scutellariae pollen |
-
2013
- 2013-11-13 CN CN201310565165.9A patent/CN103571789B/en active Active
Also Published As
| Publication number | Publication date |
|---|---|
| CN103571789A (en) | 2014-02-12 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN103571789B (en) | Jasmine pollen in-vitro germination liquid culture medium and method for measuring activity of jasmine pollen | |
| CN101717747B (en) | Liquid medium for Radix scutellariae pollen in-vitro germination and method for testing activity of Radix scutellariae pollen | |
| Sangster | Intracellular silica deposition in immature leaves in three species of the Gramineae | |
| CN102415253B (en) | Method for screening cold-resistant variety of steeplebush | |
| CN101513167B (en) | Method for obtaining a distant hybrid between cultivating chrysanthemum and related genera | |
| CN102550405B (en) | Breeding method of poplar haploid | |
| CN101243776A (en) | Cultivation method for cabbage type rape Isolated microspore plant strain | |
| CN101283669A (en) | Breeding technique of obtaining triploid grape and ploidy early identification using embryo | |
| CN105230491A (en) | Breeding method for tetraploid ricinus communis | |
| CN103396981B (en) | In vitro culture and vitality determination method for water lily pollen | |
| CN109122306A (en) | A method of obtaining high cannabidiol content industrial hemp plant | |
| CN104160953B (en) | The method of mutagenesis of a kind of tetraploid petunia | |
| CN105052730B (en) | A kind of breeding method of triploid petunia | |
| CN102919132B (en) | Tissue culture method for inducing populus yunnanensis dode tetraploid in combination with colchicine | |
| CN103461106B (en) | Method for breeding and planting morning glory with high ornamental value, disease resistance and high yield | |
| CN101524047B (en) | Method for producing core seeds of thermosensitive genic male sterile line of rice by using 'three-layer ear method' | |
| CN103837534B (en) | A kind of method measuring paenoiae alba Pollen Activity | |
| CN105009901A (en) | Method for detecting sterility of photo-thermo-sensitive genic male sterile rice according to multiphase and multi-sample method | |
| CN112075339B (en) | Method for cultivating peony polyploids by efficiently inducing polyploid pollen | |
| CN104480184A (en) | Method for determining viability of pollen by carrying out in-vitro germination culture on brassica rapa pollen | |
| CN104663431B (en) | Rhizoma Coptidis place, Lichuan fine-variety breeding method | |
| CN114190277A (en) | Method for promoting blooming and fructification of large root orchid test tube | |
| CN112868525A (en) | Method for cultivating tetraploid incarvillea | |
| CN108990793B (en) | Breeding method of tetraploid petunia | |
| CN106605595B (en) | It is cultivated fine seed strains the method for fiery fiber crops by stem apex |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| C06 | Publication | ||
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| C14 | Grant of patent or utility model | ||
| GR01 | Patent grant |