CN104067821B - A kind of preparation method of Sweetpotato Viruses Elimination seedling - Google Patents
A kind of preparation method of Sweetpotato Viruses Elimination seedling Download PDFInfo
- Publication number
- CN104067821B CN104067821B CN201410291953.8A CN201410291953A CN104067821B CN 104067821 B CN104067821 B CN 104067821B CN 201410291953 A CN201410291953 A CN 201410291953A CN 104067821 B CN104067821 B CN 104067821B
- Authority
- CN
- China
- Prior art keywords
- seedling
- preparation
- potato
- stem
- medium
- 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.)
- Expired - Fee Related
Links
- 244000017020 Ipomoea batatas Species 0.000 title claims abstract description 48
- 235000002678 Ipomoea batatas Nutrition 0.000 title claims abstract description 48
- 241000700605 Viruses Species 0.000 title claims abstract description 38
- 230000008030 elimination Effects 0.000 title claims abstract description 21
- 238000003379 elimination reaction Methods 0.000 title claims abstract description 21
- 238000002360 preparation method Methods 0.000 title claims abstract description 15
- 244000061456 Solanum tuberosum Species 0.000 claims abstract description 53
- 235000002595 Solanum tuberosum Nutrition 0.000 claims abstract description 53
- 229920003023 plastic Polymers 0.000 claims abstract description 38
- 239000004033 plastic Substances 0.000 claims abstract description 38
- 241000196324 Embryophyta Species 0.000 claims abstract description 29
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Chemical compound O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims abstract description 24
- 238000001784 detoxification Methods 0.000 claims abstract description 20
- 239000002689 soil Substances 0.000 claims abstract description 20
- 238000012360 testing method Methods 0.000 claims abstract description 20
- 210000001161 mammalian embryo Anatomy 0.000 claims abstract description 19
- 238000000034 method Methods 0.000 claims abstract description 15
- 235000018290 Musa x paradisiaca Nutrition 0.000 claims abstract description 12
- XSQUKJJJFZCRTK-UHFFFAOYSA-N Urea Chemical compound NC(N)=O XSQUKJJJFZCRTK-UHFFFAOYSA-N 0.000 claims abstract description 10
- 239000004202 carbamide Substances 0.000 claims abstract description 10
- 239000003337 fertilizer Substances 0.000 claims abstract description 10
- 230000001954 sterilising effect Effects 0.000 claims abstract description 8
- 238000004659 sterilization and disinfection Methods 0.000 claims abstract description 8
- 238000003973 irrigation Methods 0.000 claims abstract description 7
- 230000002262 irrigation Effects 0.000 claims abstract description 7
- 230000000392 somatic effect Effects 0.000 claims abstract description 6
- 239000008223 sterile water Substances 0.000 claims abstract description 5
- 230000004069 differentiation Effects 0.000 claims abstract description 3
- 230000000763 evoking effect Effects 0.000 claims abstract description 3
- 240000008790 Musa x paradisiaca Species 0.000 claims abstract 3
- 230000003203 everyday effect Effects 0.000 claims description 11
- 239000004576 sand Substances 0.000 claims description 11
- 206010020649 Hyperkeratosis Diseases 0.000 claims description 10
- 239000005556 hormone Substances 0.000 claims description 9
- 229940088597 hormone Drugs 0.000 claims description 9
- 241000238631 Hexapoda Species 0.000 claims description 8
- 238000002054 transplantation Methods 0.000 claims description 8
- 238000009395 breeding Methods 0.000 claims description 6
- 230000001488 breeding effect Effects 0.000 claims description 6
- 238000012258 culturing Methods 0.000 claims description 6
- 241000607479 Yersinia pestis Species 0.000 claims description 4
- 238000009400 out breeding Methods 0.000 claims description 4
- 230000001172 regenerating effect Effects 0.000 claims description 4
- 230000035784 germination Effects 0.000 claims description 3
- 210000002257 embryonic structure Anatomy 0.000 claims description 2
- 230000001605 fetal effect Effects 0.000 claims description 2
- 230000004083 survival effect Effects 0.000 abstract description 13
- 230000008929 regeneration Effects 0.000 abstract description 11
- 238000011069 regeneration method Methods 0.000 abstract description 11
- 239000000463 material Substances 0.000 abstract description 10
- 230000008569 process Effects 0.000 abstract description 8
- 230000003612 virological effect Effects 0.000 abstract description 5
- 230000007423 decrease Effects 0.000 abstract description 4
- 238000003745 diagnosis Methods 0.000 abstract description 3
- 230000035800 maturation Effects 0.000 abstract 1
- 239000002609 medium Substances 0.000 description 12
- 241000234295 Musa Species 0.000 description 9
- 230000000442 meristematic effect Effects 0.000 description 8
- 210000001519 tissue Anatomy 0.000 description 8
- 239000006870 ms-medium Substances 0.000 description 7
- 230000005540 biological transmission Effects 0.000 description 6
- 230000000694 effects Effects 0.000 description 5
- 244000037666 field crops Species 0.000 description 5
- 235000019504 cigarettes Nutrition 0.000 description 4
- 230000018044 dehydration Effects 0.000 description 4
- 238000006297 dehydration reaction Methods 0.000 description 4
- VGGSQFUCUMXWEO-UHFFFAOYSA-N Ethene Chemical compound C=C VGGSQFUCUMXWEO-UHFFFAOYSA-N 0.000 description 3
- 244000025221 Humulus lupulus Species 0.000 description 3
- 235000008694 Humulus lupulus Nutrition 0.000 description 3
- 230000035584 blastogenesis Effects 0.000 description 3
- 230000032823 cell division Effects 0.000 description 3
- 239000011159 matrix material Substances 0.000 description 3
- 238000011169 microbiological contamination Methods 0.000 description 3
- 241000894006 Bacteria Species 0.000 description 2
- 230000000844 anti-bacterial effect Effects 0.000 description 2
- 230000002155 anti-virotic effect Effects 0.000 description 2
- 238000004140 cleaning Methods 0.000 description 2
- 201000010099 disease Diseases 0.000 description 2
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 2
- 238000005516 engineering process Methods 0.000 description 2
- 230000004060 metabolic process Effects 0.000 description 2
- 235000015097 nutrients Nutrition 0.000 description 2
- 239000005631 2,4-Dichlorophenoxyacetic acid Substances 0.000 description 1
- 206010064571 Gene mutation Diseases 0.000 description 1
- 238000009825 accumulation Methods 0.000 description 1
- 230000000840 anti-viral effect Effects 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 230000008901 benefit Effects 0.000 description 1
- 230000015572 biosynthetic process Effects 0.000 description 1
- 238000005253 cladding Methods 0.000 description 1
- 238000005520 cutting process Methods 0.000 description 1
- 238000009826 distribution Methods 0.000 description 1
- 210000002615 epidermis Anatomy 0.000 description 1
- 239000011521 glass Substances 0.000 description 1
- 238000000338 in vitro Methods 0.000 description 1
- 230000002779 inactivation Effects 0.000 description 1
- 238000011534 incubation Methods 0.000 description 1
- 208000015181 infectious disease Diseases 0.000 description 1
- 230000002503 metabolic effect Effects 0.000 description 1
- 244000005700 microbiome Species 0.000 description 1
- 230000004899 motility Effects 0.000 description 1
- 230000005305 organ development Effects 0.000 description 1
- 239000010451 perlite Substances 0.000 description 1
- 235000019362 perlite Nutrition 0.000 description 1
- 239000000575 pesticide Substances 0.000 description 1
- 230000035755 proliferation Effects 0.000 description 1
- 230000000644 propagated effect Effects 0.000 description 1
- 239000013049 sediment Substances 0.000 description 1
- 229960005486 vaccine Drugs 0.000 description 1
- 239000010455 vermiculite Substances 0.000 description 1
- 235000019354 vermiculite Nutrition 0.000 description 1
- 229910052902 vermiculite Inorganic materials 0.000 description 1
- 238000005406 washing Methods 0.000 description 1
Landscapes
- Breeding Of Plants And Reproduction By Means Of Culturing (AREA)
- Cultivation Of Plants (AREA)
Abstract
The invention provides a kind of preparation method of Sweetpotato Viruses Elimination seedling, put together with the banana of maturation after being basked seeds by potato block 3-5 days, then by potato block plantation vernalization in incubator in sandy soil, arranges higher temperature.Get bud seedling top 1-2cm long, cut off macroscopic blade, carry out surface sterilization and use rinsed with sterile water.Strip the stem apex with l-2 leaf primordium, be inoculated on corresponding medium, differentiation and the somatic embryo of difference evoking adventive bud occur.Viral diagnosis is carried out to regeneration plant, virus-free seedling through squamous subculture, hardening, directly plant after plastic tunnel, broad irrigation.Transplanting initial 2 Zhou Zhongwu prevents the sun directly to shine, and spreads fertilizer over the fields urea and water immediately and namely obtain detoxic seedling after transplanting 3 weeks.Detoxification test tube plantlet plastic tunnel of directly planting after domestication can be significantly improved survival rate by the present invention, decreases operation sequence, and do not need the process through the domestication of domestication room, use manpower and material resources sparingly financial resources.
Description
Technical field
The invention belongs to sweet potato seedling Cultivating techniques field, be specifically related to a kind of preparation method of Sweetpotato Viruses Elimination seedling.
Background technology
Sweet potato is one of China 4 generalized grain crop, and sweet potato is asexually propagated crop, and susceptible viral infects in the course of cultivation, is just difficult to remove once infection.The accumulation of virus can cause Different Varieties to degenerate, and quality and yield reduces, and even loses commodity value, produces cause serious harm to sweet potato.There is no the practical sweet potato variety bringing out high antiviral disease so far, also without the highly effective pesticide preventing and treating virus disease both at home and abroad.The distribution of virus in plant corpus is uneven, from the tip of a root and stem apex more near, viral density is lower; Otherwise, higher.
And meristematic tissue is generally virus-free infects.Why stem apex (and tip of a root) meristematic tissue is not with virus to have following reason, the first, and the route of transmission one of virus in plant corpus is by skeleton, and not yet forms skeleton in meristematic tissue; Two is by protoplasmic connection, but this approach virus movement speed slowly, is difficult to the meristematic tissue pulling up to active growth.The second, in the meristematic tissue of vigorous division, metabolic activity is very high, and virus cannot be copied.3rd, in stem apex, there is high-level endogenous hormones, the propagation of virus can be suppressed.4th, in plant corpus, have one " virus inactivation system ", its activity in meristematic tissue should be the highest, thus makes meristematic tissue do not infected.Therefore stem-apex Meristem culture can be utilized to produce detoxic seedling.If but only got sediment diversion ratio, would often not easily survive.
Utilize sweet potato stem tip to cultivate and produce the existing research of detoxic seedling and report, but in the past the stem of multiplex land for growing field crops or greenhouse culture climing on draw materials and strip stem apex; Also the stem apex getting in vitro cuttings plant is had to make virus-free culture material; Also the vernalization of useful potato block is drawn materials and is stripped Shoot Tip Culture, but is also vernalization under normal temperature conditions.Select these methods above-mentioned, grow under normal conditions and cultivate, because the Material growth for detoxification is slower, metabolism is not strong, and strip Shoot Tip Culture detoxification efficiency poor, stem apex is bad to be stripped (because leaf primordium internode is short, leaf primordium is crowded together), and regeneration rate is low.
In addition, detoxification test tube plantlet rooting culture is extremely important, generally selects turfy soil, vermiculite, perlite etc. as transplanting medium, carries out domestication cultivation and could transplant plastic tunnel or greenhouse in about 3 weeks in domestication room.At substantial manpower, material resources and financial resources are cultivated in test-tube plantlet domestication, and poor growth, easily microbiological contamination cause survival rate to reduce.And need transplant through 2 times, complex operation.
Summary of the invention
The invention provides a kind of preparation method of Sweetpotato Viruses Elimination seedling, can solve the detoxification efficiency that prior art exists poor, stem apex is bad to be stripped, and regeneration rate is low, and detoxic seedling rooting culture complex operation, survival rate be low, grow slow problem.
For reaching the object solved the problems of the technologies described above, the present invention is achieved by the following technical solutions:
A preparation method for Sweetpotato Viruses Elimination seedling, it comprises the following steps:
(1) choose free from insect pests undamaged sweet potato potato block, be exposed to the sun under the sun 2 ~ 5 days;
(2) put together the potato block after being exposed to the sun and ripe banana 3-5 days;
(3) potato block clear water is cleaned, potato block covers sand, put into incubator and carry out vernalization, temperature 35 ~ 37 DEG C; Dark culturing 1 week before sprouting, after sprouting, every day 13 h light, 1000-2000lx;
(4) when bud seedling grows to more than 10cm, get top 1-2cm long, cut off macroscopic blade, first rinse with clear water, then carry out surface sterilization, finally use rinsed with sterile water 3-5 time;
(5) under stereoscopic anatomical lens, strip the stem apex with l-2 leaf primordium, be inoculated on the medium of interpolation 0.1-0.2mg/LNAA and 1.0-2.0mg/LBAP, cultivation temperature is 25-27 DEG C, every day 13 h light, 2000-3000lx, cultivates 3-4 week, the differentiation of evoking adventive bud;
Or described stem apex is inoculated into add 0.2-2.0mg/L2,4-D medium on dark culturing, cultivation temperature is 25-27 DEG C, and somatic embryos fetal hair is raw;
(6) cultivate on the medium adding 0.1-0.2mg/LNAA and 1.0-2.0mg/LBAP, when regenerating seedling and growing to more than 2cm, cut from base portion, be transferred in the medium not adding hormone and obtain complete detoxic seedling;
Or cultivate 4-5 week on the medium adding 0.2-2.0mg/L2,4-D after, explant successively starts to form embryo callus and body embryo, is transferred to by the explant of organizator embryo and does not add in the medium of hormone, body embryo germination is impelled to form detoxic seedling;
(7) stem climing segment method Fast-propagation is adopted to obtain detoxification test tube plantlet;
(8) domestication of detoxification test tube plantlet: Plantlet subculture is cultivated 1 month, opens bottleneck and tames;
(9) transplanting of detoxification test tube plantlet: test-tube plantlet is cleaned medium, a point individual plant is directly planted in the soil of plastic tunnel, and plastic tunnel two adds fly net;
(10) detoxification test tube plantlet is planted after in the soil of booth, first water-sprinkling, then broad irrigation;
(11) 2 Zhou Zhongwu that detoxicating cuvette transplantation of seedlings is initial take shading screen, and prevent the sun directly to shine, remove shading screen after 2 weeks, noon leaks informaton;
(12) after transplanting 3 weeks, spread fertilizer over the fields urea, water immediately, promote growth of seedling;
(13) breeding of detoxic seedling in plastic tunnel: grow to more than 30cm when stem is climing, shears that stem is climing plants in plastic tunnel, carries out breeding again of detoxic seedling, cut at every turn climing after, spread fertilizer over the fields urea, water immediately, namely obtain Sweetpotato Viruses Elimination seedling.
Compared with prior art, advantage of the present invention and good effect are:
1, potato block is put together with ripe banana and is processed 3-5 days by the present invention, utilizes ripe banana to discharge ethene, impels potato block early to germinate, because just the potato block of results germinates slow, and causes easy rotten potato owing to germinateing slow.
2, the present invention utilizes high temperature vernalization and Shoot Tip Culture to combine and produces Sweetpotato Viruses Elimination seedling, vernalization arranges higher temperature (35-37 DEG C), according to virus to sensitive, sweet potato is high temperature resistant (because sweet potato is torrid zone origin plant), utilize this difference to select suitable temperature, sweet potato body inner virus concentration can be made to reduce, and transmission speed slows down or loses activity, and sweet potato meristem cell division is accelerated, potato blastogenesis is long to be accelerated.Therefore the stem apex detoxify stripped is effective, and because leaf primordium internode elongates, stem apex easily strips (because leaf primordium internode elongates, easily sees meristematic tissue under anatomical lens, stem apex easily strips), regeneration rate high (because higher temperature growth metabolism is vigorous).The results showed, in incubator, after vernalization 8-10 days, potato BOB(beginning of block) sprouts, and just can draw materials after 11-13 days strips stem apex.When stripping stem apex under anatomical lens, because leaf primordium internode is long, be easy to stripping.By adventitious organogenesis, the frequency of stem apex regeneration plant reaches more than 90%.
3, carry out the vernalization of potato block after sweet potato rewarding autumn of the present invention at once, strip stem apex and cultivation, detoxic seedling can be obtained early.Through tube rapid propagation, Second Year plastic tunnel after domestication is bred and is directly planted land for growing field crops, decreases the time of cultivating in blake bottle, thus the cultivation caused because incubation time is long can be avoided to make a variation.Because if when the detoxic seedling obtaining regeneration is more late, Second Year detoxic seedling has little time to plant land for growing field crops, and wait the 3rd year ability plantation land for growing field crops, the time of cultivating in blake bottle is like this elongated, and easily produces gene mutation.
4, plastic tunnel of directly being planted after domestication by detoxification test tube plantlet can significantly improve survival rate.Be all first little transplantation of seedlings is placed in nutrient matrix domestication room in the past, after surviving, transplanted plastic tunnel or glass greenhouse again.This process approximately needs 3 time-of-weeks.And the present invention is by little seedling direct transplantation in the soil of plastic tunnel, can avoid transplanting easy microbiological contamination in matrix, the difficulty that survival rate is low.Because the microorganism in soil is in poised state, beneficial bacterium can suppress the procreation of harmful bacteria.And in nutrient matrix, due to the more weak very easy microbiological contamination of test-tube plantlet, thus be lowered into motility rate.Survival rate of the present invention can reach more than 95%.
5, by detoxicating cuvette seedling direct transplantation booth, decrease operation sequence, do not need the process through the domestication of domestication room, use manpower and material resources sparingly financial resources.And owing to reducing program, directly grow in soil, decrease the impact of transplanting seedling.Due to the direct planting soil of seedling, root system extending space is large, well established and vigorously developing, well developed root system, impels aerial growth healthy and strong soon.Transplant little seedling leaf after 9 days to stretch, and grown new blade, after 3 weeks, main climing beginning extends, and grows lateral bine, and after 5 ~ 6 weeks, stem is climing grows to 30 ~ more than 40cm, can shear the climing cuttage of stem and breed.
Accompanying drawing explanation
Fig. 1 cultivates by sweet potato stem tip the indefinite bud that the callus that formed grows in the present invention, left: business's potato 19; Right: Xu-shu No.22.
Fig. 2 is the whole plant figure grown up to by axillalry bud in the present invention.
Fig. 3 is the detoxification test tube plantlet after washing medium in the present invention off.
Fig. 4 is that in the present invention, detoxic seedling transplants process.
Fig. 5 is the detoxic seedling of just having transplanted in the present invention in plastic tunnel soil.
Fig. 6, in the present invention, plastic tunnel takes shading screen.
Fig. 7 is the detoxic seedling of transplanting in the present invention after 9 days.
Fig. 8 is the detoxic seedling of transplanting in the present invention 3 weeks.
Fig. 9 is the detoxic seedling of transplanting in the present invention 6 weeks.
Figure 10 cultivates by sweet potato stem tip the embryo callus formed in the present invention.
Figure 11 is by the somatic embryo that embryo callus is formed in the present invention.
Figure 12 is the somatic embryo sprouted in the present invention.
Figure 13 is the detoxification test tube plantlet grown up to after being sprouted by somatic embryo in the present invention.
Figure 14 is that in the present invention, height is the potato block of No. 14 detoxic seedling knots.
Embodiment
Below in conjunction with drawings and Examples, technical scheme of the present invention is described further.
Embodiment 1
The preparation method of Sweetpotato Viruses Elimination seedling of the present invention specifically comprises the steps:
1. the process before the vernalization of sweet potato potato block:
(1), after results in autumn, choose current main popularization sweet potato variety Xu-shu No.22 number, cigarette potato 25, business's potato 19 (commercially available kind) free from insect pests, undamaged potato block, being exposed to the sun under the sun, (first had bactericidal action in 2-3 days; Second can promote the rudiment of potato block).
(2) ripe banana is prepared, potato block after being exposed to the sun and ripe banana are put together and is put in plastics collection case, build lid, place 3-5 days (room temperature is not less than 13 DEG C) in laboratory and (utilize ripe banana to discharge ethene, promote the rudiment of potato block.Because just the potato block of results germinates comparatively slow, cause easy rotten potato).
2. vernalization:
Cleaned by above-mentioned potato block clear water, put into sand, on potato block, cladding thickness is the sand of 1-2cm, puts into incubator and carries out vernalization.Temperature arranges 35 ~ 37 DEG C.(according to virus to sensitive, because sweet potato is torrid zone origin plant, so sweet potato is high temperature resistant plant, utilize this difference, the present invention is through the suitable high temperature of experimental selection, and sweet potato body inner virus concentration can be made to reduce, and transmission speed slows down, and sweet potato meristem cell division is accelerated, potato blastogenesis is long to be accelerated.Therefore the stem apex detoxify stripped is effective, and due to the elongation of leaf primordium internode, stem apex easily strips, and regeneration rate is high).Dark culturing 1 week before sprouting, after sprouting, every day 13 h light, 1000-2000lx.Humidity according to sand is sprayed water in right amount, and in sand, water content is unsuitable excessive, otherwise easy rotten kind.Cultivate 8-10 days in incubator after, potato BOB(beginning of block) sprouts, and the bud seedling had after 11 ~ 13 days is long reaches 10cm.
3. stem apex strips and cultivation
(1) sterilization of material: when bud seedling grows to more than 10cm, get top 1-2cm long, cut off macroscopic blade, first rinse with clear water, then in super quiet workbench with 70% alcohol-pickled 10-20 second, soak 5 points with the liquor natrii hypochloritis that mass percent is 2% again and carry out surface sterilization in-7 minutes, finally use rinsed with sterile water 3-5 time.
(2) stem apex strips and cultivation: under stereoscopic anatomical lens, strip the stem apex with l-2 leaf primordium, be inoculated into add 0.2mg/LNAA and 2.0mg/LBAP MS medium on cultivate.Cultivate stem apex after 1 week to start to form callus, start after 3-4 week to form indefinite bud from callus wound, as shown in Figure 1.Adventitious bud formation frequency lists in table 1.3 experimental cultivar stem apexs form the frequency of indefinite bud all higher than 90%.
Table 1 sweet potato different cultivars Shoot Tip Culture shoot regeneration frequency
| Kind | Xu-shu No.22 number | Cigarette potato 25 | Business's potato 19 |
| Regeneration rate (%) | 95.6 | 92.3 | 93.8 |
The medium adding 0.2mg/LNAA and 2.0mg/LBAP is cultivated after one month, when regenerating seedling and growing to more than 2cm, cut from base portion, be transferred in the MS medium not adding hormone and all grow up to complete detoxic seedling.
Condition of culture is 25-27 DEG C, every day 13 h light, 2000-3000lx.
4. the Viral diagnosis of detoxic seedling
Carry out Viral diagnosis to detoxic seedling, testing result is in table 2, and 3 kind virus elimination rates all reach 100%.
Table 2 sweet potato different cultivars regeneration plant virus elimination rate
| Kind | Xu-shu No.22 number | Cigarette potato 25 | Business's potato 19 |
| Virus elimination rate (%) | 100 | 100 | 100 |
5. the aseptic Fast-propagation of detoxic seedling
The breeding of detoxic seedling adopts the climing segment method of stem, and be 1 segment by a detoxic seedling 1-2 leaf segment, cuttage enters not add in the MS medium of hormone, axillary bud sprouting, within 1 month, may have grown into whole plant again, as shown in Figure 2.So constantly segment proliferation and subculture is cultivated, reproduction speed with geometric growth, can in blake bottle amount reproduction detoxic seedling.Condition of culture is 25-27 DEG C, every day 13 h light, 2000-3000lx.
6. the domestication of detoxification test tube plantlet and transplanting:
(1) the squamous subculture test-tube plantlet of 1 month, opens bottleneck and tames, and first day slightly opens some mouths, within second day, open again a bit, so continuous hardening 2-3 days, until last bottleneck opens half (because the hardening time is short, all opens easy dehydration and wilt).
(2) test-tube plantlet is cleaned medium, a point individual plant is directly planted in the soil of plastic tunnel, and strain spacing 20 ~ 25cm, as shown in Fig. 3,4,5.Note not hindering root in cleaning and transplanting process as far as possible, can transplanting seedling time be shortened like this, and little shoot survival percent and growth rate can be improved.Plastic tunnel two add fly net (for pre-anti-virus infects again, because virus mainly insect transmission, add fly net can in advance protection against insect enter plastic tunnel).
(3) detoxification test tube plantlet is planted after in the soil of booth, first water-sprinkling (preventing seedling from having been rushed), then broad irrigation.The moisture that the effect first of broad irrigation can provide seedling to need, second can provide safeguard for keeping the air humidity in plastic tunnel.Because test-tube plantlet air humidity in test tube is very large, transplant and should keep certain air humidity early stage, otherwise easily dehydration is wilted.
7. the management of detoxic seedling in plastic tunnel:
(1) initial 2 Zhou Zhongwu of detoxicating cuvette transplantation of seedlings (i.e. at 10 in the morning to afternoon 3 point) take shading screen, prevent the sun directly to shine, as shown in Figure 6.And after taking shading screen, the temperature of not leaking informaton in plastic tunnel noon also can not be too high, can keep the air humidity in plastic tunnel.Remove shading screen after 2 weeks, noon leaks informaton.Water according to soil moisture content.
(2) after transplanting 3 weeks, spread fertilizer over the fields urea, fertilizing amount, by 8 ~ 10 jin every mu, is watered immediately, promotes growth of seedling.The present invention is due to the direct planting soil of seedling, and root system extending space is large, well established and vigorously developing, well developed root system, impels aerial growth healthy and strong soon.Transplant and observe after 9 days, little seedling leaf stretches, and has grown new blade, and as shown in Figure 7, statistics survival rate is in table 3, and 3 experimental cultivar survival rates are all more than 95%.After transplanting 3 weeks, main climing beginning extends, and some plant have grown lateral bine, as shown in Figure 8.To transplant after 5 ~ 6 weeks that stem is climing grows to 30-40cm, as shown in Figure 9.
Table 3 detoxic seedling adds up survival rate after transplanting 9 days
| Kind | Transplanted seedling number | Become number of live vaccine | Survival rate (%) |
| Xu-shu No.22 | 240 | 231 | 96.25 |
| Cigarette potato 25 | 113 | 109 | 96.46 |
| Business's potato 19 | 148 | 142 | 95.95 |
8. the breeding of detoxic seedling in plastic tunnel:
Grow to more than 30cm when stem is climing, shear the climing cuttage of stem in the soil of plastic tunnel, carry out breeding again of detoxic seedling, cultivation space between plants is 20 ~ 25cm, cut at every turn climing after, spread fertilizer over the fields urea, fertilizing amount also by 8 ~ 10 jin every mu, is watered immediately, can obtain a large amount of healthy and strong detoxic seedling.
Embodiment 2
Described in the present embodiment, the preparation method of Sweetpotato Viruses Elimination seedling specifically comprises the steps:
1. the process before the vernalization of sweet potato potato block:
(1), after results in autumn, choosing sweet potato variety high is No. 14 (commercially available kind) frees from insect pests, undamaged potato block, and being exposed to the sun under the sun, (first had bactericidal action in 2-3 days; Second can promote the rudiment of potato block).
(2) ripe banana is prepared, potato block after being exposed to the sun and ripe banana are put together and is put in plastics collection case, build lid, place 3-5 days (room temperature is not less than 13 DEG C) in laboratory and (utilize ripe banana to discharge ethene, promote the rudiment of potato block.Because just the potato block of results germinates comparatively slow, cause easy rotten potato).
2. vernalization:
Above-mentioned potato block clear water is cleaned, puts into sand, potato block covers the sand of about 1-2cm, put into incubator and carry out vernalization.Temperature arranges 35 ~ 37 DEG C.(according to virus to sensitive, because sweet potato is torrid zone origin plant, so sweet potato is high temperature resistant plant, utilize this difference, select suitable high temperature, sweet potato body inner virus concentration can be made to reduce, transmission speed slows down, and sweet potato meristem cell division is accelerated, and potato blastogenesis is long to be accelerated.Therefore the stem apex detoxify stripped is effective, and due to the elongation of leaf primordium internode, stem apex easily strips, and regeneration rate is high.) sprout before dark culturing 1 week, after sprouting, every day 13 h light, 1000-2000lx.Humidity according to sand is sprayed water in right amount, and in sand, water content is unsuitable excessive, otherwise easy rotten kind.In incubator, cultivate potato BOB(beginning of block) after 10 days sprout, the bud had after 13 days is long reaches 10cm.
3. stem apex strips and cultivation
(1) sterilization of material: when bud seedling grows to about 10cm, get top 1-2cm long, cut off macroscopic blade, first rinse with clear water, then in super quiet workbench with 70% alcohol-pickled 10-20 second, the liquor natrii hypochloritis of 2% soaks 5 points and carries out surface sterilization in-7 minutes, finally uses rinsed with sterile water 3-5 time.
(2) stem apex strips and cultivation: under stereoscopic anatomical lens, strip the stem apex with l-2 leaf primordium, be inoculated into add 0.2-2.0mg/L2,4-D MS medium on cultivate (present case 2,4-D concentration is 2.0mg/L).Start to form embryo callus after cultivating 3 weeks, as shown in Figure 10, the frequency that stem apex forms embryo callus is 76.7%.From organizator embryo embryo callus after cultivation 4-5 week, as shown in figure 11, cultivate at 25-27 DEG C, carry out under dark condition.
Being transferred to by the explant of organizator embryo does not add in the MS medium of hormone, body embryo germination seedling, as shown in figure 12.When regenerating seedling and growing to more than 2cm, cut to be transferred on new MS medium and grow up to whole plant, as shown in figure 13.Condition of culture is 25-27 DEG C, every day 13 h light, 2000-3000lx.
4. the aseptic Fast-propagation of detoxic seedling
The breeding of detoxic seedling adopts the climing segment method of stem, and be 1 segment by a detoxic seedling 1-2 leaf segment, cuttage enters not add in the MS medium of hormone, axillary bud sprouting, within 1 month, may have grown into again complete detoxic seedling, as shown in Figure 2.So constantly segment Multiplying culture, reproduction speed with geometric growth, can in blake bottle amount reproduction detoxic seedling.Condition of culture is 25-27 DEG C, every day 13 h light, 2000-3000lx.
5. the domestication of detoxification test tube plantlet and transplanting:
(1) the squamous subculture test-tube plantlet of 1 month, opens bottleneck and tames, and first day slightly opens some mouths, within second day, open again a bit, so continuous hardening 2-3 days, until last bottleneck opens half (because the hardening time is short, all opens easy dehydration and wilt).
(2) test-tube plantlet is cleaned medium, a point individual plant is directly planted in the soil of plastic tunnel, and strain spacing 20 ~ 25cm, as shown in Fig. 3,4,5.Note not hindering root in cleaning and transplanting process as far as possible, can transplanting seedling time be shortened like this, and little shoot survival percent and growth rate can be improved.Plastic tunnel two add fly net (for pre-anti-virus infects again, because virus mainly insect transmission, add fly net can in advance protection against insect enter plastic tunnel).
(3) detoxification test tube plantlet is planted after in the soil of booth, first water-sprinkling (preventing seedling from having been rushed), then broad irrigation.The moisture that the effect first of broad irrigation can provide seedling to need, second can provide safeguard for keeping the air humidity in plastic tunnel.Because test-tube plantlet air humidity in test tube is very large, transplant and should keep certain air humidity early stage, otherwise easily dehydration is wilted.
6. the management of detoxic seedling in plastic tunnel:
(1) initial 2 Zhou Zhongwu of detoxicating cuvette transplantation of seedlings (i.e. at 10 in the morning to afternoon 3 point) take shading screen, prevent the sun directly to shine, as shown in Figure 6.And after taking shading screen, the temperature of not leaking informaton in plastic tunnel noon also can not be too high, can keep the air humidity in plastic tunnel.Remove shading screen after 2 weeks, noon leaks informaton.Water according to soil moisture content.
(2) after transplanting 3 weeks, spread fertilizer over the fields urea, fertilizing amount, by 8 ~ 10 jin every mu, is watered immediately, promotes growth of seedling, promotes growth of seedling.
Transplant and observe after 9 days, little seedling leaf stretches, and has grown new blade, and as shown in Figure 7, statistics survival rate is 96%.After transplanting 3 weeks, main climing beginning extends, and some plant have grown lateral bine, as shown in Figure 8.To transplant after 5 ~ 6 weeks that stem is climing grows to 30-40cm, as shown in Figure 9.
7. the breeding of detoxic seedling in plastic tunnel:
Grow to more than 30cm when stem is climing, shear the climing cuttage of stem in the soil of plastic tunnel, carry out breeding again of detoxic seedling, plant space between plants 20 ~ 25cm, cut at every turn climing after, spread fertilizer over the fields urea, fertilizing amount also by 8 ~ 10 jin every mu, is watered immediately, can obtain a large amount of healthy and strong detoxic seedling.
Behind the Virus-free Sweetpotato transplantation of seedlings land for growing field crops in plastic tunnel, carry out conventional cultivation management, potato seedling robust growth is vigorous, vane extension, and the present invention does not all observe variation plant for 4 kinds of examination.Autumn observes after results, and potato block epidermis smooth and beautiful appearance, adds commodity value, as shown in figure 14.
Above embodiment only in order to technical scheme of the present invention to be described, but not is limited; Although with reference to previous embodiment to invention has been detailed description, for the person of ordinary skill of the art, still can modify to the technical scheme described in previous embodiment, or equivalent replacement is carried out to wherein portion of techniques feature; And these amendments or replacement, do not make the essence of appropriate technical solution depart from the spirit and scope of the present invention's technical scheme required for protection.
Claims (7)
1. a preparation method for Sweetpotato Viruses Elimination seedling, is characterized in that, it comprises the following steps:
(1) choose free from insect pests undamaged sweet potato potato block, be exposed to the sun under the sun 2 ~ 5 days;
(2) put together the potato block after being exposed to the sun and ripe banana 3-5 days;
(3) potato block clear water is cleaned, potato block covers sand, put into incubator and carry out vernalization, temperature 35 ~ 37 DEG C; Dark culturing 1 week before sprouting, after sprouting, every day 13 h light, 1000-2000lx;
(4) when bud seedling grows to more than 10cm, get top 1-2cm long, cut off macroscopic blade, first rinse with clear water, then carry out surface sterilization, finally use rinsed with sterile water 3-5 time;
(5) under stereoscopic anatomical lens, strip the stem apex with l-2 leaf primordium, be inoculated on the medium of interpolation 0.1-0.2mg/LNAA and 1.0-2.0mg/LBAP, cultivation temperature is 25-27 DEG C, every day 13 h light, 2000-3000lx, cultivates 3-4 week, the differentiation of evoking adventive bud;
Or described stem apex is inoculated into add 0.2-2.0mg/L2,4-D medium on dark culturing, cultivation temperature is 25-27 DEG C, and somatic embryos fetal hair is raw;
(6) cultivate on the medium adding 0.1-0.2mg/LNAA and 1.0-2.0mg/LBAP, when regenerating seedling and growing to more than 2cm, cut from base portion, be transferred in the medium not adding hormone and obtain complete detoxic seedling;
Or cultivate 4-5 week on the medium adding 0.2-2.0mg/L2,4-D after, explant successively starts to form embryo callus and body embryo, is transferred to by the explant of organizator embryo and does not add in the medium of hormone, body embryo germination is impelled to form detoxic seedling;
(7) stem climing segment method Fast-propagation is adopted to obtain detoxification test tube plantlet;
(8) domestication of detoxification test tube plantlet: Plantlet subculture is cultivated 1 month, opens bottleneck and tames;
(9) transplanting of detoxification test tube plantlet: test-tube plantlet is cleaned medium, a point individual plant is directly planted in the soil of plastic tunnel, and plastic tunnel two adds fly net;
(10) detoxification test tube plantlet is planted after in the soil of booth, first water-sprinkling, then broad irrigation;
(11) 2 Zhou Zhongwu that detoxicating cuvette transplantation of seedlings is initial take shading screen, and prevent the sun directly to shine, remove shading screen after 2 weeks, noon leaks informaton;
(12) after transplanting 3 weeks, spread fertilizer over the fields urea, water immediately, promote growth of seedling;
(13) breeding of detoxic seedling in plastic tunnel: grow to more than 30cm when stem is climing, shears that stem is climing plants in plastic tunnel, carries out breeding again of detoxic seedling, cut at every turn climing after, spread fertilizer over the fields urea, water immediately, namely obtain Sweetpotato Viruses Elimination seedling.
2. the preparation method of Sweetpotato Viruses Elimination seedling according to claim 1, is characterized in that: the temperature that the potato block after being exposed to the sun in described step (2) and ripe banana are placed is not less than 13 DEG C.
3. the preparation method of Sweetpotato Viruses Elimination seedling according to claim 1, is characterized in that: the thickness covering sand in described step (3) is 1-2cm.
4. the preparation method of Sweetpotato Viruses Elimination seedling according to claim 1, is characterized in that: with 70% alcohol-pickled 10-20 second in described step (4), the liquor natrii hypochloritis of 2% soaks and carries out surface sterilization in 5 minutes-7 minutes.
5. the preparation method of Sweetpotato Viruses Elimination seedling according to claim 1, is characterized in that: in described step (6) and (7), condition of culture is temperature 25-27 DEG C, every day 13 h light, 2000-3000lx.
6. the preparation method of Sweetpotato Viruses Elimination seedling according to claim 1, is characterized in that: the strain spacing of planting in described step (9) and (13) is 20 ~ 25cm.
7. the preparation method of Sweetpotato Viruses Elimination seedling according to claim 1, is characterized in that: the fertilizing amount of spreading fertilizer over the fields urea in described step (12) and (13) is 8 ~ 10 jin every mu.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201410291953.8A CN104067821B (en) | 2014-06-26 | 2014-06-26 | A kind of preparation method of Sweetpotato Viruses Elimination seedling |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201410291953.8A CN104067821B (en) | 2014-06-26 | 2014-06-26 | A kind of preparation method of Sweetpotato Viruses Elimination seedling |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN104067821A CN104067821A (en) | 2014-10-01 |
| CN104067821B true CN104067821B (en) | 2016-03-23 |
Family
ID=51589751
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201410291953.8A Expired - Fee Related CN104067821B (en) | 2014-06-26 | 2014-06-26 | A kind of preparation method of Sweetpotato Viruses Elimination seedling |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN104067821B (en) |
Families Citing this family (21)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104429953A (en) * | 2014-11-19 | 2015-03-25 | 西南大学 | Stem tip detoxification method for sweet potato virus seedling |
| CN104719164B (en) * | 2015-03-30 | 2017-06-16 | 青岛农业大学 | A kind of rapid propagation method of Sweetpotato Viruses Elimination original silkworm egg potato |
| CN105340971A (en) * | 2015-11-13 | 2016-02-24 | 刘亚平 | Sweet potato seedling detoxifying solution and preparation method thereof |
| CN105830698A (en) * | 2016-03-29 | 2016-08-10 | 镇远县百福农业科技示范有限公司 | Seedling hardening method for sweet potato tissue-cultured test-tube plantlets |
| CN105900838A (en) * | 2016-04-25 | 2016-08-31 | 吉林师范大学 | Purple sweet potato detoxification method |
| CN106386494B (en) * | 2016-09-29 | 2018-08-24 | 紫云自治县紫香源农林科技有限责任公司 | A kind of sweet potato stem tip detoxification and breeding method |
| CN106508644B (en) * | 2016-10-27 | 2020-05-15 | 南充市农业科学院 | Sweet potato virus-free seed potato propagation method |
| CN107980517A (en) * | 2017-11-22 | 2018-05-04 | 江苏徐淮地区徐州农业科学研究所(江苏徐州甘薯研究中心) | The method for improving cold district sweet potato potato seed emergence rate and growth potential |
| CN108901846A (en) * | 2018-07-12 | 2018-11-30 | 龙岩市农业科学研究所 | A kind of preparation manipulation technology of simple quick Sweetpotato Viruses Elimination seedling |
| CN108552060A (en) * | 2018-07-19 | 2018-09-21 | 江苏徐淮地区徐州农业科学研究所(江苏徐州甘薯研究中心) | The sterilizing methods of sweet potato tissue cultures explant |
| CN109105256A (en) * | 2018-07-30 | 2019-01-01 | 浙江大学 | Sweet potato stem tip poison-removing method |
| CN108739190A (en) * | 2018-08-02 | 2018-11-06 | 山东农业大学 | A kind of cultural method improving fresh type sweet potato root tuber exterior quality |
| CN108935108A (en) * | 2018-09-29 | 2018-12-07 | 河南云帮农业科技有限公司 | A kind of sweet potato tissue-cultured seedling detoxification tissue culture method |
| CN109329026B (en) * | 2018-11-30 | 2021-01-26 | 青岛农业大学 | Method for obtaining sweet potato virus-free seedlings through high-temperature and variable-temperature treatment |
| CN109392721B (en) * | 2018-12-20 | 2021-11-30 | 江苏徐淮地区徐州农业科学研究所(江苏徐州甘薯研究中心) | Method for inducing sweet potato plant regeneration |
| CN112753513B (en) * | 2021-02-05 | 2022-12-23 | 湖北恩施中国南方马铃薯研究中心 | Water culture rapid propagation method of selenium-rich leaf vegetable type sweet potatoes |
| CN113875537B (en) * | 2021-11-04 | 2022-05-13 | 华南农业大学 | Method for promoting branches of cajuput trees by high-temperature treatment |
| CN114051927A (en) * | 2021-11-09 | 2022-02-18 | 延安市农业科学研究所 | Simplified breeding method for virus-free sweet potato seedlings |
| CN113973716A (en) * | 2021-11-26 | 2022-01-28 | 广东海洋大学 | Tissue culture and rapid propagation method of sweet potatoes |
| CN115136769B (en) * | 2022-07-26 | 2024-03-12 | 石家庄市农林科学研究院 | Sweet potato latent virus removal method |
| CN115644063A (en) * | 2022-11-11 | 2023-01-31 | 青岛农业大学 | Rapid propagation method of virus-free raw seedling of sweet potato |
Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101611697A (en) * | 2009-07-13 | 2009-12-30 | 周玉玲 | Sweet potato merchant 19 detoxifying fast breeding technique and cultivation material |
| CN102138531A (en) * | 2011-04-26 | 2011-08-03 | 马宗新 | Rapid propagation technology and culture medium composition of Fuyang sweet potato 24 virus-free plantlet |
Family Cites Families (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| KR100516984B1 (en) * | 2003-06-24 | 2005-09-27 | 동양물산기업 주식회사 | Method for production of seed Dioscorea opposita Thunb. by use of tissue culture technology |
-
2014
- 2014-06-26 CN CN201410291953.8A patent/CN104067821B/en not_active Expired - Fee Related
Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101611697A (en) * | 2009-07-13 | 2009-12-30 | 周玉玲 | Sweet potato merchant 19 detoxifying fast breeding technique and cultivation material |
| CN102138531A (en) * | 2011-04-26 | 2011-08-03 | 马宗新 | Rapid propagation technology and culture medium composition of Fuyang sweet potato 24 virus-free plantlet |
Non-Patent Citations (5)
| Title |
|---|
| 甘薯脱毒技术在福建省的研究与应用;梁金平;《广西农业科学》;20060531;第37卷(第05期);503-505 * |
| 甘薯茎尖培养与脱毒技术研究;何新民等;《广西农业科学》;20090831;第40卷(第08期);964-968 * |
| 紫色甘薯的茎尖培养与脱毒;杨贤松等;《热带作物学报》;20070930;第28卷(第03期);68-73 * |
| 紫色甘薯茎尖脱毒与快繁及试管苗移植技术研究;王碧琴等;《江西科学》;20100430;第28卷(第02期);196-198、202 * |
| 重庆市主栽优良甘薯品种的茎尖培养脱毒研究;邓暑燕等;《西南师范大学学报(自然科学版)》;20050831;第30卷(第04期);711-714 * |
Also Published As
| Publication number | Publication date |
|---|---|
| CN104067821A (en) | 2014-10-01 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN104067821B (en) | A kind of preparation method of Sweetpotato Viruses Elimination seedling | |
| CN101983554B (en) | Reproduction method of Chinese cymbidium seedlings | |
| CN105191805B (en) | A kind of micro-propagation method of tilia miqueliana | |
| CN104719164B (en) | A kind of rapid propagation method of Sweetpotato Viruses Elimination original silkworm egg potato | |
| CN101926259B (en) | Orchid germchit propagating method | |
| CN108157180B (en) | Open type factory rapid propagation method for potato virus-free seedlings | |
| CN101904262B (en) | Ex vitro rooting method of sugarcane test tube plantlets | |
| CN105123529A (en) | Rapid propagation and efficient cultivation method of Bletilla striata | |
| CN103125386B (en) | Industrial horseradish planting method | |
| CN102805035A (en) | Common head cabbage tissue culture method | |
| CN104365318A (en) | Standardized myrtle planting technology | |
| CN110558172A (en) | Strawberry virus-free tissue culture seedling breeding method | |
| CN103651141B (en) | The method that Bo chrysanthemum batch production test tube seedling is the most numerous | |
| CN107197746A (en) | A kind of mating system of China fir field excellent resources | |
| CN108967192A (en) | A kind of Sweetpotato Viruses Elimination bottle seedling acclimatization and transplants method | |
| CN108377911A (en) | The tissue culture mating system of manglietia glauca | |
| CN103843664A (en) | Lycium exsertum tissue culture and rapid propagation method | |
| CN104303765B (en) | The high-yield planting method of the stem of noble dendrobium | |
| CN103430822B (en) | Aquaculture seed reproduction method for micro seed tubers of konjac | |
| CN102919124A (en) | Rapid propagation method for dendrocalamus giganteus tissue culture seedling industrial production | |
| CN104488711A (en) | Method for obtaining intervarietal hybrids of eustoma grandiflorum rapidly | |
| CN103053429B (en) | Method for regenerating semen pharbitidis in vitro embryonic axis plant | |
| CN106172012A (en) | A kind of Fructus Mori tree group culturation rapid propagating technology | |
| CN106069773B (en) | The renovation process of sleep cloth bag tissue cultures | |
| CN110583269A (en) | Production method of scion for grafting camellia oleifera in Hainan region |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| C06 | Publication | ||
| PB01 | Publication | ||
| C10 | Entry into substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| CB03 | Change of inventor or designer information |
Inventor after: Wang Jingshan Inventor after: Guo Baotai Inventor after: Wang Peng Inventor after: Ji Ruirui Inventor after: Kong Xiangyuan Inventor after: Yu Mingyang Inventor after: Zhang Dandan Inventor after: Li Guan Inventor after: Sui Jiong Ming Inventor after: Qiao Lixian Inventor after: Jiang Defeng Inventor after: Sun Shimeng Inventor before: Wang Jingshan Inventor before: Wang Peng Inventor before: Ji Ruirui Inventor before: Sui Jiong Ming Inventor before: Qiao Lixian Inventor before: Guo Baotai Inventor before: Sun Shimeng |
|
| COR | Change of bibliographic data | ||
| C14 | Grant of patent or utility model | ||
| GR01 | Patent grant | ||
| CF01 | Termination of patent right due to non-payment of annual fee |
Granted publication date: 20160323 |