CN105493972A - Method for efficiently removing CTLV (citrus tatter leaf virus) - Google Patents
Method for efficiently removing CTLV (citrus tatter leaf virus) Download PDFInfo
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- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01G—HORTICULTURE; CULTIVATION OF VEGETABLES, FLOWERS, RICE, FRUIT, VINES, HOPS OR SEAWEED; FORESTRY; WATERING
- A01G13/00—Protecting plants
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- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01G—HORTICULTURE; CULTIVATION OF VEGETABLES, FLOWERS, RICE, FRUIT, VINES, HOPS OR SEAWEED; FORESTRY; WATERING
- A01G2/00—Vegetative propagation
- A01G2/30—Grafting
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- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01G—HORTICULTURE; CULTIVATION OF VEGETABLES, FLOWERS, RICE, FRUIT, VINES, HOPS OR SEAWEED; FORESTRY; WATERING
- A01G7/00—Botany in general
- A01G7/06—Treatment of growing trees or plants, e.g. for preventing decay of wood, for tingeing flowers or wood, for prolonging the life of plants
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Abstract
The invention belongs to the field of citrus planting and particularly relates to a method for efficiently removing a CTLV (citrus tatter leaf virus). The method comprises steps as follows: (1) a citrus plant infected with the CTLV is subjected to heat treatment by the aid of the pulse temperature; (2) stem tips of the citrus plant well treated in the step (1) are grafted. By means of the method, more non-toxic tender stem tips can be germinated under the condition that normal growth of the plant is maintained.
Description
Technical field
The invention belongs to oranges and tangerines field of planting, be specifically related to the method for the broken mosaic virus of a kind of efficient removal oranges and tangerines.
Background technology
The broken leaf disease of oranges and tangerines is by the broken mosaic virus (Citrustatterleafvirus of oranges and tangerines, CTLV) a kind of serious plant disease caused, 1962 by Wallace and Drake at California, USA Late Cambrian (Wallace & Drake, 1962), generation (Miyakawaetal., 1976 are all had at present in countries such as Australia, Japan, South Africa, China; Maraisetal., 1986; Zhangetal., 1988; Broadbentetal., 1994).CTLV propagates mainly through machinery, grafting and juice friction, also can seed dispersal (Roistacheretal., 1980).It is not affine that it can cause trifoliate orange and hybrid thereof to make the mandarin tree grafting mouth of stock, and yellowing leaf, even causes whole strain plant death (Grout, B.W.W., 1990) time serious.Due to virus type disease systemic infection and there is no effective methods for the treatment of, generally should to put prevention first.The common practice of our times various countries prevention citrus virus type disease adopts Shoot-tip Grafting or thermal treatment to add Shoot-tip Grafting mode to remove virus, then plants virus-free nursery stock (RoistacherandKitto, 1977).The method that early stage researchers' Successful utilization thermal treatment and Shoot-tip Grafting combine removes the broken mosaic virus of oranges and tangerines (Koizumi, 1986; He, 1993; Songetal., 1994; Heetal., 1996; Zhang, 1997), but lack the research for variations in temperature and CTLV relation with contents, also find the problem because the not resistance to long heat treatment of oranges and tangerines plant causes detoxification to postpone in practical operation.
Therefore, a kind of method of efficient removal CTLV is badly in need of.
Summary of the invention
In order to overcome problem existing in prior art, the object of the present invention is to provide the method for the broken mosaic virus of a kind of efficient removal oranges and tangerines.
To achieve these goals and other relevant objects, the present invention adopts following technical scheme:
The invention provides the method for the broken mosaic virus of a kind of efficient removal oranges and tangerines, shown method comprises the steps:
(1) pulse temperature is adopted to heat-treat the oranges and tangerines plant having infected the broken mosaic virus of oranges and tangerines;
(2) stem apex of the oranges and tangerines plant handled well in step (1) is adopted to carry out Shoot-tip Grafting.
Preferably, in step (1), described oranges and tangerines plant is Fengjie sweet orange plant.
Further preferably, in step (1), described oranges and tangerines plant is Fengjie 72-1 navel orange plant.
Preferably, in step (1), described employing pulse temperature is heat-treated, can specifically: the oranges and tangerines plant having infected the broken mosaic virus of oranges and tangerines is first processed a period of time T1 under 40 DEG C of conditions, and the scope of described T1 is 2 ~ 6h; Then continue to process a period of time T2 under 25 ~ 30 DEG C of conditions, the scope of described T2 is 2 ~ 6h; Alternate cycles process like this 30 ~ 50 days.More preferably, alternate cycles process like this 40 ~ 50 days.
Further preferably, described employing pulse temperature is heat-treated, can specifically: the oranges and tangerines plant having infected the broken mosaic virus of oranges and tangerines is first processed a period of time T1 under 40 DEG C of conditions, and the scope of described T1 is 2 ~ 6h; Then continue to process a period of time T2 under 27 DEG C of conditions, the scope of described T2 is 2 ~ 6h; Alternate cycles process like this 30 ~ 50 days.More preferably, alternate cycles process like this 40 ~ 50 days.
Further preferably, described employing pulse temperature is heat-treated, can specifically: the oranges and tangerines plant having infected the broken mosaic virus of oranges and tangerines is first processed a period of time T1 under 40 DEG C of conditions, and the scope of described T1 is 2 ~ 6h; Then continue to process a period of time T2 under 25 DEG C of conditions, the scope of described T2 is 2 ~ 6h; Alternate cycles process like this 30 ~ 50 days.More preferably, alternate cycles process like this 40 ~ 50 days.
Further preferably, described employing pulse temperature is heat-treated, can specifically: the oranges and tangerines plant having infected the broken mosaic virus of oranges and tangerines is first processed a period of time T1 under 40 DEG C of conditions, and the scope of described T1 is 2 ~ 6h; Then continue to process a period of time T2 under 30 DEG C of conditions, the scope of described T2 is 2 ~ 6h; Alternate cycles process like this 30 ~ 50 days.More preferably, alternate cycles process like this 40 ~ 50 days.
Further preferably, the scope of T1 described above is 4h; The scope of described T2 is 4h.
In the preferred embodiments of the present invention, the oranges and tangerines plant having infected the broken mosaic virus of oranges and tangerines is first processed a period of time T1 under 40 DEG C of conditions, and the scope of described T1 is 4h; Then continue to process a period of time T2 under 27 DEG C of conditions, the scope of described T2 is 4h; Alternate cycles process like this 40 ~ 50 days.
In the preferred embodiments of the present invention, the oranges and tangerines plant having infected the broken mosaic virus of oranges and tangerines is first processed a period of time T1 under 40 DEG C of conditions, and the scope of described T1 is 4h; Then continue to process a period of time T2 under 25 DEG C of conditions, the scope of described T2 is 4h; Alternate cycles process like this 50 days.
In the preferred embodiments of the present invention, the oranges and tangerines plant having infected the broken mosaic virus of oranges and tangerines is first processed a period of time T1 under 40 DEG C of conditions, and the scope of described T1 is 4h; Then continue to process a period of time T2 under 30 DEG C of conditions, the scope of described T2 is 4h; Alternate cycles process like this 40 days.
Preferably, in step (2), the stem apex adopted comprises the leaf primordium of more than 2 ~ 3.
Compared with prior art, the present invention has following beneficial effect:
(1) conventional high-temperature thermal treatment Citrus Plants can not normal growth, usually causes plant heat injury, sprouts Non-toxic tender shoots quantity few; And technical scheme of the present invention sprouts the Non-toxic tender shoots stem apex of greater number when holding temperature process normal plants.
(2) concrete, the present invention with Fengjie sweet orange plant for object, design two kinds of short time intermittent warming conditions (40 DEG C 4h/25 DEG C 4h and 40 DEG C 4h/30 DEG C 4h), with typical heat treat condition (40 DEG C of 10h/30 DEG C of 14h) in contrast, real-time fluorescence quantitative RT-PCR is utilized to detect CTLV content, adopt Shoot-tip Grafting technology to carry out detoxification, research short time intermittent warming is on the impact of CTLV detoxification.After result shows two kinds of process sweet orange plant 30d, tender shoots germination physiology is obviously faster than contrast, and germination rate is also high than contrast; During 40 DEG C 4h/25 DEG C 4h process sweet orange plant 50d, stem sharp virus elimination rate reaches 60%, during 40 DEG C 4h/30 DEG C 4h process sweet orange plant 40d, stem sharp virus elimination rate reaches 71.43%, there is no detoxification stem sharp during normative heat treatment sweet orange plant 30d, show that two kinds of short time intermittent warming have the effect accelerating to remove CTLV thus.
Accompanying drawing explanation
Fig. 1: the melting curve figure of CTLV and 18sRNA in the first five leaf of intermittent warming sweet orange tender shoots.
Fig. 2 .1: normative heat treatment (40 DEG C of 10h/30 DEG C of 14h) processes the first five leaf CTLV relative expression quantity of 30d tender shoots.
The first five leaf CTLV relative expression quantity of Fig. 2 .2:40 DEG C 4h/25 DEG C 4h process 30d and 50d sweet orange plant tender shoots, get 9 and 5, the first five leaf of each tender shoots each position sample during this temperature match curing conditions process 30d and 50d respectively, 30d and 50d gets 7, Lao Ye sample near tender shoots and 4 respectively.
Fig. 2 .3:40 DEG C 4h/30 DEG C 4h process 30d and the first five leaf of 40d tender shoots CTLV relative expression quantity.
The broken mosaic virus relative expression quantity of Fig. 2 .4:40 DEG C 10h/30 DEG C 14h, 40 DEG C 4h/25 DEG C 4h and 40 DEG C 4h/30 DEG C the first five leaf of 4h process sweet orange plant tender shoots compares, wherein, 40 DEG C of 10h/25 DEG C of 14h intermittent warming 30 days, 40 DEG C of 4h/25 DEG C of 4h intermittent warming 30 days and 50 days, 40 DEG C of 4h/30 DEG C of 4h temperature match curing conditions process 30 days and 40 days time get 4, the first five leaf of each tender shoots each position sample respectively, 9 and 5,10 and 5, get 4, the Lao Ye sample near tender shoots respectively, 7 and 4,3 and 3.Under these 3 kinds of Temperature Treatment, the sample number that first leaf is 0 to the 5th leaf CTLV content is 33 respectively, 28,25,23,19,40 DEG C of 4h/25 DEG C of 4h temperature condition process 50 days, when 40 DEG C of 4h/30 DEG C of 4h temperature condition process 30 days and 40 DEG C of 4h/25 DEG C of 4h temperature condition process 40 days, respectively there is 1 Lao Ye near tender shoots CTLV not detected.
Fig. 2 .5: stem sharp under temperature match curing conditions process.
Fig. 2 .6: temperature match curing conditions process stem sharp virus elimination rate detects.
Embodiment
Main innovative point of the present invention is: under infection citrus tatter leaf virus plant is positioned over pulse temperature treatment conditions, utilize real time quantitative PCR method to monitor viral level in susceptible Citrus Plants.Pulse temperature process refers to as required Temperature Treatment programming be become different temperatures height and the change of different time length, the program process of similar pulse change.In pulse temperature handling procedure, high temperature is the effect with the broken mosaic virus breeding of passivation, low temperature is the temperature of applicable Citrus Plants normal growth, real time quantitative PCR method monitoring is adopted to obtain suitable pulse temperature treatment conditions, to reach such effect: the megathermal period makes that the broken mosaic virus in Citrus Plants is passivated and fertility is very low, be maintained to when not yet causing Citrus Plants heat injury and proceed to hypothermic phase, hypothermic phase Citrus Plants can sprout tender shoots by normal growth, be maintained to when broken mosaic virus propagating active not yet recovers and proceed to the megathermal period again, alternate cycles process like this, make broken mosaic virus in Citrus Plants body, be in very low content level and almost do not have fertility state always, and Citrus Plants can normal growth rudiment, significantly improve sprouted tender tip stem apex quantity and nontoxic rate, Shoot-tip Grafting method is utilized to obtain nontoxic stem sharp further.
Before further describing the specific embodiment of the invention, should be understood that protection scope of the present invention is not limited to following specific specific embodiments; It is also understood that the term used in the embodiment of the present invention is to describe specific specific embodiments, instead of in order to limit the scope of the invention.The test method of unreceipted actual conditions in the following example, usually conveniently condition, or according to the condition that each manufacturer advises.
When embodiment provides number range, should be understood that except non-invention is otherwise noted, between two end points of each number range and two end points, any one numerical value all can be selected.Unless otherwise defined, all technology used in the present invention are identical with the meaning that those skilled in the art of the present technique understand usually with scientific terminology.Except the concrete grammar used in embodiment, equipment, material, according to those skilled in the art to the grasp of prior art and record of the present invention, any method of prior art that is similar with the method described in the embodiment of the present invention, equipment, material or that be equal to, equipment and material can also be used to realize the present invention.
Unless otherwise indicated, disclosed in the present invention experimental technique, detection method, preparation method all adopt the routine techniques of the molecular biology of the art routine, biochemistry, chromatin Structure and analysis, analytical chemistry, cell chulture, recombinant DNA technology and association area.These technology are existing in existing document improves explanation, specifically can see the MOLECULARCLONING:ALABORATORYMANUAL such as Sambrook, Secondedition, ColdSpringHarborLaboratoryPress, 1989andThirdedition, 2001; Ausubel etc., CURRENTPROTOCOLSINMOLECULARBIOLOGY, JohnWiley & Sons, NewYork, 1987andperiodicupdates; TheseriesMETHODSINENZYMOLOGY, AcademicPress, SanDiego; Wolffe, CHROMATINSTRUCTUREANDFUNCTION, Thirdedition, AcademicPress, SanDiego, 1998; METHODSINENZYMOLOGY, Vol.304, Chromatin (P.M.WassarmanandA.P.Wolffe, eds.), AcademicPress, SanDiego, 1999; And METHODSINMOLECULARBIOLOGY, Vol.119, ChromatinProtocols (P.B.Becker, ed.) HumanaPress, Totowa, 1999 etc.
Embodiment
1 materials and methods
1.1 material
The 10 years raw trifoliate orange anvil Fengjie 72-1 navel orange plant and the 10 years healthy raw trifoliate orange anvil Fengjie 72-1 navel orange plant that have infected CTLV provide by Citrus Research Institute Of Chinese Academy Of Agricultural Sciences.Prepare 10 strain examination materials in experiment altogether, select wherein to grow 3 more consistent strain examination materials for experiment, basin number is respectively 822,809,1076, and reference numeral is 3,5, No. 6 strains.
1.2 experimental technique
Temperature is set as 40 DEG C of 10h/30 DEG C of 14h (No. 3 strains), 40 DEG C of 4h/25 DEG C of 4h (No. 5 strains) and 40 DEG C of 4h/30 DEG C of 4h (No. 6 strains) respectively.When trying material and sprouting tender shoots to about 1-3cm, the position long according to blastogenesis is numbered, samples.Tender shoots top about 0.15-0.20mm is for Shoot-tip Grafting, and remaining tender shoots part to base portion select progressively 5 blade extracting CTLV, detects CTLV content for real-time fluorescence quantitative RT-PCR according to top.
Tender shoots method for numbering serial:
321 → No. 3 strain the 2nd branches the 1st bud
529 → No. 5 strain the 2nd branches the 9th bud
635 → No. 6 strain the 3rd branches the 5th bud
Other numbering by that analogy
Recall rate=(sample number/experiment gross sample number of CTLV tests positive) × 100%
Virus elimination rate=(CTLV detects the stem sharp number/survive stem sharp sum of being negative) × 100%
Using oranges and tangerines reference gene 18sRNA (GeneID:12486911) as reference, adopt real-time fluorescent quantitative RT-PCR method (concrete grammar reference: Journal of Fruit Science, use real-time fluorescence RT-PCR technology for detection oranges and tangerines broken mosaic virus Liu Ke grand, Zhou Changyong, Song Zhen, Zhou Yan, Li Zhongan, Tang Kezhi) mensuration of relative expression quantity is carried out to CTLV in intermittent warming sweet orange plant tender shoots tender leaf sample.The amplification curve performance simple spike of sample, water contrast does not have amplified peak, and interpret sample amplified reaction is normal, can be used for the expression analysis (Fig. 1) to CTLV.
2 results and analysis
2.1 typical heat treat condition (40 DEG C of 10h/30 DEG C of 14h) process the CTLV distribution situation of 30d sweet orange tender shoots tender leaf
Plant germination rate after 40 DEG C of 10h/30 DEG C of 14h process is low, obtains tender shoots 4, it shifted out from this condition during 50d during 30d.First and second sheet leaf of tender shoots CTLV recall rate is zero, third and fourth, five leaf CTLV recall rates 25%, tender shoots CTLV content to present from top to base portion trend from low to high generally, shows as the phenomenon that CTLV content obviously raises from tender shoots to aging blade.
In addition, from single bud CTLV containing taking temperature, No. 301, No. 342 the first five leaf of bud are not all with poison, the 3rd leaf band poison of No. 321 buds, 4th and the 5th leaf band poison of No. 341 buds, the tender shoots stem apex sprouted under this intermittent warming condition is described has very large probability and is removed CTLV (Fig. 2 .1).
The CTLV distribution situation of 2.240 DEG C of 4h/25 DEG C of 4h process 30d and 50d sweet orange plant tender shoots
40 DEG C 4h/25 DEG C 4h process sweet orange plant 30d, sweet orange rudiment is many, and tender shoots is healthy and strong, during 50d, starts to grow blade, no longer extend when most tender shoots poor growth and length are less than 2cm.During 30d, in 9 buds, CTLV recall rate 20%, 1 bud CTLV recall rate 40%, 4 bud CTLV recall rates 80% in 3 bud-leaf sheet CTLV recall rates, 0%, 1 bud-leaf sheet; During 50d, in 5 buds, CTLV recall rate 60% in CTLV recall rate 20%, 1 bud-leaf sheet in 1 bud CTLV verification and measurement ratio, 40%, 3 bud-leaf sheets, illustrates that the processing time is slightly long stronger to the inhibitory action of CTLV.Tender shoots CTLV content also to present from top to base portion trend from low to high generally.Tender shoots side aging leaf CTLV recall rate under this condition process is 100%, illustrates that this temperature match curing conditions can not remove the CTLV in Lao Ye.
From single bud CTLV contents level, No. 544, No. 546, No. 529, No. 536 buds of 30d process and No. 5310 the first five leaf of bud CTLV content rules of 50d process and other buds different, No. 544, No. 546, No. 529 buds the 3rd leaf CTLV content other leaves a little higher than, No. 536 bud the 4th leaf CTLV content other leaves a little higher than, No. 5310 buds the 5th leaf CTLV content is slightly lower than other leaves, and this illustrates that each position of tender leaf CTLV content also exists individual difference.
In addition, from position, tender leaf place, during process 30d, first leaf CTLV recall rate is 0%, second leaf CTLV recall rate 44.44%, third and fourth, the CTLV recall rate of five leaves is 55.56%; Process 50d time, first leaf CTLV recall rate 0%, second and third, four leaf CTLV recall rates the 20%, five leaf CTLV recall rate 60%, the CTLV content of first, second and third sheet leaf of all buds is all lower than rear two leaves.Infer from the first five leaf band poison situation of tender shoots, in Shoot-tip Grafting process, seem also to cut the stem apex (Fig. 2 .2) of leaf primordium more than a conventional 2-3 leaf primordium.
The CTLV distribution situation of 2.340 DEG C of 4h/30 DEG C of 4h process 30d and 40d sweet orange tender shoots tender leaf
40 DEG C 4h/30 DEG C 4h process sweet orange plant 30d, germination rate is high, and tender shoots sprouts more than 10.During process 40d, the rudiment of sweet orange plant is slow, tender shoots contraction in length.During process 30d, in 10 buds, 6 bud different leaves CTLV recall rates are that in 0%, 2 bud-leaf sheets, CTLV recall rate is that in 20%, 1 bud-leaf sheet, CTLV recall rate is that in 40%, 1 bud-leaf sheet, CTLV recall rate is 60%; During process 50d, 3 bud-leaf sheet CTLV recall rates, 0%, 2 CTLV blade recall rates 20% in 5 buds, ditto the same, in tender shoots, CTLV content to present from top to base portion trend from low to high generally.In the aging blade of tender shoots side, there are 2 Lao Ye CTLV not detected, other aging leaf CTLV recall rate 100%, illustrate that this temperature match curing conditions also can not suppress the CTLV in Lao Ye completely, but compared with 40 DEG C of 4h/25 DEG C of 4h process, seem that there is the effect better suppressing CTLV.
From single bud, No. 635 buds of process 30d are different with 6310 buds and other buds processing 40d, a little higher than 5th leaf of 4th leaf CTLV content of No. 635 buds, 6310 buds the 4th leaf CTLV content other leaves a little higher than, result shows the difference of the individual CTLV content of tender shoots equally.
From the bud position at tender leaf place, during process 30d, first and second sheet leaf CTLV recall rate is the zero, three leaf CTLV recall rate the 10%, four leaf CTLV recall rate the 20%, five leaf CTLV recall rate 40%; During process 50d, first, second and third sheet leaf CTLV recall rate is the zero, the 45 leaf CTLV recall rate 20%, the CTLV content of first and second sheet leaf of all buds lower than third and fourth, five leaves.This condition seems the same a kind of temperature match curing conditions supposition unanimously, can attempt the stem apex (Fig. 2 .3) cutting leaf primordium more than a conventional 2-3 leaf primordium in Shoot-tip Grafting process.
The comparison that 2.4 3 kinds of intermittent warming affect sweet orange tender shoots CTLV content distribution
Analysis is compared to sweet orange tender shoots CTLV content distribution under 3 kinds of temperature match curing conditions, result following (Fig. 2 .4)
(1) comparison at same tender shoots position
Under three kinds of temperature match curing conditions process, tender shoots first leaf is not all detected CTLV.For tender shoots second leaf, during process 30d, the CTLV recall rate of 40 DEG C 10h/30 DEG C 14h and 40 DEG C 4h/30 DEG C 4h condition is zero, and the CTLV recall rate of 40 DEG C of 4h/25 DEG C of 4h treatment conditions is 44.44%.For tender shoots third and fourth, five leaves, the CTLV content of 40 DEG C of 4h/25 DEG C of 4h conditions is higher than 40 DEG C of 10h/30 DEG C of 14h conditions, and 40 DEG C of 10h/30 DEG C of 14h conditions are again higher than 40 DEG C of 4h/30 DEG C of 4h process.These results display often kind of temperature match curing conditions seems to produce inhibitory action to the breeding of CTLV on stem apex tender shoots position, all has the potentiality necessarily removing CTLV.Meanwhile, in three kinds of temperature match curing conditions, 40 DEG C 4h/30 DEG C 4h process having the greatest impact to sweet orange tender shoots CTLV content, adopts this condition, likely obtains higher virus elimination rate during detoxification.
40 DEG C 4h/25 DEG C 4h process sweet orange plant 50d, indivedual tender shoots is detected higher toxic amount, and all the other buds CTLV content is roughly the same with process 30d bud situation, but during 50d, the growth of tender shoots is not as 30d.40 DEG C 4h/30 DEG C 4h process sweet orange plant 40d, the first five leaf toxic amount of the tender shoots of acquisition is all extremely low, particularly third and fourth, the content of five leaves is lower than 40 DEG C of 4h/25 DEG C of 4h process.
(2) compare between the tender and aging position of children
The content part of the tender shoots tender leaf processed under 3 temperature match curing conditions is higher than Lao Ye CTLV content, part is lower than Lao Ye CTLV content, result display intermittent warming is huge on the impact of CTLV in tender shoots, to the tender shoots different parts CTLV inhibitory action also property of there are differences, the CTLV changes of contents amplitude in Lao Ye that simultaneously seems is relatively little.
2.5 intermittent warming sweet orange plant survival stem sharp band poison situations
Stem sharp growing state under temperature match curing conditions process, as shown in Fig. 2 .5.
Under typical heat treat condition, germination rate is low, and survival rate is high but all do not remove CTLV, and possible sweet orange plant needs the process longer time just can reach detoxification efficiency.Under 40 DEG C of 4h/25 DEG C of 4h treatment conditions, stem sharp 30d and 50d survival rate are suitable, during 30d virus elimination rate 14.29%, 50d time virus elimination rate 60%.Under 40 DEG C of 4h/30 DEG C of 4h treatment conditions, the stem sharp survival rate of 30d and 40d is suitable, does not have stem sharp by detoxification, stem sharp virus elimination rate 71.43% during 40d during 30d.Comprehensive 3 kinds of temperature match curing conditions, sharing 2 strain examination materials in normative heat treatment, is 2 times of other 2 kinds of intermittent warming, and because the time of each high temperature treatment is longer, institute tender shoots of obtaining is few, affects Shoot-tip Grafting time and success rate.Under 40 DEG C 4h/25 DEG C 4h and 40 DEG C 4h/30 DEG C of 4h condition process, in the sweet orange plant short time, germination rate is high, and institute's shoot-tip grafting plantlet that obtains is many, and this every 4h frequency conversion method is once more suitable for carrying out detoxification (Fig. 2 .6, table 2.1) to sweet orange plant.
Table 2.1 intermittent warming sweet orange shoot-tip grafting plantlet is added up
3 discuss
Under 3 kinds of temperature match curing conditions process, the stem sharp survival rate of normative heat treatment is high, but germination rate and virus elimination rate are all lower, and possible sweet orange plant grows under this condition and sustains damage.Under 40 DEG C 4h/25 DEG C 4h and 40 DEG C 4h/30 DEG C 4h treatment conditions, plant strain growth is good, and germination rate is high, and stem sharp virus elimination rate is also high.This experiment expends from processing time, examination material, germination rate and virus elimination rate Integrated comparative, and 40 DEG C 4h/25 DEG C 4h and 40 DEG C 4h/30 DEG C 4h treatment conditions are all better, the stem sharp survival rate of acquisition and virus elimination rate high.The nontoxic nursery stock that comprehensive macro-economic rersults need is basic consideration, during summer, temperature is higher, ensure that plant environment 25 DEG C is more difficult, can 40 DEG C of 4h/30 DEG C of 4h during summer, temperature is relatively higher, aestival aspect to good control some, save the energy simultaneously, can adopt 40 DEG C of 4h/25 DEG C of 4h during winter, winter temperature is lower, environmental temperature control 25 DEG C easier.3 temperature match curing conditions process sweet orange 30d detoxification efficiencies are not best, and start to take a turn for the better more than effect after 30d.In addition, the present invention is 40 DEG C of 4h/27 DEG C of 4h, 40 DEG C of 2h/30 DEG C of 6h, 40 DEG C of these several combination of variable temperature poison-removing methods of 6h/25 DEG C of 2h further by verification experimental verification also, result achieves the effect similar with 40 DEG C 4h/25 DEG C 4h and 40 DEG C 4h/30 DEG C 4h, is all better than the contrast method of normative heat treatment.
In R&D process of the present invention, inventor also attempted: the Citrus Plants not doing high temperature treatment, gets tender shoots stem apex detoxify be difficult to successfully from plant.Do the Citrus Plants of high temperature treatment, if temperature condition is 14 hours 40 degree of 10 hours+nights 30 degree of conventional daytime, it is little that plant sprouts tender shoots, and evil of being easily heated is withered.The Citrus Plants of strobe pulse Temperature Treatment, if temperature condition is the repeated transformations of 4 hours of 35 degree of 4 hours+25 degree, it is many that plant sprouts tender shoots, but very low by tender shoots stem apex detoxify rate.
The present invention finds that real-time quantitative RT-PCR is difficult to detect first and second sheet leaf CTLV content in the first five leaf of tender shoots, consider in conjunction with Shoot-tip Grafting, when applying temperature match curing conditions process sweet orange plant afterwards, suggestion cuts the leaf primordium slightly larger than a conventional 2-3 leaf primordium and carries out Shoot-tip Grafting detoxification, and production practices meaning is larger.
The present invention additionally uses other citrus varieties, all can realize technique effect of the present invention.That is the method for the broken mosaic virus of efficient removal oranges and tangerines of the present invention is not limited in Fengjie 72-1 navel orange plant, but is applicable to all citrus varieties.
The above; be only preferred embodiment of the present invention; not to any formal and substantial restriction of the present invention; should be understood that; for those skilled in the art; under the prerequisite not departing from the inventive method, also can make some improvement and supplement, these improve and supplement and also should be considered as protection scope of the present invention.All those skilled in the art, without departing from the spirit and scope of the present invention, a little change made when utilizing disclosed above technology contents, the equivalent variations of modifying and developing, be Equivalent embodiments of the present invention; Meanwhile, all according to substantial technological of the present invention to the change of any equivalent variations that above-described embodiment is done, modify and differentiation, all still belong in the scope of technical scheme of the present invention.
Claims (10)
1. a method for the broken mosaic virus of efficient removal oranges and tangerines, shown method comprises the steps:
(1) pulse temperature is adopted to heat-treat the oranges and tangerines plant having infected the broken mosaic virus of oranges and tangerines;
(2) stem apex of the oranges and tangerines plant handled well in step (1) is adopted to carry out Shoot-tip Grafting.
2. method according to claim 1, is characterized in that, in step (1), described oranges and tangerines plant is Fengjie sweet orange plant.
3. method according to claim 1, is characterized in that, in step (1), described oranges and tangerines plant is Fengjie 72-1 navel orange plant.
4. method according to claim 1, it is characterized in that, in step (1), described employing pulse temperature is heat-treated, can specifically: the oranges and tangerines plant having infected the broken mosaic virus of oranges and tangerines is first processed a period of time T1 under 40 DEG C of conditions, and the scope of described T1 is 2 ~ 6h; Then continue to process a period of time T2 under 25 ~ 30 DEG C of conditions, the scope of described T2 is 2 ~ 6h; Alternate cycles process like this 30 ~ 50 days.
5. method according to claim 1, it is characterized in that, in step (1), described employing pulse temperature is heat-treated, can specifically: the oranges and tangerines plant having infected the broken mosaic virus of oranges and tangerines is first processed a period of time T1 under 40 DEG C of conditions, and the scope of described T1 is 2 ~ 6h; Then continue to process a period of time T2 under 25 ~ 30 DEG C of conditions, the scope of described T2 is 2 ~ 6h; Alternate cycles process like this 40 ~ 50 days.
6. the method according to claim 4 or 5, is characterized in that, in step (1), the scope of described T1 is 4h; The scope of described T2 is 4h.
7. the method according to claim 4 or 5, is characterized in that, in step (1), then continues to process a period of time T2 under 27 DEG C of conditions.
8. method according to claim 1, it is characterized in that, in step (1), described employing pulse temperature is heat-treated, can specifically: the oranges and tangerines plant having infected the broken mosaic virus of oranges and tangerines is first processed a period of time T1 under 40 DEG C of conditions, and the scope of described T1 is 4h; Then continue to process a period of time T2 under 25 DEG C of conditions, the scope of described T2 is 4h; Alternate cycles process like this 50 days.
9. method according to claim 1, it is characterized in that, in step (1), described employing pulse temperature is heat-treated, can specifically: the oranges and tangerines plant having infected the broken mosaic virus of oranges and tangerines is first processed a period of time T1 under 40 DEG C of conditions, and the scope of described T1 is 4h; Then continue to process a period of time T2 under 30 DEG C of conditions, the scope of described T2 is 4h; Alternate cycles process like this 40 days.
10. method according to claim 1, is characterized in that, in step (2), the stem apex adopted comprises the leaf primordium of more than 2 ~ 3.
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN107509540A (en) * | 2017-09-28 | 2017-12-26 | 钟山县龙岛种养专业合作社 | A kind of method that citrus prevents and treats broken leaf disease |
| CN114391410A (en) * | 2022-01-19 | 2022-04-26 | 象山县农业经济特产技术推广中心 | Method for rapidly removing virus diseases of multi-embryo citrus varieties |
Citations (8)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| DD114207A1 (en) * | 1974-10-04 | 1975-07-20 | ||
| SU1658929A1 (en) * | 1988-12-30 | 1991-06-30 | Научно-исследовательский зональный институт садоводства Нечерноземной полосы | Method to obtain healthy strawberry planting stock |
| CN1331907A (en) * | 2001-06-28 | 2002-01-23 | 廉江市农业局 | Disease-free red orange seedling breeding method |
| CN1903016A (en) * | 2005-07-29 | 2007-01-31 | 潘利军 | Virus removing method for lily tissue culturing seedling by heat treatment |
| CN101061790A (en) * | 2007-04-13 | 2007-10-31 | 浙江省农业科学院 | Quick virus-free group-cultivating and propagating method of OT type lily of hybrid group |
| CN103404397A (en) * | 2013-07-26 | 2013-11-27 | 华南农业大学 | Method for preventing and treating candidatus liberibacter asiaticus by heat treatment |
| CN103891531A (en) * | 2013-12-12 | 2014-07-02 | 广西大学 | Method for cultivating citrus germplasm infected with citrus vein phloem degeneration |
| CN104813935A (en) * | 2015-04-20 | 2015-08-05 | 云南省农业科学院经济作物研究所 | Method for removing PVS viruses of potato test-tube plantlets |
-
2015
- 2015-12-04 CN CN201510896416.0A patent/CN105493972A/en active Pending
Patent Citations (8)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| DD114207A1 (en) * | 1974-10-04 | 1975-07-20 | ||
| SU1658929A1 (en) * | 1988-12-30 | 1991-06-30 | Научно-исследовательский зональный институт садоводства Нечерноземной полосы | Method to obtain healthy strawberry planting stock |
| CN1331907A (en) * | 2001-06-28 | 2002-01-23 | 廉江市农业局 | Disease-free red orange seedling breeding method |
| CN1903016A (en) * | 2005-07-29 | 2007-01-31 | 潘利军 | Virus removing method for lily tissue culturing seedling by heat treatment |
| CN101061790A (en) * | 2007-04-13 | 2007-10-31 | 浙江省农业科学院 | Quick virus-free group-cultivating and propagating method of OT type lily of hybrid group |
| CN103404397A (en) * | 2013-07-26 | 2013-11-27 | 华南农业大学 | Method for preventing and treating candidatus liberibacter asiaticus by heat treatment |
| CN103891531A (en) * | 2013-12-12 | 2014-07-02 | 广西大学 | Method for cultivating citrus germplasm infected with citrus vein phloem degeneration |
| CN104813935A (en) * | 2015-04-20 | 2015-08-05 | 云南省农业科学院经济作物研究所 | Method for removing PVS viruses of potato test-tube plantlets |
Non-Patent Citations (1)
| Title |
|---|
| 贾敏: "变温处理对叶片中柑橘碎叶病毒含量影响和脱毒效应", 《万方数据库》 * |
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN107509540A (en) * | 2017-09-28 | 2017-12-26 | 钟山县龙岛种养专业合作社 | A kind of method that citrus prevents and treats broken leaf disease |
| CN114391410A (en) * | 2022-01-19 | 2022-04-26 | 象山县农业经济特产技术推广中心 | Method for rapidly removing virus diseases of multi-embryo citrus varieties |
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