CN102326542B - New use of palmitic acid as meloidogyne incognita repellent - Google Patents
New use of palmitic acid as meloidogyne incognita repellent Download PDFInfo
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- CN102326542B CN102326542B CN 201110325372 CN201110325372A CN102326542B CN 102326542 B CN102326542 B CN 102326542B CN 201110325372 CN201110325372 CN 201110325372 CN 201110325372 A CN201110325372 A CN 201110325372A CN 102326542 B CN102326542 B CN 102326542B
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Abstract
The invention discloses new use of palmitic acid as a meloidogyne incognita repellent. According to the new use, the palmitic acid can be used for preparing the meloidogyne incognita repellent and is used as an active ingredient or an inactive ingredient; and when the palmitic acid is used as the active ingredient, the palmitic acid can be separately used, and can also be used together with other active ingredients. The repellent is a methanol solution of the palmitic acid; and the concentration of the palmitic acid in the repellent is 0.5 to 4.00mM. The palmitic acid or the methanol solution of the palmitic acid used for preventing and controlling the meloidogyne incognita is safe, nontoxic and effective; and when the concentration of a palmitic acid solution is 0.5 to 4.00mM, the meloidogyne incognita can be repelled, and the tendency index can reach -0.20.
Description
Technical field
The present invention relates to a kind of new purposes of palmitic acid, particularly palmitic acid is as the purposes in the Meloidogyne incognita repellant.
Background technology
Root-knot nematode is one of worldwide important causal organism, and production has brought serious loss to world agriculture for it, and the agricultural economy loss that therefore the annual whole world cause is up to 157,000,000,000.Kind surplus the root-knot nematode of having reported in the world at present has 90, and Meloidogyne incognita (Meloidogyne incognita), javanese root knot nematode (M.javanica), peanut root-knot nematode (M.arenaria) and northern root-knot nematode (M.hapla) account for 95% of whole root-knot nematode.A series of problems that Meloidogyne incognita can cause facility to produce, and make a large amount of economic crops underproduction.In northern China, based on Meloidogyne incognita, the host plant of root-knot nematode is in extensive range, surpass 3000 kinds of plants, comprise monocotyledon, dicotyledon, herbaceous plant and woody plant, remunerative crops such as the Solanaceae in the vegetables, Curcurbitaceae particularly, the general underproduction 30%～40% in back but tomato wherein and cucumber are injured, even total crop failure.Current, the control of root-knot nematode is mainly based on the chemicals control, and it is cancelled by legislative branch just gradually to environment, person poultry safety's the serious harm that exists.Therefore, safely, effectively, the medicine of low toxicity and the method for preventing and treating are extremely urgent.
Root-knot nematode mainly comprises six breeding times, and in its two length of times, can utilize the flourishing cerebral organ that innervates, (the AWA of olfactory nerves unit by head, AWC) semiochemicals that comes the perception root system to give out, carry out the taxis migration to host plant, infect plant, thereby finish its life cycle.Therefore, in two length of times of root-knot nematode, can play the purpose of nematoda control by disturbing the taxis migration of root-knot nematode.Along with finishing of the beautiful nematode of model animal (Caenorhabditis elegans) gene order-checking work, at present separated and identified the gene of 19800 6627 protein families of can encoding, and the function of related gene has been carried out detailed and deep research.And, we find in previous work, E-value≤1 o'clock, beautiful nematode and Meloidogyne incognita homologous gene are near 6000, wherein the mpk-1 gene is one of major function gene in the Ras approach, relevant with the taxis migration of nematode, and mpk-1 gene autoploidy in beautiful nematode and Meloidogyne incognita is higher, so the taxis migration of Meloidogyne incognita can be supported and reflect to mpk-1 expression of gene situation.
Palmitic acid is as a kind of natural acid, and is nontoxic, at aspects such as food additives, perfume industries many application arranged all, has not yet to see the report that is useful on the control Meloidogyne incognita.
Summary of the invention
The new purposes that the purpose of this invention is to provide a kind of palmitic acid.This new purposes is that palmitic acid is for the preparation of the Meloidogyne incognita repellant; In this repellant, palmitic acid can also can carry out compatibility jointly as active component with other composition with control Meloidogyne incognita activity separately as active component.
Described repellant is the methanol solution of palmitic acid.
The concentration of palmitic acid described in the described repellant in described repellant is 0.5mM-4.00mM.
The present invention also provides a kind of preparation method of described repellant, comprises the steps: palmitic acid is dissolved in the methanol solution that obtains palmitic acid behind the methyl alcohol.
The present invention also provides a kind of method of preventing and treating Meloidogyne incognita, comprises the steps: described repellant is used for the control Meloidogyne incognita; Described control Meloidogyne incognita is the repellent Meloidogyne incognita.
Use the methanol solution control Meloidogyne incognita of palmitic acid or palmitic acid, safe, nontoxic, effective, but when the concentration of palmitic acid solution is 0.50mM-4.00mM the repellent Meloidogyne incognita, the taxis index can reach-0.20.
Description of drawings
Fig. 1 is the schematic diagram that the methanol solution of variable concentrations palmitic acid detects the Meloidogyne incognita taxis.Wherein, A represents to add the circle of methyl alcohol, and B represents to add the circle of palmitic acid methanol solution, and C represents to place the nematode position.
Fig. 2 is that the methanol solution of variable concentrations palmitic acid is to the influence of Meloidogyne incognita taxis.
Fig. 3 is the influence that the methanol solution of variable concentrations palmitic acid is expressed Meloidogyne incognita Mi-mpk-1.
Embodiment
Employed experimental technique is conventional method if no special instructions among the following embodiment.
Used material, reagent etc. if no special instructions, all can obtain from commercial channels among the following embodiment.
The palmitic acid of embodiment 1, variable concentrations is to the influence of Meloidogyne incognita
1, material: with palmitic acid (CAS:57-07-3, Sigma) be made into the solution that concentration is 0.50mM, 1.00mM, 2.00mM and 4.00mM with chromatographically pure methyl alcohol, obtain the methanol solution (repellant 1) of 0.50mM palmitic acid, the methanol solution (repellant 2) of 1.00mM palmitic acid, the methanol solution (repellant 3) of 2.00mM palmitic acid and the methanol solution (repellant 4) of 4.00mM palmitic acid respectively.Preserve standby in-20 ℃ of refrigerators.
Meloidogyne incognita (Meloidogyne incognita) (force is raised, Zheng Jingwu, Shang Hanwu. root-knot nematode classification and evaluation approach and progress. Zhejiang agricultural journal .2005,17:106-110, the public can obtain from China Agricultural University) second instar larvae.
2, method: use repellant 1-4 and chromatographically pure methyl alcohol (contrast) to handle the second instar larvae of Meloidogyne incognita (Meloidogyne incognita) respectively, utilize the result of taxis index, Mi-mpk-1 gene expression, illustrate that from physiological level and molecular level palmitic acid is as the effect of Meloidogyne incognita repellant.
Under the laboratory condition, according to Ward (Ward Samuel.Chemotaxis by the nematode Caenorhabditis elegans:Identification of attractants and analysis of the response by use of mutants.Proc.Nat.Acad.Sci.USA.1973,70,817-821) and (Bargmann C.I. such as Bargmann, Hartweig E., Horvitz H.R.Odorant selective genes and neurons mediate olfaction in C.elegans.Cell, 1993,74, the method that 515-527) provides, adopting diameter is the culture dish of 6cm, is the circle of 1cm at a diameter apart from 1cm place, culture dish edge label diameter respectively.The agar that adds 10ml 0.8% in each culture dish.In the culture dish of the two circles of mark, the chromatographically pure methyl alcohol that adds 10ml in the side circle is contrast, and opposite side adds the palmitic acid solution of 10ml variable concentrations for handling.In the circle of both sides, all add 10 μ L chromatographically pure methyl alcohol as negative control.Culture dish behind the adding compound, put into 25 ℃ of incubator lucifuge hatchings 2 hours earlier, take out the back in the middle part of each culture dish, the position equidistant with the circle of both sides (Fig. 1) adds the second instar larvae of 50 Meloidogyne incognitas that just hatched, 25 ℃ of incubator lucifuges were hatched 3 hours again, according to Bargmann and Horvitz (Bargmann C.I., Horvitz H.R.Chemosensory neurons with overlapping functions direct chemotaxis to multiple chemicals in C.elegans.Neuron, 1991,7, method statistic taxis index (Chemotaxis Index) 729-742), its computational methods are: the quantity of adding nematode in the palmitic acid solution circle with add the poor of nematode population in the methanol solvate circle, with the ratio that adds line insect number, i.e. taxis index=(nematode population in nematode population in the palmitic acid solution circle-methyl alcohol circle)/the add total amount of nematode); Taxis index 〉=0.2 o'clock is to attract; 0≤taxis index＜0.2 o'clock is faint attraction, and taxis index≤-0.02 o'clock is to repel.
The palmitic acid solution-treated of each concentration is made 3 wares and is repeated, and adds up the quantity of adding nematode in palmitic acid methanol solution (B) circle in every ware and the nematode population of adding in methanol solvate (A) circle respectively, and the mean value that calculates the taxis index and respectively repeat the results are shown in Table 1.
The methanol solution of the palmitic acid of table 1. variable concentrations is handled the statistics of Meloidogyne incognita
According to the method for Ward and Bargmann etc., collect the second instar larvae of 2000 different disposal simultaneously, be used for the analysis of Mi-mpk-1 gene expression.The primer that gene information and Real-time PCR adopt, as shown in table 2.
Actin and mpk-1 gene and primer information among the table 2.Real-time PCR
RNease Kit (Tiangen) is adopted in the extraction of the total RNA of nematode, and total RNA adopts RNase-free DNase (Takara) to carry out purifying, utilizes M-MLV (Promega) to reverse, and obtains article one cDNA chain.Be template (2 μ g) with the cDNA chain, carry out Real-time PCR, fluorescent dye adopts 2 * SYBR Green mix (Toyobo), and the upstream primer of 10 μ M and downstream primer be 1 μ L respectively, complements to 25 μ l with high purity water.The PCR reaction condition is two step method:
The first step: 50 ℃ of 2min, 95 ℃ of 1min
Second step: 95 ℃ of 15s, 52 ℃ of 15s, 72 ℃ of 45s, 40 circulations, and at 72 ℃ of collection fluorescence.
3, result
(1) the palmitic acid methanol solution of variable concentrations is to the influence of Meloidogyne incognita taxis
Between scope 0.5mM～4.00mM that test detects, palmitic acid all shows as the effect of repellent to Meloidogyne incognita, and repellent effect strengthens along with the increase of palmitic acid concentration, and taxis index variation scope is-0.20～-0.067, all shows as the effect (Fig. 2) of repulsion.And in contrast (0mM), the taxis index is 0, between-0.02～0, gets rid of solvent methanol to the influence of Meloidogyne incognita.
(2) the palmitic acid methanol solution of variable concentrations is to the influence of Meloidogyne incognita Mi-mpk-1 expression
Palmitic acid shows as inhibitory action (Fig. 3) to the influence that Meloidogyne incognita Mi-mpk-1 expresses, and inhibitory action increases with the increase of palmitic acid concentration (0.50mM-4.00mM), and maximum suppression effect reaches 77%.Shown that by molecular data the palmitic acid of 0.50mM-4.00mM repels Meloidogyne incognita.
4, conclusion
Physiological data and molecular data show: but the methanol solution repellent Meloidogyne incognita of 0.50mM～4.00mM palmitic acid, thus play the effect of preventing and treating Meloidogyne incognita, and control efficiency is along with the increase that increases to of palmitic acid concentration.So palmitic acid can be used as the repellant of Meloidogyne incognita, the using dosage scope is: 0.50mM～4.00mM.
Claims (2)
1. the application of palmitic acid in preparation Meloidogyne incognita repellant, it is characterized in that: described Meloidogyne incognita repellant is the methanol solution of palmitic acid; The concentration of described palmitic acid in described repellant is 0.5 mM-4.00mM.
2. a method of preventing and treating Meloidogyne incognita comprises the steps: the methanol solution of palmitic acid is used for the control Meloidogyne incognita; Described control Meloidogyne incognita is the repellent Meloidogyne incognita; Described palmitic acid concentration is 0.5 mM-4.00mM.
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| US9526242B2 (en) * | 2013-07-18 | 2016-12-27 | Fmc Corporation | Nematode repellent composition |
| CN107216169A (en) * | 2016-03-21 | 2017-09-29 | 李俊峰 | Root-knot nematode suppresses film and its production technology |
| CN108157368A (en) * | 2017-12-30 | 2018-06-15 | 中国农业大学 | Application of the methyl hexadecanoate in Meloidogyne incognita is prevented |
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Inventor after: Zuo Yuanmei Inventor after: Dong Linlin Inventor after: Li Xiaolin Inventor after: Chen Qing Inventor before: Zuo Yuanmei Inventor before: Dong Linlin Inventor before: Li Xiaolin |
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