CN103875607B - The authentication method of a kind of soybean aphid biological strain - Google Patents
The authentication method of a kind of soybean aphid biological strain Download PDFInfo
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- CN103875607B CN103875607B CN201410095024.XA CN201410095024A CN103875607B CN 103875607 B CN103875607 B CN 103875607B CN 201410095024 A CN201410095024 A CN 201410095024A CN 103875607 B CN103875607 B CN 103875607B
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Abstract
The invention provides the authentication method of a kind of soybean aphid biological strain, adopt aphid breeding quantity and the plant diseases of differential host under greenhouse experiment to show the method combining, the soybean aphid biological strain that can Rapid identification goes out separate sources, carry out the research such as Soybean Resistance aphid Screening of Germplasm and breeding of new variety targetedly, for the integrated control of soybean aphid and the establishment of breeding for pest resistance target provide foundation, be a kind of fast, the authentication method of strong operability, have a good application prospect.
Description
Technical field
The invention belongs to plant pest authenticate technology field, be specifically related to qualification and the aphid of soybean aphid biological strainWorm Resistance Identification, particularly relates to a kind of method to soybean aphid Race Identification.
Background technology
Soybean aphid [AphisglycinesMatsumura (Hemiptera:Aphididae)] is the main of soybean seedlingOne of insect, belongs to Semiptera Aphidiadae. Before 2000 the harm of soybean aphid mainly occur in China, Philippine,The countries such as Thailand, Korea, Korea S, Indonesia and Russia, successively invade the U.S., Canada and Australian afterwards.Along with the aggravation of climate warming trend, soybean aphid presents the trend growing in intensity, and has become the world of extensive concernProperty agricultural pests. Soybean aphid is a kind of oligophagy, different host insect. The identified summer host of China have soybean,The minority bean plants such as black soya bean (Glycinesp.) and wild soybean (GlycinesojaSieb.etZucc); Winter hostOnly has sandlwood (RhamnusdavuricaPallus) one. At America & Canada, its host types is to domestic similar.The harm of soybean aphid is mainly manifested in and causes plant atrophy, depauperation, leaf rolling, loment rate to reduce, produceThe lower degradation of amount aspect, the honeydew of aphid secretion causes mould very easily to grow in addition, and can propagate multiple virus, as greatlyBean mosaic virus, dwarf virus etc., further endanger Soybean production, and the serious time of aphid damage can cause soybean to subtractProduce more than 50%. North America region because of soybean aphid endanger the economic loss causing be annual 36 to 4,800,000,000 dollars itBetween.
For the harm of soybean aphid, Chinese scholars has been carried out large quantity research, mainly concentrates on resistance resource at presentQualification, resistant gene genetic development is resolved, the meticulous location of resistant gene and the qualification of aphid bion. At presentThrough located 7 Dominant genes dominant aphid-resistant gene (Rag, Rag1, Rag2, Rag3, Rag3b, Rag5,Rag6). Foreign scholar according to North America separate sources aphid to containing Rag1, Rag2, Rag1/Rag2 resistant geneThe different infection abilities of soybean resource mark off 4 soybean aphid bions: bion 1 can not be infected and be containedThe soybean germplasm of Rag1; Bion 2 can be infected Rag1 but can not be infected Rag2; Bion 3 can be infectedRag2, but can not infect the germplasm of Rag1/Rag2,4 of bions can be infected Rag1, Rag2, Rag1/Rag2Soybean germplasm, but at present domestic also not about the research report of the qualification of soybean aphid biological strain.
The domestic screening of the resistance resource with regard to soybean aphid is done a lot of work, and has filtered out some resistant materials, butAlso do not cultivate at present the cultivated soybean kind with stable resistance. The qualification of soybean aphid bion is for various placesSelect targetedly resistant soybean kind and cultivate resistant soybean resource most important, but at present domesticly also there is no thisThe system research of aspect, one of its reason is that qualification workload is larger, two be to lack the clear and definite soybean of aphid-resistant geneResource. This laboratory, through long-term and arduous work, filters out the soybean material that partial resistance is stronger, also receives simultaneouslyCollection has arrived the clear and definite resistance resource of domestic and international partial resistance gene. On this basis, carry out the life of domestic soybean aphidManage the qualification work of microspecies, specify the pest-resistant scope of aphid-resistant gene (Rag), carry for cultivating anti-aphid new soybean varietiesFor foundation.
Summary of the invention
The present invention proposes the authentication method of soybean aphid biological strain in a kind of greenhouse, and the present invention adopts differential host to existWhat in greenhouse, through artificial infection aphid, the Aphed population after 14 days and the aphid infection ability classification after 21 days combinedMethod, Rapid identification soybean aphid biological strain, is integrated control and the establishment soybean breeding for pest resistance order of soybean aphidMark provides foundation.
Biological strain (Physiologicrace), its definition refers in biological species or in mutation identical in form, butIn physiological character, particularly there is a class bion colony of significant difference on to the virulence of Different Crop kind.
The present invention is achieved by the following technical solutions:
The authentication method of a kind of soybean aphid biological strain is provided, comprises that (1) cultivation qualification uses soybean host; (2)Inoculation soybean aphid; (3) add up the quantity of the upper aphid of described soybean host and/or carry out aphid and infect classification; (4)Identify the biological strain of described soybean aphid.
Further,, in soybean seedling V1 growth period i.e. the first trifoliolate leaf phase, connect described soybean hostPlant soybean aphid.
Preferably, described soybean host at about 11-14 days after planting, inoculation soybean aphid. More preferably, existInoculation in after planting approximately 12 days.
Further, in step (2), utilize Inoculation Method, with moistening writing brush soybean described in every strainThe upper inoculation of host soybean aphid. Preferably, in step (2), inoculate 5 soybean aphids that bred in soybeanHost's trifoliolate leaf front.
Further, the quantity of the upper aphid of described soybean host is added up in the 14th day after soybean aphid.
Further, the aphid that carries out for the 21st day after inoculation soybean aphid infects classification.
Further, qualification comprises 5 resistant variety " Dowling that anti-aphides gene is clear and definite with soybean host(Rag1) ", " PI200538 (Rag2) ", " PI567543C (Rag3) ", " PI567301B (Rag5) " and“P203(Rag6)”。
Further, qualification also comprises 2 sense worm contrast soybean varieties " eastern agriculture 47 " and " Williams with soybean host82”。
Further, the application's method is for identifying the biological strain of described soybean aphid in greenhouse.
Advantage of the present invention: fast, strong operability, qualification result is reliable. Authentication method provided by the invention is passableRealize the object of Rapid identification soybean aphid biological strain in 40 days, after artificial infection aphid, post in conjunction with qualificationThe quantity of Your Majesty soybean aphid and two indexs of aphid infection ability classification are carried out the judgement of biological strain, can ensureQualification result reliable. Meanwhile, whole qualification process material requested is simple and easy to get, strong operability.
Detailed description of the invention
The authentication method of soybean aphid biological strain in greenhouse, comprises following sequential steps:
1) in greenhouse, cultivate 7 differential hosts: 2 sense worm kinds " eastern agriculture 47 " and " Williams82 "And 5 resistant varieties " Dowling (Rag1) " that anti-aphides gene is clear and definite, " PI200538 (Rag2) ", " PI567543C (Rag3) ", " PI567301B (Rag5) " and " P203 (Rag6) ". Each identification of species is established 5Repeat, greenhouse conventional cultivation management, treat growth of seedling to V1 growth period (the first trifoliolate leaf phase, after planting approximately12 days);
2) artificial infection soybean aphid, every strain qualification host inoculates 5 soybean aphids with moistening writing brush;
3) inoculate latter 14 days, the quantity of adding up aphid on each host plant;
4) inoculate latter 21 days, carry out the classification of aphid infection ability;
5) determine the anti-sense energy of aphid to different aphid-resistant gene Rag in conjunction with Aphed population and the classification of aphid infection abilityPower, the difference qualification soybean aphid biological strain with aphid to the anti-sense ability of different aphid-resistant genes.
Embodiment 1
The qualification of Shanghai soybean aphid and Jinan soybean aphid biological strain
1, in greenhouse, isolate two apart from each others, have " separate space " that double-deck gauze separates (can be also2 independently little greenhouses), plastic tub (dark 15cm, diameter 11cm, peat that substrate composition is 1:1:1,Vermiculite, perlite) in each sowing 7 soybean qualification hosts (eastern agriculture 47, Williams82, Dowling, PI200538, PI567543C, PI567301B, P203), each qualification host arranges 5 repetitions, approximately 12 daysAfter, Soybean Seedlings Growth is to V1 growth period, and the first trifoliolate leaf launches completely;
2, artificial infection aphid, will be the upper cultured Shanghai of sense worm soybean varieties " eastern agriculture 47 " and Jinan soybeanAphid, with the trifoliolate leaf front of 5 aphid to 7 soybean plant strains of the each inoculation of moistening writing brush, after inoculation aphidThe cultivation step management of watering routinely;
3, inoculation aphid after 14 days, add up 2 soybean aphids (Shanghai and Jinan) of originating 7 hosts separatelyQuantity on plant;
4, inoculation aphid after 21 days, according to following standard to aphid the infection conditions on host plant carry out classification,The index D of being injured I (Damageindex): DI1: several isolated aphids, and with dead aphid; DI2:A small amount of isolated aphid, follows the aphid larva of some new breedings; DI3: dense aphid clone, spreads all over hostPlant; DI4: dense aphid is cloned, and follow the symptom of being obviously injured of host plant, as plant is downgraded, leafCurling and the atrophy of sheet etc. Wherein DI≤2 of host plant, are defined as pest-resistant; DI > 2 is defined as sense worm.
5, judge that in conjunction with Aphed population and the index of being injured Shanghai aphid and Jinan aphid are to different aphid-resistant gene Rag'sInfection ability;
6, according to Shanghai aphid and Jinan aphid, the difference of the infection ability to different aphid-resistant gene Rag is identified the twoBiological strain.
Qualification result
Table 1. is inoculated the Aphed population of aphid each host plant after 14 days
* different letters represent significant difference (P < 0.05, LSD)
Table 2. inoculate aphid after 21 days soybean host be subject to aphid infect classification
The anti-sense performance to different host plant of table 3. Jinan and Shanghai soybean aphid
* S, sense worm; R, pest-resistant
Greenhouse qualification result shows, Jinan soybean aphid and Shanghai soybean aphid are infected different aphid-resistant gene Rag,Jinan aphid can be infected Rag1, can not infect Rag5; And Shanghai aphid can not be infected Rag1 gene, but canTo infect Rag5. Therefore, the different manifestations by two places aphid on Rag1 or Rag5 host, can distinguishThese two soybean aphids. The two belongs to different soybean aphid biological strains.
Embodiment 2
The qualification of Changchun soybean aphid and Kweiyang soybean aphid biological strain
1, in greenhouse, isolate two apart from each others, have " separate space " that double-deck gauze separates (can be also2 independently little greenhouses), plastic tub (dark 15cm, diameter 11cm, peat that substrate composition is 1:1:1,Vermiculite, perlite) in each sowing 7 soybean qualification hosts (eastern agriculture 47, Williams82, Dowling, PI200538, PI567543C, PI567301B, P203), each qualification host arranges 5 repetitions, approximately 12 daysAfter, Soybean Seedlings Growth is to V1 growth period, and the first trifoliolate leaf launches completely;
2, artificial infection aphid, will be the upper cultured Changchun of sense worm soybean varieties " eastern agriculture 47 " and Kweiyang soybeanAphid, with the trifoliolate leaf front of 5 aphid to 7 soybean plant strains of the each inoculation of moistening writing brush, after inoculation aphidThe cultivation step management of watering routinely;
3, inoculation aphid after 14 days, add up 2 soybean aphids (Changchun and Kweiyang) of originating 7 hosts separatelyQuantity on plant;
4, inoculation aphid after 21 days, according to following standard to aphid the infection conditions on host plant carry out classification,And the index D I(Damageindex that is injured): DI1: several isolated aphids, and with dead aphid; DI2:A small amount of isolated aphid, follows the aphid larva of some new breedings; DI3: dense aphid clone, spreads all over hostPlant; DI4: dense aphid is cloned, and follow the symptom of being obviously injured of host plant, as plant is downgraded, leafCurling and the atrophy of sheet etc. Wherein DI≤2 of host plant, are defined as pest-resistant; DI > 2 is defined as sense worm.
5, judge that in conjunction with Aphed population and the index of being injured Shanghai aphid and Jinan aphid are to different aphid-resistant gene Rag'sInfection ability;
6, the biological strain that the difference of the infection ability to different aphid-resistant gene Rag is identified the two according to two places aphid.
Qualification result
Table 4. is inoculated the Aphed population of aphid each host plant after 14 days
* different letters represent significant difference (P < 0.05, LSD)
Table 5. inoculate aphid after 21 days soybean host be subject to aphid infect classification
The anti-sense performance to different host plant of table 6. Changchun and Kweiyang soybean aphid
* S, sense worm; R, pest-resistant
Greenhouse qualification result shows, Changchun soybean aphid and Kweiyang soybean aphid are infected different aphid-resistant gene Rag,Changchun aphid can be infected Rag1, Rag2, Rag3, Rag5; And Kweiyang aphid can not be infected Rag2 and Rag3Gene. Therefore, the different manifestations by two places aphid on Rag2 or Rag3 host, can distinguish these twoSoybean aphid. The two belongs to different soybean aphid biological strains.
More than describe preferred embodiment of the present invention in detail. Should be appreciated that those of ordinary skill in the artJust can design according to the present invention make many modifications and variations without creative work. Therefore, all technology neckIn territory, technical staff is under this invention's idea on the basis of existing technology by logic analysis, reasoning or limitedTest available technical scheme, all should be in by the determined protection domain of claims.
Claims (7)
1. an authentication method for soybean aphid biological strain, is characterized in that, comprises that (1) cultivation qualification posts with soybeanMain; (2) inoculation soybean aphid; (3) add up the quantity of the upper aphid of described soybean host and/or carry out aphid infectionClassification; (4) identify the biological strain of described soybean aphid;
The quantity of adding up aphid on described soybean host on the 14th day after inoculation soybean aphid, at inoculation soybean aphidThe aphid that carries out for the 21st day after worm infects classification, and adopts differential host inoculating the Aphed population of aphid after 14 daysWith the method that the aphid infection ability classification after 21 days combines, qualification soybean aphid biological strain;
Wherein, qualification comprises that with soybean host 5 clear and definite resistant varieties " Dowling " of anti-aphides gene are Rag1Gene, " PI200538 " be Rag2 gene, " PI567543C " be Rag3 gene, " PI567301B "Rag5 gene and " P203 " are Rag6 gene, also comprise 2 sense worms contrast soybean varieties " eastern agriculture 47 " and“Williams82”。
2. the authentication method of a kind of soybean aphid biological strain according to claim 1, is characterized in that, in instituteState soybean host in soybean seedling V1 growth period, i.e. the first trifoliolate leaf phase, inoculation soybean aphid.
3. the authentication method of a kind of soybean aphid biological strain according to claim 1, is characterized in that, in instituteState soybean host at 11-14 days after planting, inoculation soybean aphid.
4. the authentication method of a kind of soybean aphid biological strain according to claim 1, is characterized in that, step(2) in, utilize Inoculation Method, described in every strain, on soybean host, inoculating soybean aphid with moistening writing brush.
5. the authentication method of a kind of soybean aphid biological strain according to claim 1, is characterized in that, step(2), in, inoculate 5 soybean aphids that bred in soybean host's trifoliolate leaf front.
6. the authentication method of a kind of soybean aphid biological strain according to claim 1, is characterized in that, described inAuthentication method is for identifying the biological strain of described soybean aphid in greenhouse.
7. the authentication method of a kind of soybean aphid biological strain according to claim 1, is characterized in that, described inSoybean aphid is from Shanghai, Jinan, Changchun and Kweiyang.
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| 大豆抗蚜资源筛选及大豆蚜生物型鉴定初探;张俊杰;《中国优秀硕士学位论文全文数据库》;20130715(第07期);第9-10,13,19，24-28，46页 * |
| 大豆蚜虫抗性鉴定技术及抗性资源筛选;孟凡立等;《大豆科学》;20100625;第29卷(第3期);第457-460页 * |
| 大豆蚜虫生物型鉴定初探;张俊杰等;《大豆科学》;20130425;第32卷(第2期);第234-237页 * |
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