CN103081797A - Method for inducing corn haploid - Google Patents
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Abstract
The present invention discloses a method suitable for commercially inducing a corn haploid. The method comprises the following steps: carrying out hybridization of a corn haploid induction line I and a corn haploid induction line II to obtain a progeny recorded as an induction line hybrid, adopting the induction line hybrid as a male parent, carrying out hybridization of the induction line hybrid and a female parent material expected to obtain a haploid to harvest F0 hybridization kernel, and taking the haploid kernel to obtain the corn haploid, wherein the corn haploid induction line I and the corn haploid induction line II are different. Compared with the traditional method, the method of the present invention has the following characteristics that: on the basis of maintaining of a high induction rate, agronomy characteristics of the male parent (induction line hybridization combination) in the haploid induction is substantially increased, and especially a plant growth condition, a pollen disseminating amount, a pollen disseminating duration time, adaptability, resistance, and the like are substantially increased so as to improve haploid induction efficiency, provide a guarantee for efficient haploid induction, and provide possibility for large-scale haploid induction.
Description
Technical field
The present invention relates to a kind of haploid method of suitable commercialization inducing maize.
Background technology
The seed selection of inbred line is the important step in corn breeding, is almost determining the success or failure of hybrid variety breeding.Because corn is the different flower of monoecism, carries out amphigenetic crop, be mainly therefore to utilize the amphigenetic mode selecting and breeding corn of corn inbred line, preparation Maize Combination, screening corn hybrid seed in corn breeding work.The Breeding Process of traditional corn inbred line must be through selfing and the selection in many generations for many years, and kind of seed selection needs the time of upper ten year usually, not only the cycle long, and need to consume a large amount of human and material resources and financial resources.Therefore how shortening breeding process, improving breeding efficiency is the target that breeding men are pursued always.
The monoploid technology is as a new and high technology, for breeding man quick breeding, select inbred line to bring wide prospect.Corn monoploid is the product of gametophytic apomixis, comprises etheogenesis and the large class of parthenogenesis two.The monoploid that obtains is carried out artificial doubling, only need a generation just can obtain zygoid, greatly shorten breeding time.Since Guha (Guha S, Maheshwari S C.In vitro production of embryos from anther of Dature[J] .Nature, 1964,204:497) disclosed after approach can obtain monoploid by experiment the method that the method by group training approach Breeding of Inbred Lines is considered to get a good chance of through anther culture monoploid.But because the anther culture difficulty is large, technical operation is complicated, and the genotype dependence is very strong, and the inbred line that selects good inbred line not necessarily, is difficult to utilize aborning.
The main path that another kind of monoploid produces is that to induce be that the hybridized induction approach produces maternal (female) monoploid, referred to as Parthenogenesis haploid.Parthenogenesis haploid can produce by approach such as biotic induce and abiotic inducing (as chemical induction), but abiotic problem of inducing and a lot of difficulties of the same existence of flower pesticide group training are difficult to use in breeding.Biotic induce is to utilize the biotic induce material to make to induce the object Haploid production.Coe (Coe E H.Aline of maize with high haploid frequency[J] .Am Nat, 1959,93:381-382) found that in nineteen fifty-nine it is Stock6 that the corn parthenogenesis is induced.Constantly transformation on its basis afterwards, as change Navajo (ACR-nj) genetic marker gene over to, go out monoploid by seed coat color and plant color identification, become possibility thereby make hybridized induction monoploid carry out haploid breeding.But also find in application in practice, the inductivity of Stock6 is very low (being about 3%) also, and have a lot of serious defectives, and to responsive to temperature, self-fruitfulness is poor as male flower, and the fringe Maize Ear Rot is serious, the Navajo genetic marker a little less than.Therefore, utilizing Stock6 to induce haploid key is to overcome above-mentioned shortcoming under the prerequisite that keeps its inducibility.abroad start to walk in this respect early, and obtained greater advance, having selected some good inducing is, WS14 (Lashermes P as France, Beckert M.Genetic control of maternal haploidy in maize (Zea mays L.) and selection of haploid inducing lines[J] .Theor Appl Genet, 1988, 76 (3): 405-410), the KEMS of the former Soviet Union (Shatskaya O A.et al, Mass induction of maternal haploids in corn[J] .Maize Genet Coop Newslett, 1994, 68:51), the MHI of Moldova (Chalyk S T.Creating new haploid-inducing lines of maize[J] .Maize Genet Coop Newslett, 1999, 73:53-54), RWS (the F.K.Rober et al. of Germany, In vivo haploid induction in maize-Performance of new inducers and significance of doubled haploid lines in hybrid breeding[J] .Maydica, 2005, 50:275-283) etc.Inducing of these new choosings is not only to adapt to local environment, and its inductivity and also further raising of genetic marker, and value is well beyond Stock6.Yet although inducing of different new choosings is that its inductivity is greatly improved, the performance on economical character is still satisfied not to the utmost, and especially loose powder and self-fruitfulness are not satisfactory.Therefore, induce the loose powder and the self-fruitfulness that are extremely important guaranteeing how to improve under the prerequisite of inductivity.
Summary of the invention
An object of the present invention is to provide a kind of haploid method of inducing maize.
The haploid method of inducing maize provided by the present invention comprises the steps: that the offspring who obtains is denoted as and induces series hybrid with corn haploid inducing line I and corn haploid inducing line II hybridization; Take the described series hybrid of inducing as male parent, obtain haploid maternal material with expectation and hybridize, results hybridization in present age seed is got the monoploid seed, namely obtains corn monoploid; Corn haploid inducing line I is different with corn haploid inducing line II.
In said method, in described corn haploid inducing line I and corn haploid inducing line II hybridization, the mode of described hybridization is quadrature or reciprocal cross.
In said method, described inducing series hybrid as male parent, obtain during haploid maternal material hybridizes with expectation, the method for described hybridization is artificial pollination or spontaneous pollination.
In said method, in described spontaneous pollination, the mode of plantation is as follows: with described male parent and maternal alternate plantation, and make planting area and isolation on every side, before the male parent loose powder, female parent is carried out emasculation, then it is maternal to rely on natural force that the pollen of male parent is arrived, and then realizes pollination.
In said method, described corn haploid inducing line I and corn haploid inducing line II all are selected from: corn haploid inducing line CAUHOI, corn haploid inducing line UH400, corn haploid inducing line CAU079 and corn haploid inducing line CAU5.
In said method, described female parent is inbred line, basic population or the crossbreed that meets breeding objective.
In said method, described basic population is the maize population that forms by after a plurality of inbred lines or Cultivar mixtures pollination.
In said method, described crossbreed is single cross hybrid, triple hybrid or double cross hybrid.
Corn haploid inducing line I and corn haploid inducing line II are that two different inducing of inducibility that all have are;
Spontaneous pollination refers to the certain proportion maternal material of wrong alternate plantation of phase and induces series hybrid, the surrounding isolation, before loose powder to maternal material artificial emasculation, thereby the pollen that relies on wind-force to propagate male parent reaches the purpose of pollination to carry out haploid induction.
Above-mentioned certain proportion can be 4: 2 or 6: 3 or other ratios, and wherein the line number of maternal material is more than inducing series hybrid, and this ratio can suitably be adjusted according to the size of different crossbreed loose powder amounts;
Above-mentioned wrong phase plantation refers to according to maternal material and weaves silk sooner or later and induce the loose powder of series hybrid sooner or later to arrange male parent and maternal implantation time, can normally be pollinated with the filigree that guarantees maternal material;
Above-mentioned alternate plantation refers to male parent and female parent alternate plantation according to the above ratio, and guarantees the current plantation of paternal hybrid kind so that the operation of the fields such as emasculation;
The isolation of above-mentioned surrounding refer at least 200 meters of surroundings without other corn plantings to guarantee isolation effect;
Maternal material can be inbred line, basic population or the crossbreed that meets breeding objective or research needs.
Above-mentioned basic population refers to the maize population that formed after by the certain way mixed pollination by a plurality of inbred lines or kind.
Above-mentioned crossbreed refers to single cross hybrid, triple hybrid or double cross hybrid.
Outstanding advantages of the present invention is:
(1) induce the inductivity of series hybrid to compare with two parents, still kept higher inductivity, the inductivity of the combination that even has is higher than parents;
(2) on the basis that keeps higher inductivity, induce the economical character of series hybrid to be greatly improved: field growing is all right; The ability of resisting bad natural environment is stronger, and adaptability is wider; The loose powder performance improves greatly, and pollen amount is larger, and the duration is longer;
(3) due to induce be growing way generally a little less than, inducing when with the alternate plantation of other maternal materials (as crossbreed) is often can't normal growth; And induce series hybrid its economical character under the prerequisite that keeps higher inductivity greatly to improve (as plant growing way, loose powder amount, loose powder duration, adaptability, resistance etc.); can satisfy the requirement with other breeding material interlacing plantations; for efficient haploid induction provides assurance, provide possibility thereby carry out haploid induction for scale.
(4) with maternal material and male parent material interlacing plantation, take the mode of maternal material artificial emasculation to realize spontaneous pollination, removed the link of artificial pollination from, can greatly raise the efficiency.
Embodiment
The experimental technique that uses in following embodiment is conventional method if no special instructions.
In following embodiment, material used, reagent etc., if no special instructions, all can obtain from commercial channels.
1, High oil haploid inducer CAUHOI:CAUHOI is High oil haploid inducer (Liang Li, Xiaowei Xu, Weiwei Jin, Shaojiang Chen.Morphological and molecular evidences for DNA introgression in haploid induction via a high oil inducer CAUHOI in maize.Planta, 2009,230:367-376), inductivity is 2%, have the R-nj mark, the public can obtain from China Agricultural University.
2, high frequency haploid inducing line UH400:UH400 haploid-induction reaches 8% left and right, document " Prigge V, S á nchez C, Dhillon B S, SchipprackW, Araus JL,
M, Melchinger AE.Doubled haploids in tropical maize:I.Effects of inducers and source germplasm in vivo haploid induction rates.Crop Sci, 2011,51:1-9. " in disclosed; by the seed selection of German Huo Enhaimu university, the public can obtain from China Agricultural University.
3, high frequency haploid inducing line CAU5 and CAU079: haploid inducing line CAU5 and CAU079 be China Agricultural University's seed selection two generation haploid inducing line, average inductivity all reaches more than 10%, has the R-nj mark, " Chen Shaojiang; Li Liang; Li Haochuan. Haploid Breeding of Maize technology [M]. China Agricultyre University Press, 2009. " disclosed in a book, the public can obtain from China Agricultural University.
4, the high oil 5580 of Nongda108 and middle agricultural university is available from the greatly healthy scientific and technological development of Beijing middle peasant company.
5, Zheng Dan 958 is available from Beijing DeNong seeds Co., Ltd.
Embodiment 1, not isogeneous induction are the Agronomic of selfing and hybrid combination
Selecting CAUHOI, UH400 and inducing of two new choosings is CAU5 and CAU079.Hainan was the phase mutual cross with above-mentioned 4 in 2009, gathered in the crops contemporary seed (be F1 generation).Summer in 2010 was planted each F1 material in the Changping County, Beijing experiment station, relatively the economical character in parent and F1 generation.Economical character comprises: plant height, fringe position, loose powder phase, the phase of weaving silk etc.
Plant height: plant height refers to that ground arrives the distance at plant tassel top;
The fringe position: the fringe position is high to be referred to from ground to the distance of female fringe base portion;
Be seeded into the number of days of weaving silk: the filigree of the index futures plant gynoecium of weaving silk stretches out about 2cm from bract, is defined as when every row has 50% plant to weave silk the phase of weaving silk;
Be seeded into the loose powder number of days: the loose powder phase refers to the date of tassel middle part beginning loose powder, and every row has 50% plant loose powder to be defined as the loose powder phase;
The loose powder duration: refer to finish lasting number of days from beginning most pollen to occur to loose powder;
Pollen amount: refer to what of plant institute pollen flying.
Table 1, not isogeneous induction are and induce the economical character of series hybrid to compare
As can be seen from Table 1, inducing is that plant height itself is lower, is only on average about 130cm; And when induce be hybridization after, due to heterotic reason, plant height significantly increases, and on average is about 170cm.Except plant height, the fringe position is high also significantly to be increased, and plant is more sturdy.Inducing is that the pollen amount of middle UH400 is less, but and other to induce be that pollen amount after hybridization is all very large.The loose powder duration slightly increases, and changes breeding time (weaving silk and the loose powder number of days from being seeded into) little.In addition, be more sturdy owing to inducing the series hybrid ratio to induce, therefore stronger at capacity for the resistance to lodging.If carry out haploidly inducing in a large number; continuous increase along with human cost; necessary form large-scale production monoploid with isolated area: namely inner in the soil of a sealing (on every side without other corns); obtaining haploid material with needs does maternal; be or induce to be that hybrid combination is hybridized as male parent to induce, before loose powder, maternal artificial emasculation can be realized " natural inducing ".But, be that plant is shorter owing to inducing, and growing way a little less than, be difficult to satisfy scale and induce haploid demand, the form of therefore inducing series hybrid can satisfy well the scale isolated area is produced haploid requirement.
Case study on implementation 2, not isogeneous induction series hybrid and its parent's inductivity diversity ratio
Tested summer in 2010 and carry out in the Changping County, Beijing experiment station.Take Zheng Dan 958 as Testers (as female parent) respectively with above-mentioned table 1 in the parent induce and be and induce series hybrid to hybridize, each is induced and is or induces series hybrid to hybridize at least the maternal materials of 10 fringes; Gather in the crops afterwards cross-pollinated seed, threshing and carry out the monoploid evaluation, relatively the parent induces the inductivity difference that is, induces series hybrid.
The method that monoploid is identified is as follows:
(1) the hybridization fruit ear of Zheng Dan 958 is classified threshing to carry out haploid evaluation by different male parents sources;
(2) observe the color of hybridization seed embryo and endosperm aleurone layer, wherein embryo section and endosperm aleurone layer are all purpuric for hybridizing dliploid, when the endosperm aleurone layer purple is arranged and embryo section without the pseudohaploid that is classified as of purple, all be classified as the pollen contamination seed without purple when endosperm aleurone layer and embryo section.Statistics is hybridized the number of dliploid and pseudohaploid respectively.
(3) all pseudohaploid seeds are planted in breeding station, Hainan observation Reducing sugar in Hainan in 2010, if plant is tall and big or stem stalk purple is designated as pseudohaploid, if plant is short and small and stem stalk green is designated as monoploid, add up haploid number.
According to haploid number and hybridize diplontic number and can calculate haploid inductivity: haploid-induction=haploid number/(hybridization dliploid number+pseudohaploid number) * 100%.
Through identifying, each parent induce be and induce series hybrid induce efficient as follows:
CAUHOI，2.35％；
UH400，8％；
CAU079，9％；
CAU5，10％；
CAUHOI/UH400，4.80％；
CAUHOI/CAU079，7.14％；
CAUHOI/CAU5，8.36％；
UH400/CAU079，13.58％；
UH400/CAU5，13.62％；
CAU079/CAU5，6.03％。
By the above results as can be known, when two induce be hybridization after, its hybrid combination (we are referred to as to induce series hybrid) still has higher inducibility, its inductivity is often between parents' inductivity.When inducing the inductivity that is all high for two, it induces the inducibility of series hybrid might surpass parents, such as UH400, CAU079, the inducibility of CAU5 is all higher, UH400/CAU5, the inductivity of UH400/CAU079 has surpassed parents, reaches more than 13%.There is the inductivity of 5 to surpass 5% in above-mentioned 6 inductivities of inducing series hybrid.Therefore, carry out haploid induction to induce series hybrid as male parent, can satisfy induce in a large number haploid needs fully.
Case study on implementation 3, induce the effect of inducing of series hybrid UH400/CAU5 and different maternal material
It is that the hybridization assembly induces series hybrid UH400/CAU5 to carry out haploid induction that winter in 2009 induced with UH400 and two of CAU5 in experiment station, Hainan.
Summer in 2010 was made male parent and different hybridization of female parent with UH400/CAU5, and 3 business crossbreed of initial option hybridize as female parent, relatively induced the effect of inducing of series hybrid under different material backgrounds.
Concrete Crossing system: with male parent and maternal alternate plantation, and make planting area and isolation (surrounding at least 200 meters without other corn plantings) on every side, before the male parent loose powder, female parent is carried out emasculation, then it is maternal to rely on wind-force that the pollen of male parent is arrived, and then realizes pollination.
Different hybridization of female parent kinds are respectively:
Crossbreed 1, Nongda108 can obtain from national corn improvement center
Crossbreed 2, the high oil 5580 of middle agricultural university can be from national corn improvement center;
Crossbreed 3, Zheng Dan 958, can obtain from Henan Academy of Agricultural Sciences.
Experimental design is randomized complete-block design, establishes altogether following 10 processing.
Process 1 take crossbreed 1 as maternal, UH400/CAU5 is hybridized as male parent, hybridization at least 10 fringes;
Process 2 take crossbreed 2 as maternal, UH400/CAU5 is hybridized as male parent, hybridization at least 10 fringes;
Process 3 take crossbreed 3 as maternal, UH400/CAU5 is hybridized as male parent, hybridization at least 10 fringes;
The fruit ear threshing of the solid uniformity of each processing selecting after results.Get at random respectively 1000 of the hybridization seeds of each processing, carry out the monoploid evaluation.The monoploid authentication method is with case study on implementation 2.
The field test result shows, in 1000 hybridization seeds:
Processing has 98 to be monoploid in 1, haploid-induction is 9.8%;
Processing has 133 to be monoploid in 2, haploid-induction is 13.3%;
Processing has 125 to be monoploid in 3, haploid-induction is 12.5%.
Show from the above results, when doing maternal with the material of different background and inducing series hybrid to induce, inductivity can satisfy the haploid needs of extensive acquisition fully all more than 3%.
Claims (6)
1. the haploid method of inducing maize, comprise the steps: that the offspring who obtains is denoted as and induces series hybrid with corn haploid inducing line I and corn haploid inducing line II hybridization; Take the described series hybrid of inducing as male parent, obtain haploid maternal material with expectation and hybridize, results hybridization in present age seed is got the monoploid seed, namely obtains corn monoploid; Corn haploid inducing line I is different with corn haploid inducing line II.
2. method according to claim 1 is characterized in that: in described corn haploid inducing line I and corn haploid inducing line II hybridization, the mode of described hybridization is quadrature or reciprocal cross.
3. method according to claim 1 and 2 is characterized in that: described inducing series hybrid as male parent, obtain during haploid maternal material hybridizes with expectation, and the method for described hybridization is artificial pollination or spontaneous pollination.
4. according to claim 1 and 2 or 3 described methods, it is characterized in that: in described spontaneous pollination, the mode of plantation is as follows: with described male parent and maternal alternate plantation, and make planting area and isolation on every side, before the male parent loose powder, female parent is carried out emasculation, rely on again natural force that the pollen of male parent is arrived maternal, and then realize pollination.
5. arbitrary described method according to claim 1-4, it is characterized in that: described corn haploid inducing line I and corn haploid inducing line II all are selected from: corn haploid inducing line CAUHOI, corn haploid inducing line UH400, corn haploid inducing line CAU079 and corn haploid inducing line CAU5.
6. arbitrary described method according to claim 1-5, it is characterized in that: described female parent is inbred line, basic population or the crossbreed that meets breeding objective.
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| CN112219717A (en) * | 2020-10-14 | 2021-01-15 | 金苑(北京)农业技术研究院有限公司 | Method for inducing and identifying haploid generated by corn |
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| CN105660381A (en) * | 2016-02-25 | 2016-06-15 | 安徽省农业科学院烟草研究所 | Method for producing large-scale induction pollen sources in maize haploid breeding |
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| US11154022B2 (en) | 2018-11-27 | 2021-10-26 | The United States Of America, As Represented By The Secretary Of Agriculture | Production of haploid lolium |
| CN112005878A (en) * | 2020-08-24 | 2020-12-01 | 中国农业大学 | Method for rapidly breeding corn haploid induction line and application thereof |
| CN112219717A (en) * | 2020-10-14 | 2021-01-15 | 金苑(北京)农业技术研究院有限公司 | Method for inducing and identifying haploid generated by corn |
| CN112219717B (en) * | 2020-10-14 | 2022-03-22 | 金苑(北京)农业技术研究院有限公司 | Method for inducing and identifying haploid generated by corn |
| CN113575412A (en) * | 2021-09-02 | 2021-11-02 | 河北冀农种业有限责任公司 | Breeding method of corn haploid induction line |
| CN116671435A (en) * | 2023-07-07 | 2023-09-01 | 云南省农业科学院粮食作物研究所 | Method for enhancing induction efficiency of corn haploid |
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