CN1586134A - Polyploid rice breeding and detecting method with polyploid meiosis stability gene - Google Patents
Polyploid rice breeding and detecting method with polyploid meiosis stability gene Download PDFInfo
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Abstract
The present invention provides polyploid rice selectively breeding and identifying method with polyploid meiosis stability (PMeS) gene. The rice selectively breeding process includes: selecting parents; hybridization or complex hybridization; back crossing; doubling chromosome; identifying polyploid; selecting bearing property; observing meiosis behavior; self-crossing to form stable line; and identifying high bearing property. The identifying indexes include: polyploid line of Asia cultivated indica type rice or japonica rice; bearing rate not lower than 65 %; normal meiosis behavior; normal pollen shape and physiological activity and normal dyed pollution with 0.1 5 I-KI not lower than 95 %; bearing rate of hybrid of PMeS polyploid rice and other polyploid rice variety over 70 % and that of PMeS polyploid rice and wild rice variety over 65 %. The present invention solves the problem of low bearing rate of polyploid rice and may be used in breeding polyploid rice variety.
Description
Technical field
The present invention relates to a kind ofly have polyploid reduction division stability (Polyploid MeiosisStability, PMeS) the polyploid rice seed selection of gene and mirror survey method belong to the polyploid rice breeding
Technical field.
Background technology
Natural world higher plant about 70% all is a polyploid, main grain cotton oil crop such as wheat, cotton, rape etc. also all are polyploids, and they have experienced the process that dliploid is evolved to polyploid, and along with the increase of ploidy, their output all is doubled and redoubled.
But (Nakamori since Japanese scholar Nakamori in 1933 finds polyploid rice, E.Onthe occurrence of the tetraploid plant of rice Oryza sativa L., Proc, Imp.Acad.9,340-341,1933), have many scientists to be engaged in the research of paddy rice polyploid in the world, have the advantage that cane is sturdy, the seed change is big though have been found that polyploid rice, yet the polyploid rice breeding is made slow progress.Its main cause is that the ripening rate of polyploid rice is low, generally all is lower than 50%, do not reach the level that is applied to produce (Song Wenchang, Zhang Yuhua, four times in paddy rice change and to the influence of economical character and nutrient component, Acta Agronomica Sinica, 18 (2): 137-144,1992).Chinese patent (ZL00114471.5) inventor Cai gets field, Yuan Longping, the Lu Xinggui angle from the evolution thremmatology, proposition utilizes distant hybridization and double dominancy of polyploid seed selection super hybridization rice, it comprises that three phases is between subspecies, kind and the intergenomic hybrid vigor utilization, three kinds of technology promptly widen the affiliation distance, utilize wide compatibility gene and Ph gene (Pairinghomoeologous, homoeologous chromosomes pairing).Proposed the new approaches and the technology of polyploid breeding, enlarged the range of application of polyploid breeding, but gene relevant and strain with the high ripening rate of polyploid rice as yet not seed selection come out, the ripening rate of polyploid rice is still undesirable.
Summary of the invention
The objective of the invention is to select the polyploid rice of polyploid reduction division stability (PMeS) gene,, and make the PMeS strain of its formation be applied to the polyploid rice breeding in order to the low problem of solution polyploid rice ripening rate.
Technical scheme of the present invention is: a. selects the parent, b. hybridization or multiple cross, and c. selects the hybrid continuous backcross, d. chromosome doubling, e. polyploid identify that the f. fecundity is selected, g. meiotic behavior is observed, and strain is stablized in the h. selfing, the high fecundity hybridization assays of i..
Detailed process of the present invention is
A. select the parent
Select the parent to comprise long-grained nonglutinous rice, the japonica rice variety of Asia cultivated rice, possess the plant type compactness, big panicle many grains per panicle the close feature of grain, or has the long-grained nonglutinous rice or the japonica rice variety of wide compatibility gene and specialized character.Parent when specifically being seed selection Sg99012 (PMeS-1) is a leaf roll rice (indica type), Chinese sorghum rice (round-grained rice type), GC574 (round-grained rice type); Parent during seed selection HN2026 (PMeS-2) is a HT99104 (bare hull rice/japonica rice F
8Round-grained rice type strain), another name for Sichuan Province extensive 362.
B. hybridize or multiple cross
In the Sg99012 seed selection, obtain F with leaf roll rice * Chinese sorghum rice
1-a, leaf roll rice * GC574 obtains F
1-b is then with F
1-a and F
1-b carries out multiple cross.Can long-grained nonglutinous rice be maternal, be male parent with japonica rice, perhaps is maternal with japonica rice, is that male parent is mixed with different subspecies indica and japonica hybrids with the long-grained nonglutinous rice, then, and with two different hybrid F
1Carry out multiple cross; In the HN2026 seed selection, obtain F with HT99104 * another name for Sichuan Province extensive 362
1Hybrid.
C. select the hybrid continuous backcross
From hybridization or multiple cross offspring, select the plant type compactness, big panicle many grains per panicle, it is close grain, rice plant with 30% above ripening rate is the parent, with good japonica rice of economical character (cultivating kind to round-grained rice type direction) or long-grained nonglutinous rice (cultivating kind to indica type direction) is that backcross parent carried out for 3～5 generations and backcrosses, and obtains round-grained rice type or indica type backcross progeny that many proterties are inclined to backcross parent substantially.
D. chromosome doubling
Handle the paddy rice seedling in the tillering stage that previous step obtains with colchicine; Also can handle the callus that the rice young panicle tissue culture forms with colchicine.
E. polyploid is identified
Observe to identify that from external form promptly to show cane sturdy for polyploid, blade is abundant, and grain becomes big (bigger 1/3 to one times than dliploid), and most of grain have awns.The root tip chromosomes number of checking them is 2n=4x=48.
F. fecundity is selected
Species test method by thremmatology is carried out species test to the polyploid rice plant, selects to equal and be higher than 65% polyploid rice plant.
G. meiotic behavior is observed
Its cytogenetics feature meiotic behavior is observed.In reduction division period, to being defined as polyploid, and the pollen mother cells behavior of the polyploid rice plant of ripening rate 〉=65% is observed, normally promptly early stage, I was paired into the master with bivalent with those reduction division, 3～6 quadrivalents are arranged, and not or be lower than 1.0% univalent or high price body, mid-term, I did not have laggard,lagging chromosome, later stage I dye-free body bridge, no laggard,lagging chromosome, latter stage, I did not have the micronucleus appearance, and meiosis II is normal, and then forms normal tetrad, through 0.1%I-KI dyeing, but the polyploid rice individual plant of normal dyeing flower powder 〉=95% is picked out.
H. strain is stablized in selfing
With the polyploid rice individual plant selfing of selecting in the g process, make it stablize into strain system, breed into the PMeS strain of performance neat and consistent by strain system.
I. high fecundity hybridization assays
Stablize strain and polyploid long-grained nonglutinous rice, japonica rice variety hybridization with the PMeS in the h process, obtain F
1Hybrid promptly obtains having the polyploid hybrid rice of polyploid reduction division stability (PMeS) gene.Investigate F
1Ripening rate, general ripening rate 〉=more than 70%.
The present invention determines that the mirror survey index of polyploid rice strain, kind and the hybrid of polyploid reduction division stability (PMeS) gene is:
A. itself be long-grained nonglutinous rice in the Asia cultivated rice (Oryza sativa) or the polyploid strain of japonica rice.
B. have 〉=65% ripening rate.
C. meiotic behavior is normal.At I in early stage, chromosome pairing is preponderated with bivalent, and 3-6 quadrivalent arranged, but does not have or only have to be lower than 1.0% univalent or high price body (pairing chromosome that quadrivalent is above); At I in mid-term, later stage I and latter stage I and meiosis II in not or be lower than 0.5% laggard,lagging chromosome or micronucleus and occur.
D. POLLEN MORPHOLOGY and physiologically active are normal, through 0.1%I-KI dyeing, but normal dyeing flower powder 〉=95%.
E. with the polyploid offspring ripening rate of PMeS polyploid rice and other polyploid rice kind more than 70%, with the polyploid offspring ripening rate of A genome wild rice hybrid more than 65%.
The PMeS polyploid rice that the present invention produced has following good effect: 1, select a kind of polyploid rice that has higher ripening rate (〉=65%) itself; 2, utilize the polyploid offspring ripening rate of PMeS polyploid rice and other polyploid rice kind more than 70%, with offspring's ripening rate of A genome wild rice species hybridization more than 65%, solved the low problem of long-term unsolved polyploid rice ripening rate, made and utilize distant hybridization and double dominancy of polyploid seed selection super hybridization rice to become possibility; 3, utilize PMeS polyploid rice and polyploid rice male sterile line or polyploid rice to recover system's hybridization, can select polyploid male sterile line and recover system, provide condition for realizing the polyploid rice heterosis utilization with the high ripening rate characteristics of PMeS strain.Rice breeding is taken place by the dliploid level to the polyploid level, intervarietal cross to subspecies, plant between in addition the great change of genome intermolecular hybrid, can make full use of distant hybridization and double dominancy of polyploid, help selecting the polyploid rice kind or the hybrid of volume increase (more than 30%) significantly.
Embodiment
The present invention is further described with embodiment below.
The seed selection of embodiment 1:PMeS-1 (Sg99012)
A. select the parent
With leaf roll rice (indica type), Chinese sorghum rice (round-grained rice type), GC574 (round-grained rice type) is the parent, respectively selects 10 strains.
B. hybridize and multiple cross
Leaf roll rice * Chinese sorghum rice obtains F
1-a, leaf roll rice * GC574 obtains F
1-b is then with F
1-a and F
1-b carries out multiple cross.Select 50 hybrids.
C. select the hybrid continuous backcross
Pick out the round-grained rice type that inclines, the plant type compactness, sword-like leave is endured, and tassel is big, close hybrid individual plant 10 strains of grain, and backcrossing with GC574 as female parent obtains BC
1F
1(47 strain) therefrom selects the round-grained rice type that inclines, the plant type compactness, sword-like leave very, tassel is big, the closeer hybrid strain (30 strain) of grain and backcrosses with GC574 and obtain BC
2F
1(15 strain), continuous backcross obtains BC 3～4 times
4F
1(selecting 20 strains).
D. chromosome doubling
Handle the paddy rice seedling in the tillering stage that previous step obtains with colchicine.Get 1-2 BC of tillering of band
4F
1Hybrid rice shoot (20 strain), with blade in the horizontal partial application of base portion, reach half of base portion deeply, rice shoot is immersed in 0.2～0.3% colchicine solution then, in 1500～2000LX illumination, handle 48～56h under 20～25 ℃ of conditions, when treating that the enlargement shape appears in the rice shoot tip of a root, rice shoot is taken out, place the flowing water flushing more than 4 hours, then with shoot transplanting equipment in the earth alms bowl, in 1500～2000LX illumination, slow seedling is 10～15 days under 20 ℃ of conditions, treat newly to bear in the axil of rice shoot and tiller, select those and present blade thickness, the thicken rice shoot of feature of leaf look allows its self-sow after 10～15 days, they are transferred to outdoor planting, and these rice shoots may develop into polyploid rice.
E. polyploid is identified
Observe to identify that from external form promptly to show cane sturdy for polyploid, blade is abundant, and grain becomes big (bigger 1/3 to one times than dliploid), and most of grain have awns.The root tip chromosomes number of checking them is 2n=4x=48.
F. fecundity is selected
Species test method by thremmatology is carried out species test to the polyploid rice plant, selects to equal and be higher than 65% polyploid rice plant.
G. its cytogenetics feature meiotic behavior is observed
In reduction division period, to being defined as polyploid, and the pollen mother cells behavior of the polyploid rice plant of ripening rate 〉=65% is observed, normally promptly early stage, I was paired into the master with bivalent with those reduction division, 3～6 quadrivalents are arranged, and not or be lower than 1.0% univalent or high price body, mid-term, I did not have laggard,lagging chromosome, later stage I dye-free body bridge, no laggard,lagging chromosome, latter stage, I did not have the micronucleus appearance, and the reduction division II is normal, and then forms normal tetrad, through 0.1%I-KI dyeing, but the polyploid rice individual plant of normal dyeing flower powder 〉=95% is picked out (7 strain).
H. strain is stablized in selfing
With the polyploid rice individual plant selfing of selecting in the g process, make it stablize into strain system, breed into the PMeS strain of performance neat and consistent by strain system, name and be Sg99012.
I. high fecundity hybridization assays
Stablize strain Sg99012 and polyploid Dure (Xian), PA64s (Xian) inter breed crossing with PMeS, obtain F
1Hybrid PSR120 (Dure/sg99012), PSR012 (PA64
s/ Sg99012), investigate F
1Ripening rate be respectively 73.87%, 78.82%, strain be single plant yield all surpass 100 the gram.
The seed selection of embodiment 2:PMeS-2 (HN2026)
A. select the parent
With HT99104 (round-grained rice type), extensive 362 (indica type) in another name for Sichuan Province are the parent, respectively select 10 strains.
B. hybridize or multiple cross
HT99104 * another name for Sichuan Province is extensive 362, obtains 12 strain F
1
C. select the hybrid continuous backcross
Basic same with embodiment 1, be the backcross parent difference only with HT99104, backcrossing obtains BC after four times
4F
1Hybrid 15 strains.
D. chromosome doubling
What adopt is young fringe callus, induces to double in being added with the N6 liquid nutrient medium of 500mg/l colchicine, then is divided into the polyploid rice seedling.
E, f, g process are substantially the same manner as Example 1.
H. strain is stablized in selfing
With the polyploid rice individual plant selfing of selecting in the g process, make it stablize into strain system, breed into the PMeS strain of performance neat and consistent by strain system, name and be HN2026.
I. high fecundity hybridization assays
Stablize strain HN2026 and polyploid HT99128 (Xian), PA64s (Xian), HT99133 (Xian) inter breed crossing with PMeS, obtain F
1Hybrid PSR026 (PA64s/HN2026), PSR073 (HT99128/HN2026), PSR183 (HT99133/HN2026) investigate F
1Ripening rate be respectively 79.88%, 77.78%, 87.75%, strain be single plant yield all surpass 100 the gram.
Claims (3)
1. polyploid rice selection with polyploid reduction division stability gene is characterized in that process is:
A, select the parent, b, hybridization or multiple cross, are selected the hybrid continuous backcross at c, d, chromosome doubling, and e, polyploid identify that f, fecundity are selected, and g, meiotic behavior are observed, and strain is stablized in h, selfing, i, high fecundity hybridization assays.
2, by the described polyploid rice selection of claim 1, it is characterized in that detailed process is with polyploid reduction division stability gene:
A. select the parent
Select the parent to comprise long-grained nonglutinous rice, the japonica rice variety of Asia cultivated rice, possess the plant type compactness, big panicle many grains per panicle, the close feature of grain, or have the long-grained nonglutinous rice or the japonica rice variety of wide compatibility gene and specialized character, be specially leaf roll rice (indica type), Chinese sorghum rice (round-grained rice type), GC574 (round-grained rice type); HT99104, another name for Sichuan Province extensive 362;
B. hybridize or multiple cross
In the Sg99012 seed selection, obtain F with leaf roll rice * Chinese sorghum rice
1-a, leaf roll rice * GC574 obtains F
1-b is then with F
1-a and F
1-b carries out multiple cross, can long-grained nonglutinous rice be maternal, is male parent with japonica rice, perhaps is maternal with japonica rice, is that male parent is mixed with different subspecies indica and japonica hybrids with the long-grained nonglutinous rice, then, and with two different hybrid F
1Carry out multiple cross; Perhaps in the HN2026 seed selection, obtain F with HT99104 * another name for Sichuan Province extensive 362
1Hybrid;
C. select the hybrid continuous backcross
From the multiple cross offspring, select the plant type compactness, big panicle many grains per panicle, it is close grain, rice plant with 30% above ripening rate is the parent, with good japonica rice of economical character (cultivating kind to round-grained rice type direction) or long-grained nonglutinous rice (cultivating kind to indica type direction) is that backcross parent carried out for 3～5 generations and backcrosses, and obtains round-grained rice type or indica type backcross progeny that many proterties are inclined to backcross parent substantially;
D. chromosome doubling
Handle the paddy rice seedling in the tillering stage that previous step obtains with colchicine; Also can handle the callus that the rice young panicle tissue culture forms with colchicine;
E. polyploid is identified
Observe to identify that from external form promptly to show cane sturdy for polyploid, blade is abundant, and grain becomes big (bigger 1/3 to one times than dliploid), and most of grain have awns.The root tip chromosomes number of checking them is 2n=4x=48;
F. fecundity is selected
Species test method by thremmatology is carried out species test to the polyploid rice plant, selects to equal and be higher than 65% polyploid rice plant;
G. meiotic behavior is observed
In reduction division period, to being defined as polyploid, and the pollen mother cells behavior of the polyploid rice plant of ripening rate 〉=65% is observed, normally promptly early stage, I was paired into the master with bivalent with those reduction division, and 3～6 quadrivalents are arranged, and not or be lower than 1.0% univalent or high price body, mid-term, I did not have laggard,lagging chromosome, later stage I dye-free body bridge, no laggard,lagging chromosome, latter stage, I did not have the micronucleus appearance, and meiosis II is normal; Through 0.1%I-KI dyeing, but normal dyeing flower powder 〉=95%, and then form normal quadrantal polyploid rice individual plant and pick out;
H. strain is stablized in selfing
With the polyploid rice individual plant selfing of selecting in the g process, make it stablize into strain system, breed into the PMeS strain of performance neat and consistent by strain system;
I. high fecundity hybridization assays
Stablize strain and polyploid long-grained nonglutinous rice, japonica rice variety hybridization with the PMeS in the h process, obtain F
1Hybrid promptly obtains having the polyploid hybrid rice of polyploid reduction division stability (PMeS) gene.Investigate F
1Ripening rate, general ripening rate 〉=more than 70%.
3. by claim 1,2 described polyploid rice mirror survey methods, it is characterized in that mirror survey index is with polyploid reduction division stability gene:
A. itself be long-grained nonglutinous rice in the Asia cultivated rice (Oryza sativa) or the polyploid strain of japonica rice;
B. have 〉=65% ripening rate;
C. meiotic behavior is normal.At I in early stage, chromosome pairing is preponderated with bivalent, and 3～6 quadrivalents are arranged, but does not have or only have to be lower than 1.0% univalent or high price body (pairing chromosome that quadrivalent is above); At I in mid-term, later stage I and latter stage I and meiosis II in not or be lower than 0.5% laggard,lagging chromosome or micronucleus and occur;
D. POLLEN MORPHOLOGY and physiologically active are normal, through 0.1%I-KI dyeing, but normal dyeing flower powder 〉=95%;
E. with the polyploid offspring ripening rate of PMeS polyploid rice and other polyploid rice kind hybrid more than 70%, with the polyploid offspring ripening rate of A genome wild rice hybrid more than 65%.
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