CN107148910A - A kind of method for cultivating nearly allotetraploid corn material - Google Patents
A kind of method for cultivating nearly allotetraploid corn material Download PDFInfo
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- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01H—NEW PLANTS OR NON-TRANSGENIC PROCESSES FOR OBTAINING THEM; PLANT REPRODUCTION BY TISSUE CULTURE TECHNIQUES
- A01H1/00—Processes for modifying genotypes ; Plants characterised by associated natural traits
- A01H1/06—Processes for producing mutations, e.g. treatment with chemicals or with radiation
- A01H1/08—Methods for producing changes in chromosome number
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- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01H—NEW PLANTS OR NON-TRANSGENIC PROCESSES FOR OBTAINING THEM; PLANT REPRODUCTION BY TISSUE CULTURE TECHNIQUES
- A01H1/00—Processes for modifying genotypes ; Plants characterised by associated natural traits
- A01H1/02—Methods or apparatus for hybridisation; Artificial pollination ; Fertility
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Abstract
The invention discloses a kind of method for cultivating nearly allotetraploid corn material, this method is included using the corn allohexaploid of self-fertile as female parent, using corn as paternal hybrid, in F1Selection pollen has the plant selfing of fertility in generation；Then root tip chromosomes number and composition to offspring is identified, and pollen fertility is detected, selection meets target plant selfing, until selecting total chromosome number for 38~40, wherein number containing maize chromosome 22~28, tetraperennial teosinte chromosome number be 12~15, finger-like friction standing grain chromosome number be 0~2, and the high plant of pollen fertility, is the nearly allotetraploid corn material of the present invention.The present invention has been bred as the fertile nearly allotetraploid corn of pollen in the case of aneuploidy chromosome breakthroughly first, can be as the bridge being transferred to the desirable genes of corn Wild related germplasm in cultivated maize, and be that corn gene function and fertility research provide material.
Description
Technical field
The invention belongs to corn distant hybridization breeding method field, and in particular to one kind cultivates nearly allotetraploid corn material
The method of material.
Background technology
Corn is transition silos crop, is also the important raw material of industry.As the improvement of people's living standards, to corn
Demand is also sharply increased.It is the main path for improving corn yield to improve per unit area yield, but is due to the presence of corn germplasm hereditary basis
Narrow problem, limits the cultivation of breakthrough corn variety.Therefore, to meet the demands of production and living, in the urgent need to widening
The new corn germ plasm resource with initiative.
The corn Wild related germplasm such as teosinte and friction standing grain forms a variety of excellent in long-term evolutionary process
Gene；There is very strong resistance to corn northern leaf blight and southern leaf blight such as teosinte, it is immune to some virosis, also with strong stress resistance and suitable
The advantages of Ying Xingguang；Friction standing grain has good disease and insect resistance, such as resistant to corn root snout moth's larva, corn Yi pipe aphids, Resistant Gene To Rust,
The anti-leaf blight of corn, friction standing grain also has the good characteristics such as cold-resistant, drought resisting, Salt And Alkali Tolerance；In addition, friction standing grain also has seed nutrition
Abundant, protein content is high, the advantages of digestibility is higher.These good characteristics of corn Wild related germplasm can be used for widening jade
The Germplasm Bases of rice.But be due to affiliation farther out, have that exist after the reasons such as reproduction isolation, they and corn hybridization can not
It is solid, or filial generation is sterile or fertility is extremely low, makes the desirable genes of Wild related germplasm be difficult to imported into corn material.
MTF-1 (Tripsazea creammaize T., 2n=76) is Sichuan Agricultural University with the tetraploid quoted from the U.S.
Corn (Zea mays L., 4n=40) and tetraploid gama grass (or finger-like friction standing grain, Tripsacum
Dactyloides L., 2n=72) between intergeneric hybrid F1For female parent, with tetraperennial teosinte (Zea
Perennis, 2n=40) the corn allohexaploid that is obtained for paternal hybrid；MTF-1 chromosome number is 76, is a bag
Containing a full set of maize chromosome group (20), gama grass genome (36) and tetraperennial teosinte chromosome
The intergeneric hybrid of group (20), is the non-existent new species of nature.MTF-1 Classification And Nomenclatures are jade by Sichuan Agricultural University
Broomcorn millet standing grain belongs to Yu Qi and drenches grass seeds, and its Latin specific name is named as into Tripsazea creammaize T..
MTF-1 has female fertile characteristic, with corn (Zea mays L.), teosinte (Zea perennis L.) or
Friction standing grain (Tripsacum dactyloides L.) is that male parent can all produce solid offspring to MTF-1 pollinations.Using this characteristic,
Using MTF-1 as bridge material, the desirable genes in teosinte or friction standing grain can be transferred in corn.But it is due to that MTF-1 is deposited
The wild characteristic such as more than the perennial, tiller, its application is by larger limitation.At present, MTF-1 be mainly used in cultivate it is perennial, point
Tiller is more and forage maize kind (CN103609428A, CN103548674A) of vegetative propagation, it is difficult to directly in corn breeding
Using.
Through retrieval, do not find teosinte and friction standing grain desirable genes transformation to corn material using MTF-1 as bridge
In report.
The content of the invention
Present invention aims at provide a kind of method for cultivating nearly allotetraploid corn material.
To achieve the above object, technical scheme is as follows：
A kind of method for cultivating nearly allotetraploid corn new material, comprises the following steps：
(1) it is female parent with MTF-1 (Tripsazea creammaize T., 2n=76), with corn, in the spinning loose powder phase
(Zea mays L., 2n=20) is that male parent is hybridized, and seed is harvested when ripe, F is obtained1Generation；
(2) F, is planted1Generation, in tasseling stage with iodine-potassium iodide decoration method to F1Pollen fertility detection is carried out for material；Selection
The plant that pollen has fertility carries out selfing, and seed is harvested when ripe, obtains F2Generation；
(3) F, is planted2Generation, to the desirable tip of a root before tasseling stage since tri-leaf period, using the side of genomic in situ hybridization
Method to root tip chromosomes number and genome into identifying, and to F2Investigated for the tiller number of plant；In tasseling stage
Pollen fertility is detected with iodine-potassium iodide decoration method；It is 41~43 to select root tip chromosomes sum, wherein containing corn
(Zea mays L.) chromosome number 25~26, tetraperennial teosinte (Zea perennis L.) chromosome number 14~
15, finger-like friction standing grain (Tripsacum dactyloides L.) chromosome number 1~2, tiller number be 1~3, and pollen
The plant that fertility is more than 40% carries out selfing, and seed is harvested when ripe, obtains F3Generation；
(4) F, is planted3Generation, according to the genomic in situ hybridization method described in step (3) to root tip chromosomes number and dyeing
Body composition is identified, and pollen fertility is detected with iodine-potassium iodide decoration method in tasseling stage；Selective staining body sum is
39~40, wherein containing maize chromosome number 24~28, tetraperennial teosinte chromosome number 12~15, finger-like
Rub standing grain chromosome number 0~2, and the plant of no tiller and pollen fertility more than 50% carries out selfing, and seed is harvested when ripe,
Obtain F4Generation；
(5) F, is planted4Generation, according to step (3) the genomic in situ hybridization method to root tip chromosomes number and chromosome
Composition is identified, and pollen fertility is detected with iodine-potassium iodide decoration method in tasseling stage；Chromosome is total in testing result
Number is 38~40, wherein number containing maize chromosome is 22~28, tetraperennial teosinte chromosome number is 12~15
Bar, finger-like friction standing grain chromosome number are 0~2, the plant of no tiller and pollen fertility more than 70%；This total chromosome number
At 40 or so, and simultaneously containing corn, tetraperennial teosinte or finger-like friction standing grain two or more species dye
The plant of colour solid group, is the nearly allotetraploid corn material of the present invention；Selected plant selfing is reserved seed for planting.
Described in above method step (1) MTF-1 (Tripsazea creammaize T., 2n=76) (Su Yuegui,
Sichuan Agricultural University's Master's thesis, 2009) it is that with the tetraploid corn quoted from the U.S., (Zea mays are L.4n for Sichuan Agricultural University
=40) and tetraploid gama grass (or finger-like friction standing grain, Tripsacum dactyloides L., 2n=72) between
Intergeneric hybrid F1It is what paternal hybrid was bred as with tetraperennial teosinte (Zea perennis, 2n=40) for female parent
Corn allohexaploid；MTF-1 chromosome number is 76, is one and is rubbed comprising a full set of maize chromosome group (20), orchardgrass shape
Wipe standing grain genome (36) and tetraperennial teosinte genome (20) intergeneric hybrid, be a nature not
The new species of presence.
Described corn allohexaploid MTF-1 (Tripsazea creammaize T.) is perennial, vegetative propagation,
Plant is healthy and strong, uprightly grows thickly, well developed root system；It is that plant height reaches as high as 335cm after 310cm, tasseling stage to take out average plant height of male period,
The thick 9.8-12.9mm of stem, the tiller of tasseling stage individual plant is up to 50-60, and tiller is different not substantially, and whole life with stem stem rough error
New root tiller is continuously generated in cycle；Blade is dark green elongated, the long 61-81cm of leaf, leaf width 5.1-5.9cm, single stalk blade
Number is 20-23 pieces；Stem top male flower belongs to panicle, the long 27-30cm of inflorescence, pollen height infertility, on stalk node it is raw
5-7 branch, branch top is the female flower of spike, and 9-18 small ear is arranged raw on cob in duplicate rows or four rows.Plant
Strain tassel, leaf sheath and bract are darkviolet.MTF-1 hybridization can obtain a small amount of seed, and seed glume is hard, brown or grey brown
Color.The waterlogging-resistant ability of plant cold-resistant is strong, remains to persistently grow new tiller under the conditions of 0-5 and long-term water logging.
MTF-1 is mainly carried out numerous by asexual reproduction methods such as point pocket breeding, stalk cuttage or other vegetative manners
Grow.The public can obtain MTF-1 (Tripsazea creammaize are T.2n=76) biomaterial from Sichuan Agricultural University.
Corn described in above method step (1) refers to self-mating system, cenospecies, Local variety or collective species etc..
Iodine-potassium iodide decoration method described in above method step (2), (3), (4) or (5) is with reference to the expensive Master's thesis of Su Yue
(Sichuan Agricultural University's Master's thesis, 2009).
Genomic in situ hybridization method reference Tang Q described in above method step (3), (4) or (5) etc. 2005
Disclosed method (.Crop such as Tang Q Sci, 2005,45:717-721).
Nearly allotetraploid corn material described in above method step (5), its chromosome number is 40 or so and (is about
4 times of maize chromosome radix 10), and chromosomal origin (contains corn, the perennial class of tetraploid is beautiful simultaneously in different species
The chromosome of rice and/or finger-like friction standing grain), therefore referred to as near allotetraploid corn.
The nearly allotetraploid corn material selected in the above method, can be for assembling cross materialses, in being used as
Between bridge material, the excellent genes of Wild related germplasm are imported into common diploid corn, the Germplasm Bases of corn are widened.
The present invention has the advantage that and beneficial effect：The present invention first be bred as total chromosome number be 40 or so and comprising
Corn and tetraperennial teosinte chromosome, or it is also beautiful containing nearly allotetraploid that rub standing grain chromosome, to have fertility
Rice material, the i.e. present invention have been bred as the fertile nearly allotetraploid corn of pollen in the case of aneuploidy chromosome first.This is near
The character of allotetraploid corn is gradually intended to diploid corn, and pollen fertility is high, can be many by tetraploid as bridge
Give birth to desirable genes present in the Wild related germplasm such as class corn or friction standing grain year to be transferred in cultivated maize, to widen corn germplasm
Basis provides good instrument, while also providing good material for genetic researches such as corn gene functions.
Brief description of the drawings
Fig. 1 are F in the step of embodiment 1 (2)1The double-colored genomic in situ hybridization microphoto in generation.
Fig. 2 are F in the step of embodiment 1 (2)1The pollen fertility detection microphoto in generation.
Fig. 3 are F in the step of embodiment 1 (3)2For double-colored genomic in situ hybridization microphoto.
Fig. 4 are F in the step of embodiment 1 (3)2Microphoto is detected for pollen fertility.
Fig. 5 are F in the step of embodiment 1 (4)3For double-colored genomic in situ hybridization microphoto.
Fig. 6 are F in the step of embodiment 1 (4)3Microphoto is detected for pollen fertility.
Fig. 7 are F in the step of embodiment 1 (5)4For double-colored genomic in situ hybridization microphoto.
Fig. 8 are F in the step of embodiment 1 (5)4Microphoto is detected for pollen fertility.
Embodiment
Below by way of specific embodiment, the present invention will be further explained, but not to the scope of the present invention
Constitute any limitation.
Embodiment 1：The cultivation of nearly allotetraploid corn material of the invention
(1), in April, 2011, the maize planting self-mating system Mo17 and corn allohexaploid MTF-1 on Wenjiang farm
(Tripsazea creammaize T., 2n=76；The public can obtain MTF-1 biomaterial from Sichuan Agricultural University)；2011
In June in year, using corn allohexaploid MTF-1 as female parent, hybridized by male parent of Mo17, seed is harvested when ripe, obtain F1
In generation, it is stored in -20 DEG C of refrigerators；
(2), in April, 2012, F is taken out from -20 DEG C of refrigerators1For seed, F is planted on Sichuan Agricultural University Wenjiang farm1
Generation；In mid-May, 2012 to mid-June takes F1The tip of a root, with genomic in situ hybridization method ((referring to .Crop such as Tang Q
Sci,2005,45:Chromosome number 717-721)) is carried out with genome into identification, F is as a result found1For the chromosome of material
Sum is 35-86 bars, wherein containing maize chromosome number (hereinafter referred to as：Mz) it is 12-30 bars, tetraperennial teosinte
Chromosome number (hereinafter referred to as Zp) is 8-20 bars, and finger-like friction standing grain chromosome number (hereinafter referred to as Tr) is 15-36 bars, and not
With generation part transposition between species chromosome.In June, 2012, (with reference to Su Yuegui, sichuan agriculture was big with iodine-potassium iodide decoration method
Learn Master's thesis, 2009) to F1Pollen fertility detection is carried out for material, it is found that numbering is F1- 31 material pollen has fertility, its
Fertility is 37.5% (see Fig. 2)；F1The chromosome number of -31 materials is 54, wherein being 28 containing Mz, Zp is 11, Tr
For 15 (see Fig. 1)；By to F1- 31 Observation on Agronomic Characters, finds F1- 31 plant are annual, and plant tillering number is 3,
And seed fraction is coated with by glume, threshing is difficult；Due to F1- 31 have fertility, so carrying out selfing to it, are harvested when ripe, obtain F2
Generation；
(3), in April, 2013, F is planted on Sichuan Agricultural University Wenjiang farm2Generation, in May, 2013 to June takes tip of a root base
Because group in-situ hybridization method carry out chromosome number and genome into identify that selective staining body number is 41-43 bars, wherein
Mz is 25-26 bars, and Zp is 14-15 bars, and Tr is 1-2 bars (see Fig. 3, total chromosome number is 43, wherein 2Tr+15Zp+26Mz),
Tiller number is 1-3, and pollen fertility is the plant selfing more than 40% (see Fig. 4), 9 plants of selfing altogether, to F2Seen for economical character
Examine, find F2Middle most of plant shows as cultivated maize type, and no tiller, stem are sturdy, F2In each economical character also in madness
Separation phase, most of seed is still coated with by glume, but there have Some seeds to have begun to be exposed, and seed shatter is difficult；In
In late August, 2013 to early September harvests seed, obtains F3Generation.
(4), in April, 2014, in Sichuan Agricultural University Wenjiang plantation F3For material, in May, 2014 to June takes F3For material
The tip of a root, according to genomic in situ hybridization method described in step (2) to F3The identification of chromosome number and composition is carried out for material；
Pollen fertility is detected with iodine-potassium iodide decoration method in tasseling stage；Selective staining body number is 39-40 bars, wherein containing
Mz is 24-28 bars, and Zp is 12-15 bars, and Tr is 0-2 bars, (see Fig. 5, total chromosome number is 39, including Mz24 bars and
Zp15 bars), no tiller, pollen fertility is the plant selfing more than 50% (see Fig. 6), altogether 70 plants of selfing；By to F3Material
Observation on Agronomic Characters, finds F3Plant majority is cultivated maize type, and stem is very thin, without branch, and the seed of most of fruit ear is exposed,
Easy threshing, small part seed is coated with by glume, and threshing is difficult；Seed is harvested in late August, 2014 to early September, is obtained
F4Generation.
(5), in April, 2015, in Sichuan Agricultural University Wenjiang plantation F4For material, in May, 2015 to June takes F4For material
The tip of a root, according to the genomic in situ hybridization method described in step (2) to F4Chromosome number and genome are carried out for material
Into identification；Select F4Total chromosome number is 38-40 bars in material, wherein containing Mz 22-28 bars, Zp 12-15 bars, Tr 0-2
Bar (see Fig. 7, total chromosome number 40, including Mz24 bars and Zp15 bars)；Without tiller, pollen fertility be more than 70% (see
Plant Fig. 8), is the nearly allotetraploid corn material of the present invention；Will selected 136 plants of selfings；To F4The economical character of material
Observation, finds F4Materials most plant shows as cultivated maize type, only a few plant tillering, Ji Husuo very thin without branch, stem
There is seed completely exposed, Grain Morphology also increasingly tends to conventional corn, and threshing is relatively easy to, under in Augusts, 2015
Ten days to early September harvests seed, retains the seed of harvest.
The application of the nearly allotetraploid corn material of the present invention of embodiment 2
In October, (1) 2016, nearly heterologous the four of the gained of embodiment 1 are planted in Sichuan Agricultural University's In Xishuangbanna of Yunnan base
Times 17 plants of body corn, while planting autotetraploid corn V182.
In December, (2) 2016, the nearly allotetraploid corn using in step described in (1) is miscellaneous by male parent of V182 as female parent
Hand over pollination；Obtain cenospecies within 2 months 2017；Gained cenospecies Grain Morphology is big and full.
In March, (3) 2017,20, the cenospecies seed obtained in Sichuan Agricultural University's Wenjiang plantation step (2) is survived
18 plants, survival rate is 90%；Gained cross materialses stalk is sturdy, blade edge and it is loose, growth potential is (near different better than its parent material
Source tetraploid corn or autotetraploid corn V182), with obvious hybrid vigour.
In June, (4) 2017, cenospecies pollen fertility is detected with iodine-potassium iodide decoration method method in tasseling stage, tied
Fruit shows that its pollen has fertility, therefore, it can carry out genetic transmission.
The above results illustrate that the nearly allotetraploid corn female of aneuploidy of the present invention is fertile, and maternal and jade is used as using it
Rice hybridization, can produce fertile filial generation, thus, the nearly allotetraploid corn material of the present invention can be used as Wild related germplasm
In excellent genes be transferred to bridge material in corn.
The application of the nearly allotetraploid corn material of the present invention of embodiment 3
In October, (1) 2016, nearly heterologous the four of the gained of embodiment 1 are planted in Sichuan Agricultural University's In Xishuangbanna of Yunnan base
Times 14 plants of body corn material, while maize planting self-mating system Mo17.
In December, (2) 2016, using Mo17 as female parent, the nearly allotetraploid corn using in step described in (1) is miscellaneous as male parent
Hand over pollination；Obtain cenospecies within 2 months 2017；Gained cenospecies Grain Morphology is big and full.
In March, (3) 2017,48, the cenospecies seed obtained in Sichuan Agricultural University's Wenjiang plantation step (2) is survived
23 plants, survival rate is 48%；Gained cross materialses stalk is sturdy, blade edge and it is loose, growth potential is (near different better than its parent material
Source tetraploid corn or corn inbred line Mo17), with obvious hybrid vigour.In June, 2017, in tasseling stage iodate
Potassium decoration method method detects that its pollen of testing result has fertility to filial generation pollen fertility.
These results suggest that the nearly allotetraploid corn material of aneuploidy of the present invention has fertility, produces normal flower
Powder, gives corn material to pollinate, and can produce filial generation with it as male parent, therefore, can be as will be excellent in Wild related germplasm
Well-founded is because of the bridge material being transferred in corn.
Claims (3)
1. a kind of method for cultivating nearly allotetraploid corn new material, it is characterised in that comprise the following steps：
(1) it is female parent with MTF-1 (Tripsazea creammaize T., 2n=76), with corn (Zea, in the spinning loose powder phase
Mays L., 2n=20) hybridized for male parent, seed is harvested when ripe, F is obtained1Generation；
(2) F, is planted1Generation, in tasseling stage with iodine-potassium iodide decoration method to F1Pollen fertility detection is carried out for material；Select pollen
The plant for having fertility carries out selfing, and seed is harvested when ripe, obtains F2Generation；
(3) F, is planted2Generation, to the desirable tip of a root before tasseling stage since tri-leaf period, using the method for genomic in situ hybridization to root
Sharp chromosome number and genome are into being identified, and to F2Investigated for the tiller number of plant；In the iodo- iodine of tasseling stage
Change potassium decoration method to detect pollen fertility；It is 41~43 to select root tip chromosomes sum, wherein containing corn (Zea
Mays L.) chromosome number 25~26, tetraperennial teosinte (Zea perennis L.) chromosome number 14~15,
Finger-like friction standing grain (Tripsacum dactyloides L.) chromosome number 1~2, tiller number are 1~3, and pollen fertility
Plant more than 40% carries out selfing, and seed is harvested when ripe, obtains F3Generation；
(4) F, is planted3Generation, according to the genomic in situ hybridization method described in step (3) to root tip chromosomes number and genome into
Identified, and pollen fertility is detected with iodine-potassium iodide decoration method in tasseling stage；Selective staining body sum is 39~40
Bar, wherein containing maize chromosome number 24~28, tetraperennial teosinte chromosome number 12~15, finger-like friction standing grain
The plant that chromosome number 0~2, no tiller and pollen fertility are more than 50% carries out selfing, and seed is harvested when ripe, obtains F4
Generation；
(5) F, is planted4Generation, according to step (3) the genomic in situ hybridization method to root tip chromosomes number and genome into entering
Row identification, and pollen fertility is detected with iodine-potassium iodide decoration method in tasseling stage；Total chromosome number is 38 in testing result
~40, wherein number containing maize chromosome is 22~28, tetraperennial teosinte chromosome number is 12~15, finger-like
The standing grain chromosome number that rubs is 0~2, the plant of no tiller and pollen fertility more than 70%；This total chromosome number is on 40 left sides
The right side, and rubbed standing grain two or more species chromosome groups containing corn, tetraperennial teosinte or finger-like simultaneously
Plant, is the nearly allotetraploid corn material of the present invention；Selected plant selfing is reserved seed for planting.
2. according to the method described in claim 1, it is characterised in that the corn described in its step (1) refers to self-mating system, hybridization
Kind, Local variety or collective species.
3. according to the method described in claim 1, it is characterised in that the nearly allotetraploid corn material selected, for assembling
Cross materialses are used as middle bridge material.
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Cited By (1)
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| CN110972930A (en) * | 2019-11-22 | 2020-04-10 | 四川农业大学 | Method for cultivating high-fertility heterotetraploid corn |
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN110972930A (en) * | 2019-11-22 | 2020-04-10 | 四川农业大学 | Method for cultivating high-fertility heterotetraploid corn |
| CN110972930B (en) * | 2019-11-22 | 2022-04-22 | 四川农业大学 | Method for cultivating high-fertility heterotetraploid corn |
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