CN106561457A - Heterophylla hybrid seed artificial cultivating and breeding method - Google Patents
Heterophylla hybrid seed artificial cultivating and breeding method Download PDFInfo
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- CN106561457A CN106561457A CN201610972933.6A CN201610972933A CN106561457A CN 106561457 A CN106561457 A CN 106561457A CN 201610972933 A CN201610972933 A CN 201610972933A CN 106561457 A CN106561457 A CN 106561457A
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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
- A01H4/00—Plant reproduction by tissue culture techniques ; Tissue culture techniques therefor
- A01H4/008—Methods for regeneration to complete plants
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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
- A01H1/04—Processes of selection involving genotypic or phenotypic markers; Methods of using phenotypic markers for selection
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
- C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
- C12Q1/68—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving nucleic acids
- C12Q1/6876—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes
- C12Q1/6888—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes for detection or identification of organisms
- C12Q1/6895—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes for detection or identification of organisms for plants, fungi or algae
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
- C12Q2600/00—Oligonucleotides characterized by their use
- C12Q2600/13—Plant traits
Abstract
The invention provides a heterophylla hybrid seed artificial cultivating and breeding method. The method comprises the following specific steps of seed selection, seed disinfection, embryo stripping, embryo cultivating, proliferation and amplification, hybrid seed tissue culture seedling screening, hybrid seed excellent plant screening, hybrid seed excellent strain hereditary stability detection and hybrid seed excellent strain propagation expanding and transplanting. The method has the beneficial effects of being small in workload, high in speed, short in period, low in energy consumption and the like. The problems that the heterophylla hybrid seeds are low in germination rate, likely to suffer from insect attack, long in growth cycle, small in propagation coefficient, low in propagation expanding speed, large in breeding workload and long in breeding period and consuming labor and time are solved, and heterophylla hybrid breeding work development is promoted.
Description
Technical field
The invention belongs to medicinal plant breeding field, and in particular to a kind of Radix Pseudostellariae hybrid seed artificial culture selection-breeding side
Method.
Background technology
Radix Pseudostellariae is perennial medicinal plants, is used as medicine with tuber, is traditional strengthening by means of tonics Chinese crude drug, strong with QI invigorating
The effect of spleen, promoting the production of body fluid for nourishing the lung.Radix Pseudostellariae is mainly tuber breeding and seminal propagation, and the multiplex tuber of artificial culture carries out asexual numerous
Grow, make the accumulation of Radix Pseudostellariae disease, germplasm degenerate.Radix Pseudostellariae seminal propagation can cultivate virus-free seedling, can effectively prevent germplasm from moving back
Change.
Hybridization is that the merit of two or more kinds is concentrated in together by copulation, then through selecting and cultivating,
The method for obtaining new varieties, is breeding important method, and the new varieties of selection-breeding generally embody hybrid vigor.After hybridization pollination, harvest
Hybrid seed, generally carry out sowing in experimental plot, rudiment, transplanting, by field selection-breeding for many years, observe its growth characteristics and
Morphological characteristic, screens excellent hybridization individual plant, is enlarged breeding, forms strain, examines or check its hereditary stability and economical character,
Selection-breeding forms hybrid new breed.The Breeding Process of Hybrid is and its important, and very very long, it usually needs 10 years left
The right time.
Radix Pseudostellariae hybridization pollination was carried out in 3-4 months, fruit maturation after generally pollinating month, harvested seed.Seed enters
Row conventional seed planting, it is necessary to through the low temperature induction in winter, Second Year spring could sprout, and a seed is sprouted to form single plant
Strain, through the growth of spring and summer, in July, above-ground plant parts just turns to be yellow withered, and a little tuber is usually formed in underground,
Little tuber is planted in the fall as seed ginseng, further expanding propagation, through cultivation expanding propagation for many years, could form certain amount list
The plant of one seed culture, harvests a number of tuber, forms strain, into control cultivation, carries out individual plant selection, detects which
Hereditary stability and economical character, determine if possess excellent hybrid performance, carry out breeding of new variety.Deposit in prior art
In following shortcoming：（1）Radix Pseudostellariae hybrid seed carries out conventional seed planting, there is dormancy, through the low temperature induction in winter,
2 year spring could sprout, and the time is long, not manageability, and germination rate is low；（2）Radix Pseudostellariae hybrid seed is less, conventional seed planting, easily
Lose, also easily receive insect pest, there is invalid sowing situation；（3）Hybrid seed is sprouted and only grows into single plant, harvests single fritter
Root, breeding coefficient are low, will could form the strain of the single seed culture of certain amount, expanding propagation speed through cultivation expanding propagation for many years
Degree is slow, takes；（4）Radix Pseudostellariae hybrid seed individual plant offspring carries out conventional herd breeding, and workload is big, speed is slow, cycle length, time-consuming.
At present, the report having no in terms of Radix Pseudostellariae cross-breeding, hybrid seed selection-breeding.
The content of the invention
In order to solve hybrid seed germination rate present in existing Radix Pseudostellariae good variety selection process it is low, easily by insect pest,
Growth cycle length, breeding coefficient are low, reproductive speed is slow, selection-breeding workload big, the problem that selection-breeding cycle length, work consuming are time-consuming, improve
Hybrid seed Seedling breakneck acceleration and accuracy, shorten the cross-breeding cycle, save Hybrids Breeding cost, improve Radix Pseudostellariae hybridization and educate
Plant efficiency.The present invention provides a kind of Radix Pseudostellariae hybrid seed artificial culture selection.
To achieve these goals, the present invention is adopted the following technical scheme that：
（1）Seed is selected：Shrivelled hybrid seed is removed using wet concentration method, manually chooses the little seed of granule；
（2）Seed disinfection：The conventional disinfection method of appliable plant tissue culture technology carries out disinfecting for hybrid seed, and sterilized water is clear
It is after washing, standby on clean work station；
（3）Embryo is stripped：In gnotobasiss, the episperm of hybrid seed, endotesta, perisperm are carefully divested, is stayed complete
Embryo；
（4）Seed embryo culture：The embryonic breeding of hybrid seed is entered into light culture 7-10 days in embryo germination inducing culture, is then proceeded to weak
Optical culture, induction embryo are sprouted rapidly, and differentiate Multiple Buds；
（5）Propagation amplification：Multiple Buds were taken out after 30-60 days by embryo differentiation culture, carried out plant division, were accessed concentration of NAA and were
In the MS growth mediums of 0.2mg/L, promote bud to grow, form plant；Growth 50-70 days, single stipes of clip plant connects
Material, accesses same medium, and further expanding propagation growth forms strain；
（6）The screening of hybrid seed tissue cultured seedling：
(a) preliminary screening：Each hybrid seed strain tissue cultured seedling is compared with its male parent tissue cultured seedling, the stem and leaf life of maternal tissue cultured seedling
Long speed, the growth characteristics of robust plant degree and including shapes such as blade shape, thickness, color, stalk thickness, internode length
State feature, tentatively judges the hybrid performance that hybrid seed tissue cultured seedling has, and distinguishes hybrid tissue cultured seedling and non-hybrid tissue cultured seedling；
B () molecule is screened：Using DNA molecular marker technology, further comparison and detection analyzes each hybrid seed tissue cultured seedling, male parent group
The DNA molecular multiformity of seedlings cultivating, maternal tissue cultured seedling, accurately judges the hybrid performance of hybrid seed tissue cultured seedling, retains hybrid tissue culture
Seedling, goes unless hybrid tissue cultured seedling；
（7）The screening of hybrid elite plant strain；Select fast growth, robust plant, microlith root formed speed block, quantity it is many,
The big hybrid strain of microlith root；
（8）The hereditary stability detection of hybrid elite plant strain：By the hybrid elite plant strain tissue cultured seedling of screening, in concentration of NAA it is
Continuous 3 subcultures expanding propagation culture in the MS growth mediums of 0.2mg/L, each incubation time 30-60 days detect each subculture
The tissue cultured seedling growth concordance of expanding propagation culture and hereditary stability, the hybrid elite plant strain for selecting inheritance stability consistent with growth；
（9）Hybrid elite plant strain expanding propagation and transplanting：To the further expanding propagation of hybrid elite plant strain tissue cultured seedling for having screened, then transplant
Field production, selection-breeding new varieties.
The invention has the advantages that：（1）By the embryo for stripping Radix Pseudostellariae hybrid seed, artificial aseptic in-vitro inducing is carried out
Culture, releases dormancy characteristics of different, and the embryo of seed is sprouted immediately, germination rate nearly 100%；（2）Embryo is in inductive differentiation medium
In, it is rapid to sprout, and Multiple Buds are differentiated, the growth coefficient of hybridization embryo is greatly improved, expanding propagation growth is further organized, it is single miscellaneous
Hand over the embryo of seed go out a large amount of plant by expanding propagation rapidly, quickly form independent strain；（3）By comparing each cenospecies subgroup
Seedlings cultivating and father, the growth characteristics of maternal tissue cultured seedling and morphological characteristic, can tentatively judge the hybrid performance that hybrid seed has；Should
Use DNA molecular marker technology, relative analyses hybrid seed tissue cultured seedling, male parent tissue cultured seedling, the DNA molecular multiformity of maternal tissue cultured seedling
Analysis, further determines that the hybrid performance of hybrid seed tissue cultured seedling, goes unless hybrid strain system tissue cultured seedling, hybrid breakneck acceleration block,
Accuracy is high；（4）By compareing each hybrid strain tissue cultured seedling of culture, male parent tissue cultured seedling, maternal tissue cultured seedling, more each tissue culture is observed
Seedling plant strain growth speed, robustness, microlith root form speed, quantity, size, and detect its hereditary stability, selection-breeding hybrid
Elite plant strain, superior hybrid selection-breeding speed block, accuracy are high；（5）To the further expanding propagation of hybrid elite plant strain for having screened, then
Transplant field production, selection-breeding new varieties；Conventional hybrid selection is compared, this method workload is little, speed block, cycle is short, energy
Consumption is few.
Specific embodiment
Below in conjunction with specific embodiment, the present invention will be further described.
Embodiment 1
A kind of Radix Pseudostellariae hybrid seed artificial culture selection, which concretely comprises the following steps：
（1）Seed is selected：Shrivelled hybrid seed is removed using wet concentration method, it is artificial to reject the little seed of granule, only reservation
Hybrid seed more than grain is of medium size and medium；
（2）Seed disinfection：The conventional disinfection method of appliable plant tissue culture technology carries out disinfecting for hybrid seed, and sterilized water is clear
It is after washing, standby on clean work station；
（3）Embryo is stripped：In gnotobasiss, the episperm of hybrid seed, endotesta, perisperm are carefully divested, is stayed complete
Embryo；
（4）Seed embryo culture：The embryonic breeding of hybrid seed is entered into light culture 7 days in embryo germination inducing culture, the low light level is then proceeded to
Culture, induction embryo are sprouted rapidly, and differentiate Multiple Buds.
（5）Propagation amplification：After embryo differentiation culture 50 days, Multiple Buds are taken out, carry out plant division, accessing concentration of NAA is
In the MS growth mediums of 0.2mg/L, promote bud to grow, form plant；Growth 50 days, single stipes of clip plant connects material, connects
Enter same medium, further expanding propagation growth forms strain.
（6）The screening of hybrid seed tissue cultured seedling：
a）Preliminary screening：Each hybrid seed strain tissue cultured seedling is compared with its male parent tissue cultured seedling, the stem and leaf life of maternal tissue cultured seedling
Long speed, the growth characteristics of robust plant degree and including shapes such as blade shape, thickness, color, stalk thickness, internode length
State feature, tentatively judges the hybrid performance that hybrid seed tissue cultured seedling has, and distinguishes hybrid tissue cultured seedling and non-hybrid tissue cultured seedling；
b）Molecule is screened：Using DNA molecular marker technology, further comparison and detection analyzes each hybrid seed tissue cultured seedling, male parent group
The DNA molecular multiformity of seedlings cultivating, maternal tissue cultured seedling, accurately judges the hybrid performance of hybrid seed tissue cultured seedling, retains hybrid tissue culture
Seedling, goes unless hybrid tissue cultured seedling.
（7）The screening of hybrid elite plant strain；Select fast growth, robust plant, microlith root and form speed block, quantity
The hybrid strain many, microlith root is big；
（8）The hereditary stability detection of hybrid elite plant strain：By the hybrid elite plant strain tissue cultured seedling of screening, in concentration of NAA it is
Continuous 3 subcultures expanding propagation culture in the MS growth mediums of 0.2mg/L, each incubation time 30 days detect each subculture expanding propagation
The tissue cultured seedling growth concordance of culture and hereditary stability, the hybrid elite plant strain for selecting inheritance stability consistent with growth.
（9）Hybrid elite plant strain expanding propagation and transplanting：To the further expanding propagation of hybrid elite plant strain tissue cultured seedling for having screened, then
Transplant field production, selection-breeding new varieties.
Embodiment 2
A kind of Radix Pseudostellariae hybrid seed artificial culture selection, which concretely comprises the following steps：
（1）Seed is selected：Shrivelled hybrid seed is removed using wet concentration method, it is artificial to reject the little seed of granule, only reservation
Hybrid seed more than grain is of medium size and medium；
（2）Seed disinfection：The conventional disinfection method of appliable plant tissue culture technology carries out disinfecting for hybrid seed, and sterilized water is clear
It is after washing, standby on clean work station；
（3）Embryo is stripped：In gnotobasiss, the episperm of hybrid seed, endotesta, perisperm are carefully divested, is stayed complete
Embryo；
（4）Seed embryo culture：The embryonic breeding of hybrid seed is entered into light culture 8 days in embryo germination inducing culture, the low light level is then proceeded to
Culture, induction embryo are sprouted rapidly, and differentiate Multiple Buds.
（5）Propagation amplification：After embryo differentiation culture 30 days, Multiple Buds are taken out, carry out plant division, accessing concentration of NAA is
In the MS growth mediums of 0.2mg/L, promote bud to grow, form plant；Growth 60 days, single stipes of clip plant connects material, connects
Enter same medium, further expanding propagation growth forms strain.
（6）The screening of hybrid seed tissue cultured seedling：
a）Preliminary screening：Each hybrid seed strain tissue cultured seedling is compared with its male parent tissue cultured seedling, the stem and leaf life of maternal tissue cultured seedling
Long speed, the growth characteristics of robust plant degree and including shapes such as blade shape, thickness, color, stalk thickness, internode length
State feature, tentatively judges the hybrid performance that hybrid seed tissue cultured seedling has, and distinguishes hybrid tissue cultured seedling and non-hybrid tissue cultured seedling；
b）Molecule is screened：Using DNA molecular marker technology, further comparison and detection analyzes each hybrid seed tissue cultured seedling, male parent group
The DNA molecular multiformity of seedlings cultivating, maternal tissue cultured seedling, accurately judges the hybrid performance of hybrid seed tissue cultured seedling, retains hybrid tissue culture
Seedling, goes unless hybrid tissue cultured seedling.
（7）The screening of hybrid elite plant strain；Select fast growth, robust plant, microlith root and form speed block, quantity
The hybrid strain many, microlith root is big；
（8）The hereditary stability detection of hybrid elite plant strain：By the hybrid elite plant strain tissue cultured seedling of screening, in concentration of NAA it is
Continuous 3 subcultures expanding propagation culture in the MS growth mediums of 0.2mg/L, each incubation time 60 days detect each subculture expanding propagation
The tissue cultured seedling growth concordance of culture and hereditary stability, the hybrid elite plant strain for selecting inheritance stability consistent with growth.
（9）Hybrid elite plant strain expanding propagation and transplanting：To the further expanding propagation of hybrid elite plant strain tissue cultured seedling for having screened, then
Transplant field production, selection-breeding new varieties.
Embodiment 3
A kind of Radix Pseudostellariae hybrid seed artificial culture selection, which concretely comprises the following steps：
（1）Seed is selected：Shrivelled hybrid seed is removed using wet concentration method, it is artificial to reject the little seed of granule, only reservation
Hybrid seed more than grain is of medium size and medium；
（2）Seed disinfection：The conventional disinfection method of appliable plant tissue culture technology carries out disinfecting for hybrid seed, and sterilized water is clear
It is after washing, standby on clean work station；
（3）Embryo is stripped：In gnotobasiss, the episperm of hybrid seed, endotesta, perisperm are carefully divested, is stayed complete
Embryo；
（4）Seed embryo culture：The embryonic breeding of hybrid seed is entered into light culture 10 days in embryo germination inducing culture, the low light level is then proceeded to
Culture, induction embryo are sprouted rapidly, and differentiate Multiple Buds.
（5）Propagation amplification：After embryo differentiation culture 60 days, Multiple Buds are taken out, carry out plant division, accessing concentration of NAA is
In the MS growth mediums of 0.2mg/L, promote bud to grow, form plant；Growth 70 days, single stipes of clip plant connects material, connects
Enter same medium, further expanding propagation growth forms strain.
（6）The screening of hybrid seed tissue cultured seedling：
a）Preliminary screening：Each hybrid seed strain tissue cultured seedling is compared with its male parent tissue cultured seedling, the stem and leaf life of maternal tissue cultured seedling
Long speed, the growth characteristics of robust plant degree and including shapes such as blade shape, thickness, color, stalk thickness, internode length
State feature, tentatively judges the hybrid performance that hybrid seed tissue cultured seedling has, and distinguishes hybrid tissue cultured seedling and non-hybrid tissue cultured seedling；
b）Molecule is screened：Using DNA molecular marker technology, further comparison and detection analyzes each hybrid seed tissue cultured seedling, male parent group
The DNA molecular multiformity of seedlings cultivating, maternal tissue cultured seedling, accurately judges the hybrid performance of hybrid seed tissue cultured seedling, retains hybrid tissue culture
Seedling, goes unless hybrid tissue cultured seedling.
（7）The screening of hybrid elite plant strain；Select fast growth, robust plant, microlith root and form speed block, quantity
The hybrid strain many, microlith root is big；
（8）The hereditary stability detection of hybrid elite plant strain：By the hybrid elite plant strain tissue cultured seedling of screening, in concentration of NAA it is
Continuous 3 subcultures expanding propagation culture in the MS growth mediums of 0.2mg/L, each incubation time 50 days detect each subculture expanding propagation
The tissue cultured seedling growth concordance of culture and hereditary stability, the hybrid elite plant strain for selecting inheritance stability consistent with growth.
（9）Hybrid elite plant strain expanding propagation and transplanting：To the further expanding propagation of hybrid elite plant strain tissue cultured seedling for having screened, then
Transplant field production, selection-breeding new varieties.
Claims (1)
1. a kind of Radix Pseudostellariae hybrid seed artificial culture selection, it is characterised in that comprising following concrete steps：
（1）Seed is selected：Shrivelled hybrid seed is removed using wet concentration method, manually chooses the little seed of granule；
（2）Seed disinfection：The conventional disinfection method of appliable plant tissue culture technology carries out disinfecting for hybrid seed, and sterilized water is clear
It is after washing, standby on clean work station；
（3）Embryo is stripped：In gnotobasiss, the episperm of hybrid seed, endotesta, perisperm are carefully divested, is stayed complete
Embryo；
（4）Seed embryo culture：The embryonic breeding of hybrid seed is entered into light culture 7-10 days in embryo germination inducing culture, is then proceeded to weak
Optical culture, induction embryo are sprouted rapidly, and differentiate Multiple Buds；
（5）Propagation amplification：Multiple Buds were taken out after 30-60 days by embryo differentiation culture, carried out plant division, were accessed concentration of NAA and were
In the MS growth mediums of 0.2mg/L, promote bud to grow, form plant；Growth 50-70 days, single stipes of clip plant connects
Material, accesses same medium, and further expanding propagation growth forms strain；
（6）The screening of hybrid seed tissue cultured seedling：
(a) preliminary screening：Each hybrid seed strain tissue cultured seedling is compared with its male parent tissue cultured seedling, the stem and leaf life of maternal tissue cultured seedling
Long speed, the growth characteristics of robust plant degree and including shapes such as blade shape, thickness, color, stalk thickness, internode length
State feature, tentatively judges the hybrid performance that hybrid seed tissue cultured seedling has, and distinguishes hybrid tissue cultured seedling and non-hybrid tissue cultured seedling；
B () molecule is screened：Using DNA molecular marker technology, further comparison and detection analyzes each hybrid seed tissue cultured seedling, male parent group
The DNA molecular multiformity of seedlings cultivating, maternal tissue cultured seedling, accurately judges the hybrid performance of hybrid seed tissue cultured seedling, retains hybrid tissue culture
Seedling, goes unless hybrid tissue cultured seedling；
（7）The screening of hybrid elite plant strain；Select fast growth, robust plant, microlith root formed speed block, quantity it is many,
The big hybrid strain of microlith root；
（8）The hereditary stability detection of hybrid elite plant strain：By the hybrid elite plant strain tissue cultured seedling of screening, in concentration of NAA it is
Continuous 3 subcultures expanding propagation culture in the MS growth mediums of 0.2mg/L, each incubation time 30-60 days detect each subculture
The tissue cultured seedling growth concordance of expanding propagation culture and hereditary stability, the hybrid elite plant strain for selecting inheritance stability consistent with growth；
（9）Hybrid elite plant strain expanding propagation and transplanting：To the further expanding propagation of hybrid elite plant strain tissue cultured seedling for having screened, then transplant
Field production, selection-breeding new varieties.
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| CN109618933A (en) * | 2019-01-23 | 2019-04-16 | 贵州大学 | A kind of radix pseudostellariae Seed embryo culture base and its cultural method |
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| CN109618933A (en) * | 2019-01-23 | 2019-04-16 | 贵州大学 | A kind of radix pseudostellariae Seed embryo culture base and its cultural method |
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| CN106561457B (en) | 2019-03-15 |
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