CN101613755A - Identify the method for carnation chromosome number with bud - Google Patents
Identify the method for carnation chromosome number with bud Download PDFInfo
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- CN101613755A CN101613755A CN200910094743A CN200910094743A CN101613755A CN 101613755 A CN101613755 A CN 101613755A CN 200910094743 A CN200910094743 A CN 200910094743A CN 200910094743 A CN200910094743 A CN 200910094743A CN 101613755 A CN101613755 A CN 101613755A
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Abstract
The invention provides and a kind ofly identify the method for carnation chromosome number, comprise that flower pesticide, ovary wall and corolla tissue, mixture of ice and water pre-treatment, the stationary liquid of getting bud are fixed, dyeing, salt acid dissociation, compressing tablet and microscopy with bud.Carry out chromosome sectioning with organization of root tips and compare with traditional, solved in the root tip chromosomes film-making, few, the difficult microscopic of visual field cell, karyomit(e) disperses bad shortcoming, and it is long to have overcome Dianthus caryophyllus L. cuttage seeding cycle of taking root simultaneously, and the problem that wastes time and energy of cuttage; Compare with the stem tip chromosome film-making, overcome the interference problem of non-meristematic cell or repertory or secretory product, only need get bud can carry out chromosome sectioning, can not destroy the growth conditions of plant, draw materials conveniently, and metacinesis phase cell is many, it is little to have solved carnation chromosome, quantity is many, the difficult problem that karyomit(e) is difficult to count.Therefore, when plant blossom, utilizing bud to carry out chromosome sectioning is efficient ways.
Description
Technical field
The present invention relates to a kind of method of identifying carnation chromosome number, especially a kind of method with bud evaluation carnation chromosome number belongs to cytobiology and Cytogenetic techniques field.
Background technology
Dianthus caryophyllus L. (Dianthus caryophyllus Linn) has another name called carnation, for the perennial perennial root herbaceous plant of Caryophyllaceae Carnation, is one of world-renowned fresh cutting flower, also is one of topmost large outlet flowers in Yunnan Province.In the Dianthus caryophyllus L. cross-breeding, exist and hybridize not affinity, the phenomenon that setting percentage is on the low side.One of them reason is unclear to the cytology background of a lot of germplasms.Therefore, success ratio, the raising cross-breeding efficient of hybridization be increased, carnation chromosome ploidy analysis problem should be solved earlier.
Karyomit(e) is the carrier of gene, and the preparation chromosome specimen is undoubtedly the most basic technology of cytogenetics, and good chromosome sectioning is the prerequisite of carrying out chromosome banding, group type analysis, in situ hybridization etc.The observation of plant cell chromosome and analysis, significant for the research of genetics phenomenon such as cytometaplasia in the action of chromosome karyotype analysis, cytogamy after stain colour solid and the culturing process.
The chromosome length of Dianthus caryophyllus L. is 2.5-4.0um, belongs to the microchromosome type, has normal kinetochore, because carnation chromosome number is many, and belongs to microchromosome, the compressing tablet difficulty is bigger, therefore needs to seek the high material of cell division index and carries out chromosome counting.The tip of a root is the most frequently used material of chromosome counting, and especially the seed root-tip cells is the most reliable material of research karyomit(e).The Dianthus caryophyllus L. setting percentage is low, most plants shaky and by nourishing and generating, but cuttage seeding is taken root the cycle long, and cuttage wastes time and energy, and therefore carries out chromosome sectioning if get the tip of a root with the slotting seedling rooting of fibre, then need spend longer for some time, the difficulty of drawing materials is big.When getting stem apex in addition and carrying out chromosome sectioning, because stem apex is drawn materials and is subject to the restriction of the season of growth, and the interference of different non-meristematic cell or repertory or secretory product is arranged, make metacinesis phase cell few or do not have, stem apex strips difficulty, thereby influences chromosome sectioning and counting.Therefore, must be improved prior art.
Summary of the invention
The object of the present invention is to provide a kind of drawing materials easily, time saving and energy saving, and the many mutually methods with bud evaluation carnation chromosome number of metaphase.
The present invention finishes by following technical proposal: a kind of method with bud evaluation carnation chromosome number is characterized in that through following process steps:
A gets the bud of the long 1.0-1.9cm of Dianthus caryophyllus L.;
B puts into mixture of ice and water pre-treatment 22-26h with bud;
C after pretreated bud cleaned, immerses the fixing 2-24h of processing in the stationary liquid of following volume ratio: dehydrated alcohol: Glacial acetic acid=3: 1;
D after the bud after fixing the processing cleaned, under room temperature, immerses concentration and is to dissociate in the HCL solution of 1mol/L and handle 10-20min;
E after the bud cleaning of dissociating after handling, places on the slide glass, tears up the bud tissue with scalper, drips a 1-3 and drips the carbol fuchsin dye liquor, dyeing 5-10min;
F, the bud tissue covered after dyeing is handled, thieving paper in the covering touches the bud tissue, and cell is disperseed rapidly;
G with sealing around the cover glass, promptly gets chromosome number with observation by light microscope with nail varnish.
Described bud is specifically got its flower pesticide or ovary wall or corolla tissue.
The cleaning of described C, D, E step adopts distilled water to clean 2-3 time, and implement is a syringe.
The present invention compared with prior art has following advantage and effect: adopt such scheme, promptly carry out chromosome sectioning with immature bud tissue, thereby carnation chromosome number is counted, carry out chromosome sectioning with organization of root tips and compare with traditional, solved in the root tip chromosomes film-making, few, the difficult microscopic of visual field cell, karyomit(e) disperses bad shortcoming, it is long to have overcome Dianthus caryophyllus L. cuttage seeding cycle of taking root simultaneously, and the problem that wastes time and energy of cuttage; Compare with the stem tip chromosome film-making, overcome the interference problem of non-meristematic cell or repertory or secretory product, only need get bud can carry out chromosome sectioning, can not destroy the growth conditions of plant, draw materials conveniently, and metacinesis phase cell is many, it is little to have solved carnation chromosome, quantity is many, the difficult problem that karyomit(e) is difficult to count.Therefore, when plant blossom, utilizing bud to carry out chromosome sectioning is efficient ways.
Embodiment
Below in conjunction with embodiment the present invention is described further.
Embodiment 1
With Dianthus caryophyllus L. kind " autumn harvest " is material, wins the long bud of 1.9cm, strips out ovary, and ovary is vertically cut, and gets its ovary wall and puts into mixture of ice and water pre-treatment 24h; After drawing waste liquid with the 5ml syringe, pretreated ovary wall is cleaned 3 times with distilled water, immerse volume ratio and be: dehydrated alcohol: fixing 2h in the Ka Nuoshi I stationary liquid of Glacial acetic acid=3: 1; Behind identical syringe absorption waste liquid, the ovary wall after fixing the processing is cleaned 3 times with distilled water, under room temperature, immerse 20min in the HCL solution that concentration is 1mol/L, carry out the salt acid dissociation; Behind identical syringe absorption waste liquid, clean ovary wall 3 times with distilled water again, place on the slide glass, tear up the ovary wall tissue with scalper, drip 2 carbol fuchsin dye liquors, dyeing 10min, covered, thieving paper in the covering, knock the ovary wall tissue with chopsticks on the thieving paper cover glass gently covering, cell is disperseed rapidly, around cover glass, carry out sealing with nail varnish, use observation by light microscope immediately, its microscopy effect: under the situation of amplifying 1000 times (10 times of eyepiece * 100 times object lens), mid-term, meristematic cell was many, and karyomit(e) shrinks, the karyomit(e) good dispersion is easy to counting.Dyed body analysis of accounts, Dianthus caryophyllus L. kind " autumn harvest " chromosome number is 30, diploid.
Embodiment 2
With Dianthus caryophyllus L. kind " Huang Fei's letter " is material, wins the long bud of 1.3cm, strips out ovary, and ovary is vertically cut, and gets its ovary wall and puts into mixture of ice and water pre-treatment 22h; Behind the waste liquid after draw handling with the 5ml syringe, ovary wall is cleaned 2 times with distilled water, immerse volume ratio afterwards and be: dehydrated alcohol: fixing 4h in the Ka Nuoshi I stationary liquid of Glacial acetic acid=3: 1; Behind the waste liquid after the identical syringe absorption processing, ovary wall is cleaned 3 times with distilled water, under room temperature, immerse 15min in the HCL solution that concentration is 1mol/L, carry out the salt acid dissociation; Behind the waste liquid after the identical syringe absorption processing, ovary wall is cleaned 3 times with distilled water again, place on the slide glass, tear up the ovary wall tissue with scalper, drip 3 carbol fuchsin dye liquors, dyeing 5min, covered, thieving paper in the covering, knock the ovary wall tissue with chopsticks on the thieving paper cover glass gently covering, cell is disperseed rapidly, around cover glass, carry out sealing with nail varnish, use observation by light microscope after four days, its microscopy effect: under the situation of amplifying 1000 times (10 times of eyepiece * 100 times object lens), mid-term, the meristematic cell number was many, and karyomit(e) shrinks, the karyomit(e) good dispersion is easy to counting.Dyed body analysis of accounts, Dianthus caryophyllus L. kind " Huang Fei's letter " chromosome number is 60, tetraploid.
Embodiment 3
" being crazy about " with the Dianthus caryophyllus L. kind is material, gets the long bud of 1.5cm, and pollen was in monokaryon and kept to the side the phase this moment, took out flower pesticide, put into mixture of ice and water pre-treatment 24h; Behind the waste liquid after draw handling with the 5ml syringe, ovary wall is cleaned 3 times with distilled water, immerse volume ratio and be: dehydrated alcohol: fixing 2h in the Ka Nuoshi I stationary liquid of Glacial acetic acid=3: 1; Behind the waste liquid after the identical syringe absorption processing, ovary wall is cleaned 3 times with distilled water, under room temperature, immerse 20min in the HCL solution that concentration is 1mol/L, carry out the salt acid dissociation; Behind the waste liquid after the identical syringe absorption processing, ovary wall is cleaned 3 times with distilled water, place on the slide glass, tear up anther tissue with scalper, drip 2 carbol fuchsin dye liquors, dyeing 5min, covered, thieving paper in the covering, knock anther tissue with chopsticks on the thieving paper cover glass gently covering, cell is disperseed rapidly, around cover glass, carry out sealing with nail varnish, use observation by light microscope immediately, its microscopy effect: under the situation of amplifying 1000 times (10 times of eyepiece * 100 times object lens), mid-term, meristematic cell was many, and karyomit(e) shrinks, the karyomit(e) good dispersion is easy to counting.It is 30 that dyed body analysis of accounts, Dianthus caryophyllus L. kind " are crazy about " chromosome number, diploid.
Embodiment 4
" Luo Jiate " is material with the Dianthus caryophyllus L. kind, gets the long bud of 1.2cm, and this moment, pollen was in the subtrahend first division diakinesis stage to the subtrahend a, took out flower pesticide, put into mixture of ice and water pre-treatment 22h; Behind the waste liquid after draw handling with the 5ml syringe, ovary wall is cleaned 3 times with distilled water, immerse volume ratio and be: dehydrated alcohol: fixing 4h in the Ka Nuoshi I stationary liquid of Glacial acetic acid=3: 1; Behind the waste liquid after the identical syringe absorption processing, ovary wall is cleaned 2 times with distilled water, under room temperature, immerse 10min in the HCL solution that concentration is 1mol/L, carry out the salt acid dissociation; Behind the waste liquid after the identical syringe absorption processing, ovary wall is cleaned 2 times with distilled water, place on the slide glass, tear up anther tissue with scalper, drip 3 carbol fuchsin dye liquors, dyeing 5min, covered, thieving paper in the covering, knock anther tissue with chopsticks on the thieving paper cover glass gently covering, cell is disperseed rapidly, around cover glass, carry out sealing with nail varnish, use observation by light microscope immediately, its microscopy effect: under the situation of amplifying 1000 times (10 times of eyepiece * 100 times object lens), mid-term, meristematic cell was many, and karyomit(e) shrinks, the karyomit(e) good dispersion is easy to counting.Dyed body analysis of accounts, Dianthus caryophyllus L. kind " Luo Jiate " chromosome number is 30, diploid.
Embodiment 5
With Dianthus caryophyllus L. kind " high mallow titanium alloy " is material, gets the long bud of 1.2cm, takes out corolla, puts into mixture of ice and water pre-treatment 26h; Behind the waste liquid after draw handling with the 5ml syringe, corolla is cleaned 2 times with distilled water, immerse volume ratio and be: dehydrated alcohol: fixing 16h in the Ka Nuoshi I stationary liquid of Glacial acetic acid=3: 1; Behind the waste liquid after the identical syringe absorption processing, corolla is cleaned 2 times with distilled water, under room temperature, immerse 15min in the HCL solution that concentration is 1mol/L, carry out the salt acid dissociation; Behind the waste liquid after the identical syringe absorption processing, corolla is cleaned 3 times with distilled water, place on the slide glass, tear up the corolla tissue with scalper, drip 1 carbol fuchsin dye liquor, dyeing 5min, covered, thieving paper in the covering, knock anther tissue with chopsticks on the thieving paper cover glass gently covering, cell is disperseed rapidly, around cover glass, carry out sealing with nail varnish, used observation by light microscope on the 4th day, its microscopy effect: under the situation of amplifying 1000 times (10 times of eyepiece * 100 times object lens), mid-term, meristematic cell was many, and karyomit(e) shrinks, the karyomit(e) good dispersion is easy to counting.Dyed body analysis of accounts, Dianthus caryophyllus L. kind " high mallow titanium alloy " chromosome number is 30, diploid.
Embodiment 6
" powder beauty " is material with the Dianthus caryophyllus L. kind, wins the long bud of 1.1cm, takes out corolla, puts into mixture of ice and water pre-treatment 26h; Behind the waste liquid after draw handling with the 5ml syringe, corolla is cleaned 2 times with distilled water, immerse volume ratio and be: dehydrated alcohol: fixing 16h in the Ka Nuoshi I stationary liquid of Glacial acetic acid=3: 1; Behind the waste liquid after the identical syringe absorption processing, corolla is cleaned 2 times with distilled water, under room temperature, immerse 20min in the HCL solution that concentration is 1mol/L, carry out the salt acid dissociation; Behind the waste liquid after the identical syringe absorption processing, corolla is cleaned 3 times with distilled water again, place on the slide glass, tear up the corolla tissue with scalper, drip 1 carbol fuchsin dye liquor, dyeing 8min, covered, thieving paper in the covering, knock the corolla tissue with chopsticks on the thieving paper cover glass gently covering, cell is disperseed rapidly, around cover glass, carry out sealing with nail varnish, use observation by light microscope after two days, its microscopy effect: under the situation of amplifying 1000 times (10 times of eyepiece * 100 times object lens), mid-term, meristematic cell was many, and karyomit(e) shrinks, the karyomit(e) good dispersion is easy to counting.Dyed body analysis of accounts, Dianthus caryophyllus L. kind " powder beauty " chromosome number is 60, tetraploid.
Claims (3)
1, a kind of method with bud evaluation carnation chromosome number is characterized in that through following process steps:
A gets the bud of the long 1.0-1.9cm of Dianthus caryophyllus L.;
B puts into mixture of ice and water pre-treatment 22-26h with bud;
C after pretreated bud cleaned, immerses the fixing 2-24h of processing in the stationary liquid of following volume ratio: dehydrated alcohol: Glacial acetic acid=3: 1;
D after the bud after fixing the processing cleaned, under room temperature, immerses concentration and is to dissociate in the HCL solution of 1mol/L and handle 10-20min;
E after the bud cleaning of dissociating after handling, tears up the bud tissue, places on the slide glass dyeing 5-10min;
F, the bud tissue covered after dyeing is handled, thieving paper in the covering touches the bud tissue, and cell is disperseed rapidly;
G with sealing around the cover glass, promptly gets chromosome number with observation by light microscope with nail varnish.
2, as claimed in claim 1ly identify the method for carnation chromosome number, it is characterized in that described bud specifically gets its flower pesticide or ovary wall or corolla tissue with bud.
3, the method with bud evaluation carnation chromosome number as claimed in claim 1 is characterized in that the cleaning of described C, D, E step adopts distilled water to clean 2-3 time, and implement is a syringe.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN 200910094743 CN101613755B (en) | 2009-07-17 | 2009-07-17 | Method for identifying carnation chromosome number by bud |
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| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN 200910094743 CN101613755B (en) | 2009-07-17 | 2009-07-17 | Method for identifying carnation chromosome number by bud |
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| CN101613755A true CN101613755A (en) | 2009-12-30 |
| CN101613755B CN101613755B (en) | 2012-09-05 |
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| CN 200910094743 Expired - Fee Related CN101613755B (en) | 2009-07-17 | 2009-07-17 | Method for identifying carnation chromosome number by bud |
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Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102277423A (en) * | 2011-07-13 | 2011-12-14 | 浙江万里学院 | Quick determination method of number of chromosomes of bivalve by utilizing ovum fluorescent microscope observation |
| CN105838786A (en) * | 2016-03-24 | 2016-08-10 | 西南大学 | Method of chromosome smearing of adult plants of loquat with young tender flower buds |
| CN110012748A (en) * | 2019-04-23 | 2019-07-16 | 南京林业大学 | It polymerize monogynaecial stripping means in gynoecium |
| CN111238888A (en) * | 2020-01-16 | 2020-06-05 | 云南省农业科学院甘蔗研究所 | Efficient sugarcane or sugarcane near-edge seed stem tip chromosome flaking method |
-
2009
- 2009-07-17 CN CN 200910094743 patent/CN101613755B/en not_active Expired - Fee Related
Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102277423A (en) * | 2011-07-13 | 2011-12-14 | 浙江万里学院 | Quick determination method of number of chromosomes of bivalve by utilizing ovum fluorescent microscope observation |
| CN105838786A (en) * | 2016-03-24 | 2016-08-10 | 西南大学 | Method of chromosome smearing of adult plants of loquat with young tender flower buds |
| CN110012748A (en) * | 2019-04-23 | 2019-07-16 | 南京林业大学 | It polymerize monogynaecial stripping means in gynoecium |
| CN110012748B (en) * | 2019-04-23 | 2021-10-01 | 南京林业大学 | Method for stripping single pistil in polymeric pistil |
| CN111238888A (en) * | 2020-01-16 | 2020-06-05 | 云南省农业科学院甘蔗研究所 | Efficient sugarcane or sugarcane near-edge seed stem tip chromosome flaking method |
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| CN101613755B (en) | 2012-09-05 |
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