CN114875072B - Method for creating germplasm of purple perilla - Google Patents
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- CN114875072B CN114875072B CN202210230222.7A CN202210230222A CN114875072B CN 114875072 B CN114875072 B CN 114875072B CN 202210230222 A CN202210230222 A CN 202210230222A CN 114875072 B CN114875072 B CN 114875072B
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- 235000004347 Perilla Nutrition 0.000 title claims abstract description 131
- 238000000034 method Methods 0.000 title claims abstract description 46
- 244000124853 Perilla frutescens Species 0.000 title abstract description 112
- 241000196324 Embryophyta Species 0.000 claims abstract description 30
- 230000017260 vegetative to reproductive phase transition of meristem Effects 0.000 claims abstract description 18
- 238000003306 harvesting Methods 0.000 claims abstract description 13
- 238000012216 screening Methods 0.000 claims abstract description 10
- 241000229722 Perilla <angiosperm> Species 0.000 claims abstract 37
- 235000013399 edible fruits Nutrition 0.000 claims description 18
- 241001573881 Corolla Species 0.000 claims description 14
- 230000009418 agronomic effect Effects 0.000 claims description 13
- 239000007788 liquid Substances 0.000 claims description 13
- 241001164374 Calyx Species 0.000 claims description 10
- 239000003205 fragrance Substances 0.000 claims description 7
- 238000007605 air drying Methods 0.000 claims description 6
- 230000003698 anagen phase Effects 0.000 claims description 6
- 238000007796 conventional method Methods 0.000 claims description 6
- 239000000243 solution Substances 0.000 claims description 6
- 238000002347 injection Methods 0.000 claims description 5
- 239000007924 injection Substances 0.000 claims description 5
- LZZYPRNAOMGNLH-UHFFFAOYSA-M Cetrimonium bromide Chemical compound [Br-].CCCCCCCCCCCCCCCC[N+](C)(C)C LZZYPRNAOMGNLH-UHFFFAOYSA-M 0.000 claims description 4
- 238000000605 extraction Methods 0.000 claims description 4
- 239000012535 impurity Substances 0.000 claims description 4
- 239000000843 powder Substances 0.000 claims description 4
- 238000009331 sowing Methods 0.000 claims description 4
- 238000009395 breeding Methods 0.000 claims description 3
- 230000001488 breeding effect Effects 0.000 claims description 2
- 230000008859 change Effects 0.000 claims description 2
- 238000005259 measurement Methods 0.000 claims 1
- 230000000007 visual effect Effects 0.000 claims 1
- 235000004348 Perilla frutescens Nutrition 0.000 abstract description 10
- 238000009402 cross-breeding Methods 0.000 abstract description 5
- 230000008569 process Effects 0.000 abstract description 5
- 230000035772 mutation Effects 0.000 abstract description 4
- 238000012549 training Methods 0.000 abstract description 2
- DTOSIQBPPRVQHS-PDBXOOCHSA-N alpha-linolenic acid Chemical compound CC\C=C/C\C=C/C\C=C/CCCCCCCC(O)=O DTOSIQBPPRVQHS-PDBXOOCHSA-N 0.000 description 2
- 229920000742 Cotton Polymers 0.000 description 1
- 244000068988 Glycine max Species 0.000 description 1
- 235000010469 Glycine max Nutrition 0.000 description 1
- 241000219146 Gossypium Species 0.000 description 1
- 240000006240 Linum usitatissimum Species 0.000 description 1
- 235000004431 Linum usitatissimum Nutrition 0.000 description 1
- 240000007594 Oryza sativa Species 0.000 description 1
- 235000007164 Oryza sativa Nutrition 0.000 description 1
- 241000209140 Triticum Species 0.000 description 1
- 235000021307 Triticum Nutrition 0.000 description 1
- 235000020661 alpha-linolenic acid Nutrition 0.000 description 1
- 210000004556 brain Anatomy 0.000 description 1
- 238000012512 characterization method Methods 0.000 description 1
- 239000003153 chemical reaction reagent Substances 0.000 description 1
- 238000011161 development Methods 0.000 description 1
- 230000018109 developmental process Effects 0.000 description 1
- 230000000694 effects Effects 0.000 description 1
- 238000002474 experimental method Methods 0.000 description 1
- 244000037666 field crops Species 0.000 description 1
- 238000012214 genetic breeding Methods 0.000 description 1
- 239000004519 grease Substances 0.000 description 1
- 230000012010 growth Effects 0.000 description 1
- 238000009396 hybridization Methods 0.000 description 1
- 229960004488 linolenic acid Drugs 0.000 description 1
- 239000000463 material Substances 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 235000016709 nutrition Nutrition 0.000 description 1
- 230000035764 nutrition Effects 0.000 description 1
- 235000009566 rice Nutrition 0.000 description 1
- 238000011282 treatment Methods 0.000 description 1
- 235000021122 unsaturated fatty acids Nutrition 0.000 description 1
- 150000004670 unsaturated fatty acids Chemical class 0.000 description 1
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- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
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- C12N15/00—Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
- C12N15/09—Recombinant DNA-technology
- C12N15/63—Introduction of foreign genetic material using vectors; Vectors; Use of hosts therefor; Regulation of expression
- C12N15/79—Vectors or expression systems specially adapted for eukaryotic hosts
- C12N15/82—Vectors or expression systems specially adapted for eukaryotic hosts for plant cells, e.g. plant artificial chromosomes (PACs)
- C12N15/8201—Methods for introducing genetic material into plant cells, e.g. DNA, RNA, stable or transient incorporation, tissue culture methods adapted for transformation
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N15/00—Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
- C12N15/09—Recombinant DNA-technology
- C12N15/63—Introduction of foreign genetic material using vectors; Vectors; Use of hosts therefor; Regulation of expression
- C12N15/79—Vectors or expression systems specially adapted for eukaryotic hosts
- C12N15/82—Vectors or expression systems specially adapted for eukaryotic hosts for plant cells, e.g. plant artificial chromosomes (PACs)
- C12N15/8241—Phenotypically and genetically modified plants via recombinant DNA technology
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- Breeding Of Plants And Reproduction By Means Of Culturing (AREA)
Abstract
The invention provides a purple perilla germplasm innovation method, which comprises the following steps: selecting donor perilla and acceptor perilla; harvesting and preserving seeds; cultivating donor perilla and acceptor perilla; extracting donor perilla DNA; selecting a receptor perilla frutescens in flowering phase; creating new germplasm of purple perilla; collecting seeds; and screening variant germplasm. Compared with the prior art, the invention has the following advantages: 1) The method has wide selection of the purple perilla donor, does not have the condition of flowering period incompatibility of the donor and the receptor purple perilla, and greatly reduces the space-time restriction of the donor and the receptor purple perilla; 2) Compared with the traditional conventional crossbreeding, the method eliminates the difficulty of emasculation and reduces the damage to plants in the crossbreeding process; 3) The method has simple operability and reduces the requirements of manual training and proficiency; 4) The method can cause the perilla acceptor to generate mutation in the seed forming process, and different mutation results are the feasibility and applicability of the perilla germplasm innovation technology.
Description
Technical Field
The invention relates to the technical field of genetic breeding, in particular to a method for creating a purple perilla germplasm.
Technical Field
Perilla frutescens (Perilla frutescens L.Britt), also known as common Perilla Frutescens, has specific fragrance, and can refresh and refresh brain. The stems, leaves, seeds and flowers of the purple perilla have special activity and nutrition, and are economic crops with wide application. The perilla seed contains a large amount of grease, the oil yield can reach 45% -55%, unsaturated fatty acid accounts for more than 90% of the total oil content, and the alpha-linolenic acid content can reach 50% -70%. The perilla is used as a full-function plant, and the stems, leaves, flowers and seeds of the perilla have the value of exploitation and utilization.
At present, the market of the purple perilla germplasm is disordered, most of the purple perilla germplasm is sold in a mixed way, along with the wide application of the purple perilla, the demand for special germplasm materials is increased, and the demand for specific varieties of purple perilla is also increased. However, the number of the varieties of the purple perilla grown in the market is small, the purple perilla is used as a ta flower plant, the purple perilla flowers are arranged in a round umbrella shape on the inflorescence axis, each small flower has an opening of less than 5mm, four stamens are arranged, the emasculation is realized in the bud period, the technical operation is quite difficult, the hybridization breeding process of the purple perilla is greatly restricted, and therefore, the technical difficulty is brought to creating new varieties of the purple perilla by means of the conventional breeding method. In order to develop new germplasm creation by effectively utilizing perilla germplasm resources, a new way is necessary to be opened up, and the damage of mechanical damage to recipient plants can be effectively avoided by utilizing the pollen tube channel method technical means, and the technology is simple and easy to implement. The technology not only provides a basis for the germplasm creation of the purple perilla, but also provides a premise for the industrialized development of the purple perilla. The pollen tube channel method is a very effective technical means for creating new germplasm resources, and has a mature technical system in field crops such as rice, wheat, soybean, cotton, flax and the like. However, the application of the purple perilla in purple perilla has not been reported.
Disclosure of Invention
Aiming at the limitation of the prior art, the invention provides a method for creating the germplasm of the purple perilla, which effectively reduces the operation difficulty of conventional crossbreeding in the background technology.
In order to achieve the above purpose, the present invention adopts the following technical scheme:
a method for creating a germ plasm of a purple perilla, comprising the following steps:
1) Selection of donor and acceptor perilla:
selecting varieties with agronomic character differences in the biological field from donor perilla and acceptor perilla;
2) Harvesting and preserving seeds:
the donor perilla and the acceptor perilla which are selected in the step 1) are ripe seeds in the current year, the seeds are air-dried after being picked, the seeds are removed from impurities and shriveled, the seeds are placed at room temperature, and spring sowing is carried out in the next year;
3) Cultivation of donor and acceptor perilla:
planting the donor and acceptor perilla seeds obtained in the step 2) according to a conventional method, and normally growing;
4) Extraction of donor perilla DNA:
before the flowering period, selecting leaves of a donor perilla plant to extract DNA, and preserving at low temperature;
5) Receptor perilla flowering phase selection:
when the acceptor perilla plant is bloomed, observing and selecting the full bloom period of the floret;
6) Creation of new purple perilla germplasm:
selecting florets to be instilled, absorbing donor perilla DNA liquid extracted in advance by using a 0.5mL injection needle tube or a liquid shifter, instilling the donor perilla DNA liquid into a floret of acceptor perilla, wrapping the DNA solution by petals, fully infiltrating stigmas, repeating instilling after natural air drying, and marking;
7) Seed collection:
after the acceptor perilla is normally set, harvesting, recording and storing the marked acceptor perilla seeds;
8) Variant germplasm screening:
and (3) after planting the harvested acceptor perilla seeds, taking acceptor perilla as a control, observing agronomic characters, and screening perilla variant plants.
Further, in the above-mentioned method for creating a germ plasm of perilla, in the step 1), the agronomic trait in the biological field is any one of the following: plant shape, plant height, stalk color, leaf shape, corolla color, flowering phase, growth phase, she Baopian shape and size, maturity phase, fruit nap, fruit calyx size, fruit opening size, seed size, color and leaf fragrance.
Further, in the above-mentioned method for creating the germplasm of the purple perilla, in the step 2), the purple perilla is naturally air-dried for 3-7 days after harvesting, and the purple perilla is placed at a room temperature lower than 20 ℃.
Further, in the above-mentioned method for creating a germ plasm of perilla, in the step 3), perilla seeds are planted in a field, a greenhouse or a flowerpot by a conventional method.
In step 4), leaves newly grown for 3-5 days are selected, DNA is extracted by CTAB method, and the obtained product is preserved at-20 ℃.
Further, in the above-mentioned method for creating a germ plasm of a perilla, in the step 5), the full bloom period of the floret means: the corolla exposes the calyx, the anther cracks and breaks apart to form powder, and the stigma extends beyond the corolla.
Further, in the above-mentioned method for creating a germ plasm of Perilla frutescens, in the step 6), the instillation amount of the donor Perilla frutescens DNA liquid is 2-10 μl, and the instillation is repeated 2-3 times.
Further, in the above-mentioned method for creating a germ plasm of perilla, in the step 8), the harvested receptor perilla seeds are planted in a greenhouse, a flowerpot or a field.
In step 8), the purple perilla variant plants are screened according to biological concept by observing agronomic characters; the agricultural property is any one of the following: plant shape, plant height, stalk color, leaf shape, corolla color, flowering phase, growth phase, she Baopian shape and size, maturity phase, fruit nap, fruit calyx size, fruit opening size, seed size, color and leaf fragrance.
In step 8), the screening of the purple perilla variant plants is performed by a method of observing phenotypic changes by naked eyes, measuring actual data by a measuring tool, judging or identifying molecular level; the measuring tool measures actual data, namely length and outer diameter are measured through a ruler and a vernier caliper, and weight is measured through an electronic balance.
Compared with the prior art, the invention has the following advantages:
1) The method has wide selection of the purple perilla donor, and has no flowering phase disorder of the donor and the acceptor purple perilla, thereby greatly reducing the space-time restriction of the donor and the acceptor purple perilla.
2) Compared with the traditional conventional crossbreeding, the method eliminates the difficulty of emasculation and reduces the damage to plants in the crossbreeding process.
3) The method has simple operability and reduces the requirements of manual training and proficiency.
4) The method can cause the perilla acceptor to generate mutation in the seed forming process, and different mutation results are the feasibility and applicability of the perilla germplasm innovation technology.
Drawings
FIG. 1 shows the variation plants and leaf sizes obtained by screening with the Perilla frutescens germplasm innovation method according to the embodiment of the present invention.
Detailed Description
The present invention will be described in further detail below in order to make the objects, technical solutions and advantages of the present invention more apparent. It is to be understood that the description is only intended to illustrate the invention and is not intended to limit the scope of the invention.
Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs, and the terms used herein in this description of the invention are for the purpose of describing particular embodiments only and are not intended to be limiting of the invention. Reagents and instruments used herein are commercially available, and reference to characterization means is made to the relevant description of the prior art and will not be repeated herein.
For a further understanding of the present invention, the present invention will be described in further detail with reference to the following preferred embodiments.
Example 1
A method for creating a germ plasm of a purple perilla, comprising the following steps:
1) Selection of donor and acceptor perilla:
selecting varieties with agronomic character differences in the biological field from donor perilla and acceptor perilla;
2) Harvesting and preserving seeds:
the donor perilla and the acceptor perilla which are selected in the step 1) are ripe seeds in the current year, the seeds are air-dried after being picked, the seeds are removed from impurities and shriveled, the seeds are placed at room temperature, and spring sowing is carried out in the next year;
3) Cultivation of donor and acceptor perilla:
planting the donor and acceptor perilla seeds obtained in the step 2) according to a conventional method, and normally growing;
4) Extraction of donor perilla DNA:
before the flowering period, selecting leaves of a donor perilla plant to extract DNA, and preserving at low temperature;
5) Receptor perilla flowering phase selection:
when the acceptor perilla plant is bloomed, observing and selecting the full bloom period of the floret;
6) Creation of new purple perilla germplasm:
selecting florets to be instilled, absorbing donor perilla DNA liquid extracted in advance by using a 0.5mL injection needle tube or a liquid shifter, instilling the donor perilla DNA liquid into a floret of acceptor perilla, wrapping the DNA solution by petals, fully infiltrating stigmas, repeating instilling after natural air drying, and marking;
7) Seed collection:
after the acceptor perilla is normally set, harvesting, recording and storing the marked acceptor perilla seeds;
8) Variant germplasm screening:
and (3) after planting the harvested acceptor perilla seeds, taking acceptor perilla as a control, observing agronomic characters, and screening perilla variant plants.
In step 1), the agronomic trait in the biological field is any one of the following: plant shape, plant height, stalk color, leaf shape, corolla color, flowering phase, growth phase, she Baopian shape and size, maturity phase, fruit nap, fruit calyx size, fruit opening size, seed size, color and leaf fragrance.
In the step 2), natural air drying is carried out for 3-7 days after harvesting, and the room temperature is lower than 20 ℃.
In step 3), perilla seeds are planted in a field, a greenhouse or a flowerpot by a conventional method.
In step 4), selecting new leaves growing for 3-5 days, extracting DNA by using a CTAB method, and preserving at-20 ℃.
In step 5), the full bloom stage of the floret refers to: the corolla exposes the calyx, the anther cracks and breaks apart to form powder, and the stigma extends beyond the corolla.
In the step 6), the instillation amount of the donor perilla DNA liquid is 2-10 mu l, and the instillation is repeated for 2-3 times.
In step 8), the harvested receptor perilla seed is planted in a greenhouse, a flowerpot or a field.
In the step 8), the perilla variant plant is screened according to biological concept by observing agronomic characters; the agricultural property is any one of the following: plant shape, plant height, stalk color, leaf shape, corolla color, flowering phase, growth phase, she Baopian shape and size, maturity phase, fruit nap, fruit calyx size, fruit opening size, seed size, color and leaf fragrance.
In the step 8), the purple perilla variant plants are screened by a method of observing phenotype change by naked eyes, measuring actual data by a measuring tool, judging or identifying molecular level; the measuring tool measures actual data, namely length and outer diameter are measured through a ruler and a vernier caliper, and weight is measured through an electronic balance.
Example 2
A simple and easy-to-operate purple perilla germplasm innovation method comprises the following steps:
(1) Selection of donor and acceptor perilla: the purple perilla with green leaf color and oblong leaf shape is selected as a receptor, and the leaf area is 12.5cm multiplied by 18cm; purple leaf, wrinkled leaf, purple perilla germplasm with leaf area of 8cm multiplied by 13cm as a donor;
(2) Harvesting and preserving seeds: naturally air-drying the mature seeds in the same year for 3-7 days after harvesting, removing impurities and shriveling, standing at room temperature below 20 ℃, and spring sowing for later use;
(3) Donor and acceptor perilla cultivation: planting donor Perilla frutescens in field, and planting acceptor Perilla frutescens in flowerpot according to conventional method to extract DNA for normal growth of acceptor Perilla frutescens;
(4) Extraction of donor perilla DNA: before the flowering period, selecting newly grown 3-5 days of leaves on a donor perilla plant, extracting DNA by using a CTAB method, detecting the concentration of 50-100 ng/. Mu.l of the DNA, and storing in a refrigerator at-20 ℃ for later use;
(5) Flowering phase selection: when the acceptor purple perilla plants bloom, observing and selecting the full bloom period of florets, and the characteristics are as follows: the corolla exposes the calyx, the anther basically breaks apart to form powder, and the stigma extends beyond the corolla;
(6) Creation of new purple perilla germplasm: selecting florets to be instilled, absorbing donor DNA liquid extracted in advance by using a 0.5mL injection needle tube or a liquid shifter, instilling about 2-5 mu l of the donor DNA liquid into a floret disc of a receptor purple perilla, wrapping the DNA solution by petals without overflowing, fully infiltrating the stigmas by the DNA solution, repeating for 2 times after naturally airing, marking clearly, and hanging a label for recording;
(7) Seed collection: after the perilla is normally firm, harvesting, recording and storing the marked perilla seeds;
(8) Variant germplasm screening: the harvested perilla seeds are planted in a greenhouse flowerpot, the acceptor perilla is used as a control, the observation and recording of agronomic characters are carried out, the perilla variant plants are screened according to the biological concept, the new perilla germplasm is obtained, the leaf area of the acceptor perilla is 12.5cm multiplied by 18cm, 1 variant plant appears in the D0 generation plants, and the leaf area is 21cm multiplied by 30cm, as shown in figure 1.
Table 1 below shows statistics of the results of the different flowering phase treatments in the experiment in this example.
The foregoing description of the preferred embodiments of the invention is not intended to be limiting, but rather is intended to cover all modifications, equivalents, or alternatives falling within the spirit and principles of the invention.
Claims (10)
1. A method for breeding perilla, which is characterized by comprising the following steps:
1) Selection of donor and acceptor perilla:
selecting varieties with agronomic character differences in the biological field from donor perilla and acceptor perilla;
2) Harvesting and preserving seeds:
the donor perilla and the acceptor perilla which are selected in the step 1) are ripe seeds in the current year, the seeds are air-dried after being picked, the seeds are removed from impurities and shriveled, the seeds are placed at room temperature, and spring sowing is carried out in the next year;
3) Cultivation of donor and acceptor perilla:
planting the donor and acceptor perilla seeds obtained in the step 2) according to a conventional method, and normally growing;
4) Extraction of donor perilla DNA:
before the flowering period, selecting leaves of a donor perilla plant to extract DNA, and preserving at low temperature;
5) Receptor perilla flowering phase selection:
when the acceptor perilla plant is bloomed, observing and selecting the full bloom period of the floret;
6) Creation of new purple perilla germplasm:
selecting florets to be instilled, instilling donor perilla DNA liquid into a floret of acceptor perilla, wrapping the DNA solution by petals, fully infiltrating stigmas, repeating instilling after natural air drying, and marking;
7) Seed collection:
after the acceptor perilla is normally set, harvesting, recording and storing the marked acceptor perilla seeds;
8) Variant germplasm screening:
and (3) after planting the harvested acceptor perilla seeds, taking acceptor perilla as a control, observing agronomic characters, and screening perilla variant plants.
2. The method according to claim 1, wherein in step 1), the agronomic trait in the biological field is any one of the following: plant shape, plant height, stalk color, leaf shape, corolla color, flowering phase, growth phase, she Baopian shape and size, maturity phase, fruit nap, fruit calyx size, fruit opening size, seed size, color and leaf fragrance.
3. The method according to claim 1, wherein in step 2), the natural air drying is performed for 3-7 days after harvesting, and the temperature is kept below 20 ℃.
4. The method according to claim 1, wherein in the step 3), the perilla seeds are planted in a field, a greenhouse or a flowerpot in a conventional manner.
5. The method according to claim 1, wherein in step 4), leaves newly grown for 3-5 days are selected, DNA is extracted by CTAB method, and the obtained product is stored at-20 ℃.
6. The method according to claim 1, wherein in the step 5), the full bloom stage of the floret is: the corolla exposes the calyx, the anther cracks and breaks apart to form powder, and the stigma extends beyond the corolla.
7. The method according to claim 1, wherein in the step 6), the liquid injection amount of the donor perilla DNA is 2-10. Mu.l, and the injection is repeated 2-3 times.
8. The method of claim 1, wherein in step 8), the harvested acceptor perilla seeds are planted in a greenhouse, a flowerpot or a field.
9. The method according to claim 1, wherein in step 8), the Perilla variant plants are selected by observing agronomic traits and based on biological concepts; the agricultural property is any one of the following: plant shape, plant height, stalk color, leaf shape, corolla color, flowering phase, growth phase, she Baopian shape and size, maturity phase, fruit nap, fruit calyx size, fruit opening size, seed size, color and leaf fragrance.
10. The method according to claim 1, wherein in the step 8), the Perilla variant plants are selected by visual observation of phenotype change, measurement of actual data by measuring tool, or molecular level identification; the measuring tool measures actual data, namely length and outer diameter are measured through a ruler and a vernier caliper, and weight is measured through an electronic balance.
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| CN103109740A (en) * | 2013-03-08 | 2013-05-22 | 中国科学院近代物理研究所 | New strain breeding method of high-quality purple perilla |
| CN111972280A (en) * | 2020-08-24 | 2020-11-24 | 河北农业大学 | Pollination method for improving purple perilla hybridization success rate |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN103109740A (en) * | 2013-03-08 | 2013-05-22 | 中国科学院近代物理研究所 | New strain breeding method of high-quality purple perilla |
| CN111972280A (en) * | 2020-08-24 | 2020-11-24 | 河北农业大学 | Pollination method for improving purple perilla hybridization success rate |
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