CN114391472B - Method for improving hybrid seed setting rate of pineapple - Google Patents
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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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- A—HUMAN NECESSITIES
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- A01G—HORTICULTURE; CULTIVATION OF VEGETABLES, FLOWERS, RICE, FRUIT, VINES, HOPS OR SEAWEED; FORESTRY; WATERING
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Abstract
The invention provides a method for improving the seed setting rate of pineapple hybridization, which comprises the steps of collecting pollen after a parent strain is selected for flower forcing, spraying pollen treatment by using a pollen treatment agent containing calcium chloride, urea and ferric chloride to improve the cutinase activity in the pollen, promoting the pollen germination, improving the affinity of pineapple hybridization and further improving the insemination rate, then treating a maternal stigma by using a medicament containing cytokinin and citric acid to remove callose accumulation, promoting the pollen germination and pollen tube growth after pollination, and improving the seed setting rate. The method is simple and easy to master, does not need laboratory operation, can be completed in the field, can solve the problem of low seed setting rate of the pineapples due to hybridization affinity, and greatly improves the efficiency of pineapple hybridization breeding.
Description
Technical Field
The invention relates to the technical field of plant crossbreeding, in particular to a method for improving the hybrid seed setting rate of pineapples.
Background
The pineapple industry in China has the problems of single variety, degeneration of species and the like, and the improvement of the existing variety needs to be promoted by means of sexual hybridization and the like. However, the hybrid breeding work in China starts late, accumulates less and progresses slowly, and various problems exist in the hybrid breeding of pineapples, such as: the high self-incompatibility of the pineapples hinders the hybridization compatibility among varieties and lines, increases the difficulty of hybridization breeding, and also causes the phenomena of no or little seed formation of hybridized fruits in actual work, which seriously affect the smooth proceeding of the hybridization work, thereby hindering the hybridization breeding of the pineapples.
Chinese patent CN101861828A discloses a pineapple hybridization method, pollen which can be used for hybridization pollination is selected by reasonably allocating parents, pollination is carried out when small flower petals are open, stigma is slightly seen and honeydew is secreted on the stigma, and the pineapple hybridization breeding method is low in seed setting rate. In the hybridization breeding work, the common methods for overcoming incompatibility comprise the steps of treating stigmas with medicaments, cutting the stigmas for pollination, grafting stigmas, in vitro fertilization, hot water treatment, laser and high-voltage electrostatic field, pollen guide, pollination in ovaries and the like, the treatment effect is different due to different plant types, and the research reports about pineapple hybridization incompatibility are less at present. Therefore, it is necessary to provide a method for improving the hybridization affinity between pineapple species and the seed set rate.
Disclosure of Invention
Therefore, the invention aims to provide a method for improving the hybrid seed setting rate of pineapples.
The technical scheme of the invention is realized as follows:
a method for improving the hybrid seed setting rate of pineapples comprises the following steps:
a. flower forcing: selecting parent strains, and irrigating a flower accelerant to obtain hybrid parent strains;
b. pollen pretreatment: picking anther of male parent after flower bud blooms, taking out pollen from anther, spraying pollen with pollen treating agent, and air drying to obtain pollen; the pollen treating agent comprises calcium chloride, urea and ferric chloride;
c. treating stigma: smearing the maternal stigma with the medicament;
d. and (3) cross pollination: c, taking the pollen treated in the step b to perform artificial pollination on the stigma treated in the step c, and closing the petals of the female parent after the pollination is finished;
e. seed taking: and cutting the hybridized small fruits and collecting seeds after the fig is fully yellow and ripe.
Further, the species of the parent strain were 'shenwan' (a. Comosus cv. 'Shen Wan') and 'Yulinglong' (a. Nanus cv. 'Yulinglong').
Furthermore, each 1L of the flower forcing agent contains 8-12 g of urea and 22-28 mL of ethephon, so that the flowering of parent plants is promoted, and the flowering period of the parent plants is unified.
Furthermore, the flower forcing agent is used by pouring from the top of the plant core, and the pouring amount is 40-50 mL/plant.
Further, the collection time of the anthers is 9-10 am, the buds burst in the period, and dew on the flowers is dried.
Furthermore, the pollen is taken out by tearing the anther and applying external force to the anther to naturally disperse the pollen in the anther so as to reduce the damage to the pollen.
Furthermore, the concentration of calcium chloride in the pollen treating agent is 0.2-0.5 wt%, the concentration of urea is 0.1-0.3 wt%, and the concentration of ferric chloride is 0.2-0.4 wt%.
Further, the pollen treatment agent is used by applying the pollen treatment agent to pollen in the form of spray; the application amount is 2-5 mL/g pollen.
Further, the medicament comprises: cytokinin (CTK) and citric acid.
Furthermore, each 1L of the medicine contains 0.03-0.05 g of cytokinin and 0.1-0.2 g of citric acid.
Furthermore, the temperature of the pollen pretreatment and the hybrid pollination is 20-30 ℃.
Further, the optimal temperature of the pollen pretreatment and the cross pollination is 28-30 ℃.
Compared with the prior art, the invention has the beneficial effects that:
the pollen treating agent containing calcium chloride, urea and ferric chloride is used for spraying and treating the pollen, so that the cutinase activity in the pollen can be improved, the pollen germination is promoted, the pineapple hybridization affinity is improved, and the insemination rate is further improved, which is shown in that the seed setting rate is increased.
The invention uses the medicament containing cytokinin and citric acid to treat the stigma, can remove callose accumulation without damaging the stigma, can promote pollen grain germination and pollen tube growth after pollination, and improves the seed setting rate.
The method is simple and easy to master, does not need laboratory operation, can be completed in the field, can solve the problem of low seed setting rate of the pineapples due to hybridization affinity, and greatly improves the efficiency of pineapple hybridization breeding.
Detailed Description
For clear and complete description of the technical solutions in the present invention, it is obvious that the inventor combines the embodiments to describe, but the following embodiments describe only some embodiments of the present invention, not all embodiments. All other embodiments, which can be obtained by a person skilled in the art without making any creative effort based on the embodiments in the present invention, belong to the protection scope of the present invention.
The hybrid parent materials used in the experiment of the invention are 'Shenwan' (A.comosus cv. 'Shenwan') and 'Yulinglong' (A.nanus cv. 'Yulinglong'), respectively;
the number of the hybridized flowers in each embodiment of the invention is 50;
example 1
A method for improving the hybrid seed setting rate of pineapples comprises the following steps:
a. flower forcing: selecting a parent plant, taking 'Shenwan' pineapple as a female parent and 'Yulinglong' pineapple as a male parent, pouring every 1L of flower forcing agent containing 10g of urea and 25mL of ethephon from the top of the plant heart, wherein the pouring amount is 45mL per plant, and obtaining a hybrid parent plant;
b. pollen pretreatment: picking up anthers of male parents after flower buds bloom at 9-10 am, taking out pollen from the anthers, applying the pollen treating agent to the pollen in a spray mode at 28 ℃, applying 3mL of the pollen treating agent to every 1g of the pollen, and drying in the air to obtain the pollen; the concentration of calcium chloride in the pollen treating agent is 0.3wt%, the concentration of urea is 0.2wt%, and the concentration of ferric chloride is 0.3wt%;
c. treating stigma: smearing the stigma of the female parent with a preparation containing 0.04g cytokinin and 0.2g citric acid per 1L;
d. and (3) cross pollination: c, carrying out artificial pollination at 28 ℃, lightly poking the dissembled female parent petals with tweezers, clamping the male parent pollen processed in the step b with the tweezers, adhering the male parent pollen on the stigma processed in the step c, and lightly closing the petals;
e. seed taking: in the middle and last ten days of 6 months, when the fig is fully yellow and ripe, each hybrid fruit is cut off by a small knife, and seeds are collected.
Example 2
A method for improving the hybrid seed setting rate of pineapples comprises the following steps:
a. flower forcing: selecting a parent plant, taking 'Shenwan' pineapple as a female parent and 'Yulinglong' pineapple as a male parent, and pouring flower forcing agent containing 8g of urea and 22mL of ethephon into every 1L of the top of the plant core, wherein the pouring amount is 50mL per plant, so as to obtain a hybrid parent plant;
b. pollen pretreatment: picking up anthers of male parents after flower buds bloom at 9-10 am, taking out pollen from the anthers, applying the pollen treating agent to the pollen in a spray mode at 30 ℃, applying 2mL of the pollen treating agent to every 1g of the pollen, and air-drying to obtain the pollen; the concentration of calcium chloride in the pollen treating agent is 0.5wt%, the concentration of urea is 0.3wt%, and the concentration of ferric chloride is 0.4wt%;
c. treating stigma: smearing the stigma of the female parent with a preparation containing 0.03g cytokinin and 0.1g citric acid per 1L;
d. and (3) cross pollination: c, carrying out artificial pollination at 30 ℃, lightly poking the dissilient female parent petals with tweezers, clamping the male parent pollen processed in the step b with the tweezers, adhering the pollen to the stigma processed in the step c, and lightly closing the petals;
e. seed taking: in the middle and last ten days of 6 months, when the fig is fully yellow and ripe, each hybrid fruit is cut off by a small knife, and seeds are collected.
Example 3
A method for improving the hybrid seed setting rate of pineapples comprises the following steps:
a. flower forcing: selecting a parent plant, taking 'Shenwan' pineapple as a female parent and 'Yulinglong' pineapple as a male parent, pouring every 1L of flower forcing agent containing 10g of urea and 25mL of ethephon from the top of the plant heart, wherein the pouring amount is 45mL per plant, and obtaining a hybrid parent plant;
b. pollen pretreatment: picking up anthers of male parents after flower buds bloom at 9-10 am, taking out pollen from the anthers, applying the pollen treating agent to the pollen in a spray mode at 28 ℃, applying 3mL of the pollen treating agent to every 1g of the pollen, and drying in the air to obtain the pollen; the concentration of calcium chloride in the pollen treating agent is 0.3wt%, the concentration of urea is 0.2wt%, and the concentration of ferric chloride is 0.3wt%;
c. treating stigma: smearing maternal stigma with a drug containing 0.04gABA and 0.2g 6-BA per 1L;
d. and (3) cross pollination: c, carrying out artificial pollination at 28 ℃, slightly poking the dissilient female parent petals with forceps, clamping the male parent pollen treated in the step b with the forceps, adhering the male parent pollen on the stigma treated in the step c, and slightly closing the petals;
e. seed taking: in the middle and last ten days of 6 months, when the fig is fully yellow and ripe, each hybrid fruit is cut off by a small knife, and seeds are collected.
Example 4
A method for improving the hybrid seed setting rate of pineapples comprises the following steps:
a. flower forcing: selecting a parent plant, taking 'Shenwan' pineapple as a female parent and 'Yulinglong' pineapple as a male parent, pouring every 1L of flower forcing agent containing 10g of urea and 25mL of ethephon from the top of the plant heart, wherein the pouring amount is 45mL per plant, and obtaining a hybrid parent plant;
b. pollen pretreatment: picking anthers of male parents after flower buds bloom at 9-10 am, taking pollen out of the anthers, applying the pollen treating agent to the pollen in a spray mode at 28 ℃, applying 3mL of the pollen treating agent to 1g of the pollen, and air-drying to obtain pollen; the concentration of calcium chloride in the pollen treating agent is 0.1wt%, the concentration of urea is 0.5wt%, and the concentration of ferric chloride is 0.1wt%;
c. treating stigma: smearing the maternal stigma with a 1L preparation containing 0.04g cytokinin and 0.2g citric acid;
d. and (3) cross pollination: c, carrying out artificial pollination at 28 ℃, slightly poking the dissilient female parent petals with forceps, clamping the male parent pollen treated in the step b with the forceps, adhering the male parent pollen on the stigma treated in the step c, and slightly closing the petals;
e. seed taking: in the middle and last ten days of 6 months, when the fig is fully yellow and ripe, each hybrid fruit is cut off by a small knife, and seeds are collected.
Example 5
A method for improving the hybrid seed setting rate of pineapples comprises the following steps:
a. flower forcing: selecting a parent plant, pouring every 1L of flower forcing agent containing 10g of urea and 25mL of ethephon from the top of the plant core by taking the Gonwu pineapple as a female parent and the Yulinglong pineapple as a male parent, wherein the pouring amount is 45mL per plant, and obtaining a hybrid parent plant;
b. pollen pretreatment: picking anthers of male parents after flower buds bloom at 9-10 am, taking pollen out of the anthers, applying the pollen treating agent to the pollen in a spray mode at 28 ℃, applying 3mL of the pollen treating agent to 1g of the pollen, and air-drying to obtain pollen; the concentration of calcium chloride in the pollen treating agent is 0.3wt%, and the concentration of urea in the pollen treating agent is 0.2wt%;
c. treating stigma: smearing the stigma of the female parent with a preparation containing 0.04g cytokinin and 0.2g citric acid per 1L;
d. and (3) cross pollination: c, carrying out artificial pollination at 28 ℃, lightly poking the dissembled female parent petals with tweezers, clamping the male parent pollen processed in the step b with the tweezers, adhering the male parent pollen on the stigma processed in the step c, and lightly closing the petals;
e. seed taking: in the middle and last ten days of 6 months, when the fig is fully yellow and ripe, each hybrid fruit is cut off by a small knife, and seeds are collected.
Example 6
A method for improving the hybrid seed setting rate of pineapples comprises the following steps:
a. flower forcing: selecting a parent plant, taking 'Shenwan' pineapple as a female parent and 'Yulinglong' pineapple as a male parent, pouring every 1L of flower forcing agent containing 10 urea and 25mL ethephon from the top of the plant heart, wherein the pouring amount is 45mL per plant, and obtaining a hybrid parent plant;
b. pollen pretreatment: picking up anthers of male parents after flower buds bloom at 9-10 am, taking out pollen from the anthers, applying the pollen treating agent to the pollen in a spray mode at 28 ℃, applying 3mL of the pollen treating agent to every 1g of the pollen, and drying in the air to obtain the pollen; the concentration of calcium chloride in the pollen treating agent is 0.3wt%, the concentration of urea is 0.2wt%, and the concentration of ferric chloride is 0.3wt%;
c. and (3) cross pollination: c, carrying out artificial pollination at 28 ℃, lightly poking the dissembled female parent petals with tweezers, clamping the male parent pollen processed in the step b with the tweezers, adhering the male parent pollen on the stigma processed in the step c, and lightly closing the petals;
d. seed taking: in the middle and last ten days of 6 months, when the fig is fully yellow and ripe, each hybrid fruit is cut off by a small knife, and seeds are collected.
Example 7
A method for improving the hybrid seed setting rate of pineapples comprises the following steps:
a. flower forcing: selecting a parent plant, taking 'Shenwan' pineapple as a female parent and 'Yulinglong' pineapple as a male parent, pouring every 1L of flower forcing agent containing 10g of urea and 25mL of ethephon from the top of the plant heart, wherein the pouring amount is 45mL per plant, and obtaining a hybrid parent plant;
b. pollen collection: picking anther of male parent after flower bud blooms at 9-10 am, and taking out pollen from anther;
c. treating stigma: smearing the maternal stigma with a 1L preparation containing 0.04g cytokinin and 0.2g citric acid;
d. and (3) cross pollination: c, carrying out artificial pollination at 28 ℃, lightly poking the dissembled female parent petals with tweezers, clamping the male parent pollen processed in the step b with the tweezers, adhering the male parent pollen on the stigma processed in the step c, and lightly closing the petals;
e. seed taking: in the middle and last ten days of 6 months, when the fig is fully yellow and ripe, each hybrid fruit is cut off by a small knife, and seeds are collected.
Evaluation of hybridization Effect:
(1) The number of fruits with seeds, the number of seeds and the number of ovules of the above examples were counted, and the seed setting rate and affinity index were calculated, and the results are shown in table 1.
TABLE 1 hybridization affinity of pineapple treated by different methods
| Percentage of seed formation (%) | Affinity index | |
| Example 1 | 100 | 0.227 |
| Example 2 | 100 | 0.215 |
| Example 3 | 84.0 | 0.152 |
| Example 4 | 92.0 | 0.165 |
| Example 5 | 76.0 | 0.149 |
| Example 6 | 64.0 | 0.131 |
| Example 7 | 62.0 | 0.128 |
The result shows that the pollen treated by the pollen treating agent and the agent has high seed setting rate and affinity index for pollination, which are both obviously higher than the method of only using one of the pollen treating agent and the agent or using different components of the agent.
Test example 1
And (3) hybridization combination: 'Yulinglong' × 'Shenwan';
the experiment was set up with 2 treatments, i.e., treatments with the treatment method of the invention and without the pollen treatment agent and agent of the invention applied, the cross pollination method being as in example 1 (except pollination temperature and hybridization combination);
the pollination temperature is respectively set to be 20 ℃, 28 ℃ and 35 ℃ (artificial climate box), each group combines 6 inflorescences, and each inflorescence has 13 florets.
TABLE 2 seed set percentage of Yulinglong ' x ' Shenwan ' at different temperatures
From the table 2, it can be seen that the seed setting rate of the 'Yulinglong' x 'God Bay' is the highest when the pollination temperature is 28 ℃, the affinity of the pollen treated by the method is improved, the seed setting rate is basically doubled at different hybridization temperatures, and the pollination seed setting rate is the highest at 28 ℃ and can reach 100%.
Test example 2
Hybridization combination: 'God gulf' × 'Yu exquisite', 'Yu exquisite' × 'God gulf';
hybridization method referring to example 1, the control was not treated with a pollen treatment agent spray.
TABLE 3 hybridization affinity of pineapple
As can be seen from Table 3, the pollen treated by the pollen treating agent and the agent of the invention is used for pollination, all pollinated small fruits bear seeds, the seed bearing rate reaches 100%, the affinity index is about 2 times of that of the control, the hybridization affinity is obviously improved, and the change of the parent and the female parent has no significant influence on the hybridization effect.
In conclusion, the method for the seed setting rate of pineapple hybridization provided by the invention can effectively promote pollen germination and improve the affinity of pineapple hybridization. In addition, the preparation method is simple and easy to operate, and the cross breeding efficiency is high.
The above description is only for the purpose of illustrating the preferred embodiments of the present invention and should not be taken as limiting the scope of the present invention, which is intended to cover any modifications, equivalents, improvements, etc. within the spirit and scope of the present invention.
Claims (5)
1. A method for improving the hybrid seed setting rate of pineapples is characterized by comprising the following steps:
a. flower forcing: selecting parent strains, and irrigating a flower accelerant to obtain hybrid parent strains;
b. pollen pretreatment: picking anther of male parent after flower bud blooms, taking out pollen from anther, spraying pollen with pollen treating agent, and air drying to obtain pollen; the pollen treating agent comprises calcium chloride, urea and ferric chloride;
c. treating stigma: smearing the maternal stigma with the medicament;
d. and (3) cross pollination: c, taking the pollen treated in the step b to carry out artificial pollination on the stigma treated in the step c, and closing the petals of the female parent after the pollination is finished;
e. seed taking: cutting the hybrid small fruits and collecting seeds after the fig is yellow and ripe;
the parent strain is 'Shenwan' and 'Yulinglong';
0.03 to 0.05g of cytokinin and 0.1 to 0.2g of citric acid are contained in each 1L of the medicament;
the concentration of calcium chloride in the pollen treating agent is 0.2-0.5 wt%, the concentration of urea is 0.1-0.3 wt%, and the concentration of ferric chloride is 0.2-0.4 wt%;
the pollen treating agent is used by applying the pollen treating agent to pollen in a spray form; applying 2-5mL of the pollen treating agent to every 1g of pollen.
2. The method for improving hybrid seed production rate of pineapples according to claim 1, wherein the flower forcing agent comprises 8-12g urea and 22-28 mL ethephon per 1L of the flower forcing agent; the flower forcing agent is poured from the top of the plant core, and the pouring amount is 40-50mL per plant.
3. The method for improving hybrid seed setting rate of pineapple according to claim 1, wherein said anther is collected at 9-10 am.
4. The method for improving hybrid seed setting rate of pineapple according to claim 1, wherein the temperature for pollen pretreatment and hybrid pollination is 20 to 30 ℃.
5. The method for improving hybrid seed setting rate of pineapple according to claim 4, wherein the optimal temperature for pollen pretreatment and hybrid pollination is 28 to 30 ℃.
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| CN101861828A (en) * | 2010-05-05 | 2010-10-20 | 中国热带农业科学院南亚热带作物研究所 | Pineapple hybridization method |
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| USPP16396P3 (en) * | 2004-06-17 | 2006-03-28 | Dole Food Company, Inc. | Pineapple plant named ‘P-1972’ |
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| CN101861828A (en) * | 2010-05-05 | 2010-10-20 | 中国热带农业科学院南亚热带作物研究所 | Pineapple hybridization method |
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| Title |
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| ‘神湾’菠萝与亲缘关系不同的父本的交配亲和性研究;张伟等;《中国园艺学会2019年学术年会暨成立90周年纪念大会论文摘要集》;20191021;第90页 * |
| 菠萝主栽品种间的杂交亲和力测定;张伟等;《中国园艺学会2018年学术年会论文摘要集》;20181017;第2564页 * |
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