JP3949637B2 - Lettuce male sterile line and method for producing lettuce F1 seed - Google Patents

Description

  The present invention relates to a lettuce male sterile line and a method for producing lettuce F1 seeds using the male sterile line.

  Conventional plant varieties are widely known as fixed varieties grown through self-propagation and hybrid first varieties (hereinafter referred to as F1 varieties) using hybrid stress. The advantage of this F1 variety is that the characteristics of yield, disease resistance, etc. are superior to those of parents due to the expression of hybrid strength, the characteristics of the cultivated varieties are uniform, and the inheritance traits in the next generation This means that the profit of the breeder can be protected by the occurrence of the separation. So far, breeding methods for F1 varieties include a method using self-incompatibility and a method using male sterility (hereinafter referred to as “MS”). Incompatibility has become mainstream.

  F1 varieties have been developed and used in many crops such as Brassicaceae, Solanumaceae, and Cucurbitaceae, while lettuce (Lactuca sativa L.) is an important vegetable produced in many countries around the world. One of the few uncultivated crops. As Ryder (Non-Patent Documents 1, 2, and 3) states, there are three obstacles to the lettuce F1 variety. The first is that excellent self-incompatible individuals or male sterile individuals are not obtained as F1 parents. Second, lettuce is originally a self-propagating plant, and the rate of other breeding is low due to the structure of the flower organs. The third is that there is no flower-visiting insect with a unique relationship, and the third is that no visible traits linked to male sterility are found, and male sterile individuals cannot be selected unless they are in bloom.

In recent years, the occurrence of lettuce root rot ( Fusarium oxysporum sp. Lactucae ) has been expanding, and as a most effective measure, there is an increasing expectation for breeding resistant varieties. In addition, race differentiation is recognized in lettuce root rot and mixed contamination areas of multiple races have emerged. Therefore, breeding is inevitably progressing with the goal of combined race resistance.
Therefore, in order to allow the above-mentioned disease-resistant varieties to be cultivated more quickly, it is desired to make lettuce F1 varieties and to produce male sterile lines for that purpose.

E.J.Ryder, An epistatically controlled pollen sterile in lettuce (Lactuce sativa L.), Proc. Am. Soc. Hortic Sci., 83 (1963) 585-589 E.J.Ryder, A recessive male sterility gene in lettuce (Lactuca sativa L.), Proc.Am.Soc.Hortic.Sci.91 (1967) 366-368 E.J.Ryder, Genetic Studies in Lettuce J.Amer.Soc.Hor.Sci., 96 (6): 826-828 (1971)

Accordingly, an object of the present invention is to provide a male sterile line that serves as a basis for improving the variety of lettuce.
Another object of the present invention is to produce lettuce F1 seeds from the male sterile line.

  As a result of intensive research to solve the above-mentioned problems, the present inventors found a male sterile individual “733” that appeared as a mutation in the progeny of lettuce root rot resistance selection, and inbred A male sterile line “MS1024” was obtained in the progeny that the individual was bred after mating. Next, as a result of mating male sterile individuals with this "MS1024", the inheritance of male sterility was confirmed in the progeny, and it was successfully established as a male sterile line. The present invention has been completed based on such findings.

That is, the present invention includes the following inventions.
(1) A lettuce male sterile line having the male sterile gene of the lettuce male sterile line “MS1024” (patent biological deposit center deposit number: FERM P-19543) or a homologous gene thereof and exhibiting male sterile characteristics.
(2) The lettuce male sterile line according to (1), which is the lettuce male sterile line “MS1024” (patent biological deposit center deposit number: FERM P-19543).
(3) A lettuce male sterile line described in (1) or (2) is used as a mother, a lettuce male fertile line is used as a pollen parent, and the hybrid first seed (F1 seed) is obtained from the mother after the crossing. A method for producing lettuce F1 seeds, characterized in that
(4) Lettuce F1 seed obtained by the method according to (3).
(5) The lettuce F1 seed according to (4), deposited at the Patent Organism Depositary under the deposit number: FERM P-19542.

  According to the present invention, a lettuce male sterile line is provided. The lettuce male sterility line of the present invention shows a male sterility of a type that cannot produce pollen at all, but in crosses with other lines, there is an amount collected similar to the inbred line, and there is a problem with the female ability Absent. Therefore, the male sterile line according to the present invention can be used as a single parent of all existing lettuce lines, and a new hybrid variety can be bred in a short period of time. In particular, it is very useful for rapidly cultivating varieties resistant to diseases such as lettuce root rot, which is currently a problem in many lettuce producing areas.

1. Establishment of male sterility line According to the present invention, the male sterility gene of the lettuce male sterility line “MS1024” (patent biological deposit center deposit number: FERM P-19543) or a homologous gene thereof, A lettuce male sterile line exhibiting a spider trait is provided.

  Separation in the progeny with the male sterility gene or its homologous gene as the parent and the male wisdom individual as the pollen parent is male fertile: male sterility = 3: 1 in the F2 generation, and in the B1F1 generation Male feasible: male sterility = 1: 1.

  In addition, the above-mentioned “homologous gene” refers to a string consisting of DNA complementary to the base sequence of the male sterility gene of “MS1024” (patent biological deposit center deposit number: FERM P-19543) and stringent DNA encoding a protein that hybridizes under mild conditions and has an activity to confer male sterility. Here, stringent conditions refer to conditions under which so-called specific hybrids are formed and non-specific hybrids are not formed. For example, a nucleic acid having high homology, a base sequence having about 70% or more, preferably about 80% or more, more preferably about 90% or more, and most preferably about 95% or more with the base sequence of the male sterility gene. Examples include conditions in which complementary strands of DNA consisting of sequences hybridize and complementary strands of nucleic acids having lower homology do not hybridize. More specifically, the sodium concentration is 150 to 900 mM, preferably 600 to 900 mM, and the temperature is 60 to 68 ° C, preferably 65 ° C.

  The lettuce male sterility line of the present invention is typically the lettuce male sterility line `` MS1024 '', but `` MS1024 '' bred by crossing other lettuce varieties and selecting the progeny of the cross, etc. Also included are male sterility lines having the male sterility gene and inheriting the male sterility trait.

2. Production of lettuce first generation hybrid seeds The male sterility line obtained in (1) shows a single factor recessive inheritance pattern and can be used for seeding F1 seeds. That is, lettuce F1 seeds are obtained by crossing the male sterile line as a parent and the male fertile line as a pollen parent.

  In the mating for F1 seed production, all currently available fixed species are available as male fertile lines for use as pollen parents, such as Patriot (Nitto Agricultural Products), V Lettuce (Kaneko Seedling), Gold Cos Examples include Me (Turuta Seedling), Evening Red Fire (Takii Seedling) and Thermy (Sakata Seed). In addition to male malleability, those capable of modifying lettuce by imparting desired traits (disease resistance, night drawability, high yield, etc.) are preferred.

For the mating, pollen parental lettuce male fertile line pollen may be pollinated to the mother lettuce male sterile line pistil, and usual methods can be applied. Examples thereof include pollination with an air medium, insect medium, etc., and methods such as artificial mating in which pollen parent pollen is collected and sprayed or spotted on a mother pistil.
Moreover, a normal method can be applied in lettuce F1 seeding. For example, lettuce male sterile individuals and male flexible individuals are planted alternately in every row, mated, and then seeded from the mother only.

EXAMPLES Hereinafter, the present invention will be described more specifically with reference to examples. However, these examples do not limit the present invention.
(Example 1) Production and mating of lettuce male sterile line
(1) Establishment and characteristics of lettuce male sterile line
In the fall of 2001, we found a male sterile individual “733” that appeared as a mutation in the progeny of lettuce root rot resistance selection in the Romaine lettuce line grown in the Nagano Vegetable Flower Experiment Station. The inbred line “735” was crossed with this “733” to obtain the inbred line F1 seed “916”.

  In the spring of 2002, "916" was self-bred and F2 seed "1024" was collected. In the summer of 2002 and spring of 2003, "1024" was developed and the male sterile individuals that had been separated were acquired (hereinafter, this male sterile individual is referred to as "MS1024"). This "MS1024" was issued on September 30, 2003 at the National Institute of Advanced Industrial Science and Technology, Patent Biological Deposit Center (1st, 1st East, Tsukuba City, Ibaraki Prefecture, 6th postal code 305-8856). Has been deposited.

  In addition, male fertile individuals (Normal 1024: hereinafter referred to as “N1024”), “6056” (transferred from Tohoku Agricultural Research Center), “Patriot” (Nitto Agricultural Products Co., Ltd.), and “Gold Cosmy” (Turuta Seedling) Cultivated).

  The test plants were grown in a cell tray and then transplanted and grown in a 12 cm superpot and a drain bed in a greenhouse. The characteristics of male sterility were examined for the number of attached pollen grains per pistil, the number of self-propagating seeds per flower, and the number of hybrid seeds per flower for each generation of F1, F2 and B1F1. One to three strains per line / variety were tested, and the number of attached pollen grains per pistil was investigated for 5 pistils, and the number of seeds per flower was investigated for 10 head flowers. The petal colors and morphological abnormalities were observed sequentially during cultivation. The survey results are shown in Table 1.

  As shown in Table 1, no pollen was found on the “MS1024” pistil. Further, since no pollen grains were found in the pod sheath, the male sterility of “MS1024” is pollen sterility due to the fact that no pollen is formed. On the other hand, 442 to 582 pollen grains adhered on the female pupae of “N1024”, “Patriot”, and “Gold Cosmy”. On the “6056” pistil there was only 140.6 pollen grains, which was considerably smaller than other lines and varieties.

  A photograph of the shape of the female stigma of the male sterile individual “MS1024” and the state of pollen grain attachment is shown in FIG. In addition, FIG. 2 shows a photograph of the shape of the female stigma and the state of pollen grain adhesion of the “Patriot” male male individual.

  The number of self-propagating seeds per flower was 0 for “MS1024” and 17.3 for “N1024”. In “6056”, 0.09 seeds were difficult to harvest. Satisfactory seeds were obtained with 10.1 "Patriot" and 14.6 "Gold Cosmy".

The petal color was yellow in all lines, and there was no difference between “MS1024” and “N1024”. In addition, “6056” was observed between individual individuals, such as head flower size and petal morphological abnormalities, but “MS1024” and “N1024” did not show any abnormalities such as petal malformation. I couldn't distinguish.
Therefore, the male sterility of “MS1024” was considered to have the ability to grow lettuce F1 varieties, judging from the pollen production ability and seed production ability.

(2) Mating using male sterile lines
In the summer of 2002, the above-mentioned “MS1024” was used as a mother, and “6056”, “Patriot” and “Gold Cosmy” were crossed as pollen parents, and F1 seeds were collected.
In the fall of 2002, each F1 individual was mated to MS1024 to obtain B1F1 seeds, and F2 seeds were obtained by self-breeding of each F1 individual.
Table 2 shows the amount of F1 seed collected by hand mating, and Table 3 shows the amount of F2 seed collected by self-breeding.

  As shown in Table 2, hybrid seeds were not obtained from the cross between “MS1024”, but sufficient seeds of 15.0 were obtained from the cross between “MS1024” and “Gold Cosmy”. On the other hand, the amount of seeding was small in the cross between “MS1024” and “6056”.

  As shown in Table 3, in all of the combination lines examined, the amount of F2 generation was sufficient, 15.0-21.4 per flower.

(Example 2) Inheritance of lettuce male sterility
F2 seeds and B1F1 seeds collected in Example 1 were cultivated in a greenhouse twice in summer 2002 and summer 2003, and the state of separation of male sterility was investigated.
Confirmation of male sterility was judged by observation of exposed female pupae with a loupe and the amount of seeds collected. In addition, we observed changes in male sterility due to flowering time, and changes depending on the part of specific strains.
The separation of male sterility of the P, F1, F2 and B1F1 generations is shown in Table 4.

  As shown in Table 4, the F1 population showed sufficient fertility. “MS1024” x “Patriot” and “MS1024” x “N1024” F2 populations were investigated twice in 2002 and 2003, but in both surveys, male feasibility: male sterility = 3: 1 Separation was observed. For the B1F1 population, three lines were tested in 2003, but all obtained observations that matched a segregation ratio of 1: 1. These results indicate that male sterility is dominated by a single recessive gene. Seven male sterility genes have been reported so far, but the male sterility gene of “MS1024” was judged to be different from these and was named ms-serizawa.

  In addition, lettuce fixed seed "patriot", lettuce F1 seed (MS1024 x Patriot), as of September 30, 2003, National Institute of Advanced Industrial Science and Technology patent biological deposit center No. 6 Postal code 305-8566) are deposited as FERM P-19544 and FERM P-19542, respectively.

(Example 3) Characteristics of lettuce first-generation hybrid
In the fall of 2002, F1 seeds harvested in Example 1 were tested for lettuce root rot resistance in a greenhouse to investigate the degree of resistance. In the spring of 2003, these F1 lines were cultivated in the field, and the hybrid formation rate, yield, and core length were investigated. Table 5 shows the characteristics of the P and F1 generations.

  As shown in Table 5, the severity of lettuce root rot race 1 was 32.5 for MS1024 x Gold Cosmy, 19 for MS1024 x V lettuce, and MS1024 x 98. -0827 ”was 4. The F1 combination line with “MS1024” showed an intermediate numerical value for the race 1 with the parental lineage with respect to the race 1, but depending on the combination parent, stronger resistance can be expressed. For Race 2, the severity is 7.5 for MS1024 x Gold Cosmy, 0 for MS1024 x Night Red Fire, and 0 for MS1024 x Regina. Although sufficient resistance was exhibited, resistance could be further strongly expressed by the combined lines.

The hybrid formation rate of the tested F1 combination lines was 100%, and no single breeding individual appeared.
Therefore, the male sterility of “MS1024” was considered to have sufficient ability to produce F1 seeds with a purity of 100%.
Further, as shown in Table 5, many of the F1 combination lines showed intermediate values of the parent lines in the traits of total weight, core length, and number of leaves.

The photograph of the form of the female pupa of the male sterile individual “MS1024” is shown. The photograph of the form of the female pupa of male wisdom individual "Patriot" is shown.

Claims (4)

  Lettuce male sterile line “MS1024” (patent biological deposit center deposit number: FERM P-19543). A lettuce male sterile line according to claim 1 is used as a mother, a lettuce male fertile line is crossed as a pollen parent, and a hybrid first seed (F1 seed) is collected from the mother after the crossing. Lettuce F1 seed production method. Lettuce F1 seed obtained by the method according to claim 2 . The lettuce F1 seed according to claim 3 , which is deposited at the Patent Biological Deposit Center under the deposit number: FERM P-19542.

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