In Japan, tomatoes are produced all year round to meet the high demand for fresh tomatoes. They are cultivated in cool or highland areas under rain shelter in summer and in plastic houses during the cooler seasons. However, growing tomatoes in the same plastic houses all year round is not easy because plants lose vigor due to very high temperatures in summer. Therefore, some farmers try to cultivate tomatoes within a shorter period but more frequently per year than usual. In this cultivation system, only fruits in the first truss are harvested and the low yield per plant is compensated by frequent cropping. Thus, early flowering is a pre-requisite for an increase in fruit production.

Although tomato is an autonomous plant, flower initiation is directly or indirectly influenced by light, temperature, carbon dioxide, nutrition, moisture and growth regulators. However, no single environmental factor can be considered as critical for the regulation of flowering time in tomato. Furthermore, it is well-known that vigorous vegetative growth hinders reproductive development but the physiological and genetic background of this phenomenon is not clear. In this study, quantitative trait analysis (QTL) analysis was performed to explore the genetic control of flowering time as affected by growing season, temperature and mineral nutrition. QTLs controlling flowering time and related traits, such as number of leaves preceding the first inflorescence (LN), length of the largest leaf (LL), number of lateral shoots (LS), fresh weight (FRW), and plant height (PH), were characterized and mapped concurrently. Backross inbred lines developed from a cross between the commercial cultivar, S. lycopersicum and the wild species, S. pimpinellifolium were used as a mapping population. S. pimpinellifolium is the closest relative of the cultivated tomato and flowers earlier than the cultivated tomato.

1.QTLs controlling flowering time and related traits grown in spring and summer.

In January 2004, S. pimpinellifolium 'PI124039' (male) was crossed to S. lycopersicum cv. 'M570018' (female). The F1 was backcrossed to 'M570018' in May 2004, the latter being used as a male parent. The resultant BC1F1 plants were advanced by the single seed descent method until BC1F3 were obtained in June 2005. A linkage map was constructed using the 114 BC1F3 families, and a phenotypic evaluation was done in spring and summer using the resultant BC1F4 families.

The two parents differed significantly in all traits examined, except for PH. On the other hand, the growing season affected all traits significantly, except for LL. DTF was generally hastened in summer, with the parents and progeny flowering 7-10 days earlier than those grown in spring. Moreover, DTF was found to be highly correlated with all traits, except LS in summer.

Composite interval mapping detected 16 QTLs for the six traits evaluated in spring and summer, individually explaining 10%-42% of the phenotypic variation. In chromosome 1, a major DTF QTL was detected near marker C2_At5g49480, which accounted for about 40% of the phenotypic variation in spring with contributing alleles from S. lycopersicum. However, in summer this QTL was responsible for only 16% of the phenotypic variation, while the DTF QTL in chromosome 3 near marker C2_At5g51110 explained more of the variance for DTF (24%). The alleles derived from S. pimpinellifolium in chromosome 3 resulted in early flowering.

Most of the QTLs were co-located in a single locus in chromosomes 1 and 3, while the other QTLs were sparsely mapped in chromosomes 5, 7 and 10. The DTF QTLs in chromosome 1 were co-located with QTLs for LL, LS and FRW. This locus also corresponds to the same region to which a QTL was previously mapped for DTF, days to emergence and days to third leaf in a BC1 population derived from a cross between S. lycopersicum and S. pimpinellifolium. The number of days to third leaf is considered to reflect the rate of leaf initiation. On the other hand, in chromosome 3, the DTF QTL was found to be co-located with QTLs for LN, FRW (summer) and PH. Co-localization of DTF and LN QTLs have been also observed in previous studies, indicating that the same gene/s may be controlling these traits. These results suggest that the DTF QTLs in chromosomes 1 and 3 probably control the rate of leaf initiation and the period from the vegetative to reproductive stage, respectively.

2.QTL analysis of traits related to flowering time under low and high temperature regimes

The growing season was found to affect the flowering time of tomato. Thus, the effect of temperature was investigated in this experiment. The BC1F3 plants were selfed by the single seed descent method and BC1F5 seeds were obtained in July 2006. A linkage map was constructed using 105 BC1F5 families. For phenotyping, tomato plants from the resultant BC1F6 families were subjected to low (23/18oC) and high (30/25oC) temperature regimes for 20 days from sowing. After the temperature treatment, all plants were transplanted to the glasshouse in June 2007.

Results showed that S. esculentum differed significantly from S. pimpinellifolium in all traits evaluated. DTF, LL, FRW and PH were not significantly different between temperatures, while LN significantly increased at high temperature. The LN QTL in chromosome 7 was detected only at low temperature. It reduced the number of leaves preceding the first inflorescence, suggesting that low temperatures hastened the phase change from vegetative to reproductive growth. It has been previously reported that high temperatures stimulate flower development and hasten anthesis in tomato. Therefore, it is possible that the temperature treatments affect the time to flower initiation but flower development proceeds rapidly under high temperature treatments after flower induction, thereby masking the effects of the treatments on flower initiation.

3.QTLs controlling flowering time and related traits in tomato grown under low and high nutrient levels

Besides temperature, flowering time of tomato is affected by other factors, including mineral nutrition. Previous studies have shown that nutrient deficiencies prolong the vegetative phase, thereby delaying flower initiation. In this experiment, therefore, the genetic basis of flowering in tomato under two different nutrient levels was investigated.

The traits evaluated showed genetic variability in the parental lines. The nutrient treatments significantly affected the vegetative growth and reproductive development of the BC1F6 families. Flowering was delayed by 8-10 days in all genotypes under low nutrient condition. LN was not affected by the nutrient level, but nutrient deficiency reduced LL by almost 50%.

The QTL analysis detected a total of 24 QTLs. Six DTF QTLs were identified, half of which were detected in both nutrient treatments, while the other half were detected only in high nutrient levels. The major DTF QTLs in chromosomes 2 (near marker SSR32) and 3 (near marker C2_At5g51110) exerted more in high nutrient level than in low nutrient level, leading to early flowering in plants grown under high nutrient conditions. Moreover, the DTF QTLs co-localized with QTLs for other traits; with LL QTLs (low nutrient level) in chromosome 1, LN and PH QTLs in chromosome 2, LN (low nutrient level) and PH QTLs in chromosome 3 near marker SSR231, and with LN and PH QTLs in chromosome 3 near marker C2_At5g51110. The co-location of major QTLs for DTF and LN in chromosomes 2 and 3 in both nutrient levels further suggests that the same genes control these traits. On the other hand, 7 QTLs (1 LN, 3 LL, 1 LS, and 2 PH) were detected only in low nutrient level, whereas 6 QTLs (3 DTF, 1LL, 1FRW and 1 PH) were detected only in high nutrient level. No LL QTLs were common in both nutrient treatments, confirming previous reports that leaf elongation and shoot growth are highly responsive to differences in nitrogen levels.

In conclusion, a total of 8 DTF QTLs were detected in chromosomes 1, 2, 3, 7 and 12. Major DTF QTLs were detected in chromosomes 1, 2, and 3, while 6 DTF QTLs were found to be co-located with QTLs for other traits, indicating the presence of loci with pleiotropic effects on these traits. Most of the QTLs identified in this study were conferred by the S. pimpinellifolium alleles, indicating the great importance of this wild species in the genetic improvement of the cultivated tomato.

わが国の夏は高温過ぎるため,一年を通じて同一のハウスでトマトを栽培することは容易ではない。そこで,第一花房で収穫を打ち切り,代わりに作付け回数を増やして年間の収量増をはかる一段密植栽培が注目されている。この栽培方法では開花までの期間を短縮することが重要な要件となる。

トマトは中生植物であるが,花芽分化には光,温度,養分などの要因が影響を及ぼす。また,過度の栄養成長は生殖成長を阻害することが知られているが,これらの現象の生理的,遺伝的背景については明らかになっていない。そこで,本研究ではトマト栽培種と近縁野生種のS. pimpinellifoliumの雑種第一代に栽培種を戻し交配した系統を用い,量的形質遺伝子座 (QTL) 解析によって栽培季節,温度,栄養条件と遺伝子の交互作用が開花時期や栄養成長に及ぼす影響について調査した。調査した形質は,開花時期,第一花房下葉数,最大葉長,腋芽数,生体重,草丈の6形質である。

春と夏に栽培したトマトにおける開花時期のQTL並びに関連形質

S. pimpinellifoliumとトマト(反復親)の BC1F3 世代114系統を作出し, BC1F4 世代で春と夏に形質評価を行った。夏は春に比べて,開花までの日数が7-10日早かった。開花までの日数は夏の腋芽数を除き,他の形質と有意な相関が認められた。複合区間マッピング法によって,合計16のQTLが検出された。第1染色体のC2_At5g49480マーカーの近傍に,開花までの日数に関するQTLが検出された。このQTLは,最大葉長,腋芽数,生体重と同一の位置に座乗していた。一方,第3染色体のマーカーC2_At5g51110 近傍にも開花までの日数に関するQTLが検出された。このQTLは,第一花房下葉数,生体重 (夏),草丈のQTLと同じ位置に座乗していた。さらに,このQTLは既に明らかにされている出芽までの日数や第3葉出現までのQTLと同じ位置に存在することが明らかになった。これらの結果から,開花日数の第1,第3染色体上のQTLはそれぞれ,葉原基の分化速度と栄養成長から生殖成長への転換時期を支配していると考えられた。

異なる温度条件下における開花時期に関連する形質のQTL解析

作季の違いがトマトの開花時期に影響を及ぼすことが明らかになったので,夏に BC1F6 105系統を昼30,夜25°C(30/25°C)と23/18°Cの条件下で20日間栽培した後,ハウスに移して開花まで栽培した。 連鎖地図はBC1F5 105系統群を用いて作成した。第一花房下葉数のみは高温下で有意に増加したが,開花までの日数,最大葉長,生体重,草丈については温度の影響は認められなかった。本実験では,開花までの日数に関するQTLは検出されなかった。第一花房下葉数に関するQTLが第2,3,7染色体上に検出された。これらQTLの中,第7染色体上のQTLは低温でのみ検出された。この結果は,低温が栄養成長から生殖成長への転換を早めることと一致した。高温下では花の発達が促進され,開花時期が早まることが知られているので,本研究で開花時期のQTLが検出されなかったのは花芽分化後の高温のために花芽分化に及ぼす温度の影響が遮蔽されてしまった可能性が考えられた。

栄養条件の違いが開花時期を調節するQTL に及ぼす影響

これまでの研究において,養分欠乏下では栄養成長の期間が延び,開花が遅れることが報告されている。そこで,施肥量を変えてBC1F6系統群を栽培し,栄養条件の違いがQTLに及ぼす影響を調べた。養分欠乏下で開花は8-10日遅くなった。また,第一花房下葉数は施肥量の影響を受けなかったが,最大葉長は養分欠乏下で50%低下した。検出された24のQTL中6つが開花時期に関するQTLで,養分供給が十分な場合,第2,第3染色体上のQTLが開花に大きな影響を及ぼすことが明らかになった。さらに,開花に関連するQTLは最大葉長,花房下葉数,草丈などのQTLと同一遺伝子座に座乗していることが明らかになった。

以上要するに,本研究では,開花日数に関連するQTLとして第1,2,3,7,12染色体上に計8QTLを検出したが,このうち第1,2,3染色体上に座乗する6QTLは花房下葉数,最大葉長,草丈などのQTLと同一遺伝子座またはその近傍に座乗していることを明らかにし,今後のトマト育種に多くの知見を与えたもので,実用上,応用上資するところが少なくない。よって,審査委員一同は,本論文が博士(農学)を授与されるに相応しいと認めた。