WO1999031242A1

WO1999031242A1 - Plants with raised phytochrome expression

Info

Publication number: WO1999031242A1
Application number: PCT/EP1998/008092
Authority: WO
Inventors: Christiane Gatz
Original assignee: Kws Saat Ag
Priority date: 1997-12-12
Filing date: 1998-12-11
Publication date: 1999-06-24
Also published as: EP1037996A1; AU1967299A; DE19755101A1

Abstract

The invention relates to a plant characterized in that it overexpresses phytochrome. The invention relates particularly to a plant which overexpresses phytochrome through the introduction of a phytochrome B gene into said plant. The invention further relates to plant parts, seed and/or plant cells which, like the plant, are characterized by phytochrome overexpression. The invention also relates to the use of one and/or several phytochrome genes for the production of plants which overexpress phytochrome. The invention further relates to a method for growing a plant which overexpresses phytochrome or parts of plants, seed and/or cells which overexpress phytochrome. Lastly the invention relates to a vector containing a phytochrome gene which codes for an amino acid sequence the N-terminal of which is rich in glycine.

Description

Plants with increased phytochrome expression

The present invention relates to a plant which is characterized in that it overexpresses phytochrome. In particular, the present invention relates to a plant that overexpresses phytochrome B by introducing or activating a phytochrome B gene in the plant. The invention further relates to parts of plants, seeds and / or plant cells which, like the plant, are characterized by phytochrome overexpression. The invention also relates to the use of one and / or more phytochrome genes for the production of plants which overexpress phytochrome. The invention further relates to a method for producing a plant which overexpresses phytochrome or parts of plants, seeds and / or cells which overexpress phytochrome. Finally, the invention relates to a vector which contains a phytochrome gene which codes for an amino acid sequence whose N-terminus is hydrophobic.

The development of plants is controlled in many ways by light from germ to flower formation. Germination and greening is triggered by light. Plants can use the energy of sunlight in their green tissues to convert carbon dioxide (CO ₂ ) into high-energy organic compounds. This process, in which sugar molecules initially form, is called photosynthesis. A large part of the photosynthetically stored energy is used by the plant for growth and other metabolic processes, but a certain part can also be preserved in storage organs.

Since plants are absolutely dependent on light as an energy source, their development is controlled by the environmental factor light in such a way that they can optimally use the light. For example, seedlings only form the structures that are important for photosynthesis when they come to light. Up to this point, they prefer to grow in length at the expense of their reserve materials in order to reach the light as quickly as possible. Under shade conditions, adult plants also use a large part of their energy to stretch the stem in order to escape the shade. Light is perceived by chromoproteins, which serve as photoreceptors. Plants have three different photoreceptor systems that absorb light of different wavelengths. One of these is the phytochrome photoreceptor system, which absorbs in the red light region of the spectrum.

Phytochromes are chromoproteins that consist of a 124 kDa protein component and an elongated tetrapyrrole system as a chromophore (Kendrick and Kronenberg, 1986). Phytochromes are synthesized in the dark in the so-called Pr form, which is physiologically inactive. By absorbing light with a wavelength of 660 nm, they change into the physiologically active Pfr form, which controls a variety of functions in the plant. It is characteristic of the phytochromes that they can be converted back into the Pr form by irradiation with dark red light. Phytochrome-dependent physiological processes include:

• Formation of proteins that are involved in the photosynthetic performance of plants.

• Shortening the internodes to adapt the plants to shade conditions. (Under shadow conditions, the red light component of the sunlight is absorbed by the canopy, the less strongly absorbed dark red light causes a conversion of Pfr to the inactive Pr form, which results in an inter- node stretch.)

• Formation of anthocyanins

• Induction of flower or bulb formation

• Delay in senescence

The first phytochrome gene was cloned and sequenced in 1985 (Hershey et al., 1985). It was phytochrome A from oats, which was particularly abundant in seedlings grown in the dark. Based on an abundance of physiological studies, the existence of further phytochromes was suspected, but their isolation was very difficult because of their small amount. It was only through molecular techniques that the unique one succeeded Detection of further phytochromes. The molecular identification of distinct phytochrome genes (PhyA, PhyB, PhyC) was carried out in 1989 by sequencing various cDNAs from Arabidopsis thaliana (Sharrock and Quail, 1989), which was supplemented by the publication of the PhyD and PhyE genes in 1993 (Clack et al., 1993). PhyA, PhyB, PhyC and PhyE are identical in approx. 50% of their amino acid positions, PhyD with 70% amino acid identity is closer to PhyB than to the other phytochromes. The presence of distinct phytochrome genes could then be demonstrated in other plants. There is a phytochrome gene in the potato that is 76% identical to PhyA from Arabidopsis and was therefore classified as type A (Heyer, 1992). A gene homologous to PhyB from Arabidopsis could also be isolated from the potato (Heyer, 1992). However, from the presence of the five different phytochromes in Arabidopsis, one cannot infer the composition of the phytochrome family in other plants. A phytochrome gene was found in the tomato, which is closely related to the potato, the sequence of which cannot be grouped into classes A to E (Pratt et al., 1997). Therefore it is called PhyF.

In 1989 a cDNA for the one phytochrome (phytochrome A from oats) was first introduced in tobacco. This led to an increased synthesis of this phytochrome in the transgenic plants. It was found that plants react very sensitively to the increase in phytochrome. With strong expression of the phytochrome gene (five times increased, measured in etiolated seedlings), the following phenotypic changes appeared

• Reduced apica dominance

• Dwarfism (semi-dwarfism)

• Increased chlorophyll content per leaf area

• Smaller but thicker leaves

• Deformed chloroplasts

• Worsened diffusion of CO ₂ through the mesophyll cells

• Reduced photosynthesis rate

• Increased enzymatic activity of enzymes involved in photosynthesis (based on leaf area) • Less root mass

• Delayed senescence

Of the generally desirable goals when growing plants, these plants showed dwarfism, increased chlorophyll content per leaf area, thicker leaves, increased activity of enzymes that are involved in photosynthesis (based on leaf mass). However, they were inferior to conventional plants in some respects: they had deformed chloroplasts, which led to a deterioration in the diffusion of CO ₂ through the mesophyl cells and to a lower rate of photosynthesis. The deformed chloroplasts and the reduced photosynthetic performance were such serious defects that the other effects no longer come into play. The overall performance of the plants was no better than that of the original plants. In addition, the root mass was less pronounced than in conventional plants. They were inferior to conventional plants in terms of the increased formation of underground organs (here roots) and the photosynthetic performance. With lower expression (2-3 times increased, measured in etiolated seedlings), however, they showed a property that leads to an improvement in the so-called harvest index, especially in dense planting (harvest index: ratio of the amount of the desired product (e.g. seeds, fruit, Leaf, tuber) to the total biomass). In many crop plants, the mutual shading of the plants when densely planted causes the plants to grow in length in order to have an advantage over the neighboring plants competing for light. In the field, all plants grow in length, which is at the expense of the growth of other organs. However, the increased amounts of phytochrome A cause a shortening of the intemodes. This means that leaf growth is more pronounced in relation to stem growth and there is an increase in yield (leaf / ha) of 15-20%. These data were obtained with tobacco plants in which the leaves are the valuable raw material. An increased photosynthesis rate, delayed senescence and enlargement of the root ball was not documented in these low-expressing plants. In 1991, phytochrome B proteins were overexpressed in transgenic plants in an increased amount (18-30 times, measured in green tissue) (Wagner et al., 1991). It was phytochrome B from Arabidopsis, which was overexpressed in Arabidopsis. A shortened growth of the hypocotyl was described, later the influence on phototropism and flowering induction was described. An effect on the goals of increased photosynthesis rate, delayed senescence, enlargement of the root ball, increased yield was not observed. Arabidopsis phytochrome B was also overexpressed in tobacco. Dwarf growth and an influence on the induction of flowering have been observed. An influence on photosynthesis, root mass, yield or senescence has not been documented (Halliday et al.).

Accordingly, it is an object of the present invention to provide plants or parts of plants, seeds and / or plant cells, a) whose photosynthetic performance is increased and / or b) whose aging is delayed, as a result of which the solar energy is usable for longer, which is reflected in an increase in yield and / or c) in which the root ball is enlarged, which leads to better nutrient absorption and / or d) whose yield is increased.

In particular, it is the object of the present invention to specify plants, parts of plants, seeds and / or plant cells in which these four goals can be achieved in combination.

These tasks are solved by the objects mentioned in the independent claims. Preferred embodiments of the invention are specified in the subclaims.

According to claim 1 there is provided a plant which is characterized in that it overexpresses phytochrome.

Some of the terms used in the application are explained in more detail below in order to clarify how they are to be understood here. “Phytochromes” are understood to mean all those chromoproteins which consist of an approximately 124 kDa protein portion and an elongated tetrapyrrole system as a chromophore and which serve as photoreceptors in the plant.

“Overexpression” is understood to mean any increase in the phytochrome level in the plant, be it that it may be due to the introduction of a strong promoter together with the phytochrome gene, or to gene amplification or other possibilities known in biotechnology to increase the protein synthesis performance of a cell . Overexpression occurs when the modified (overexpressing) plant produces at least twice the amount of phytochrome, in particular PhyB, as the original plants. This also includes double the amount in just one part of the plant, e.g. the sheet. The modified plant preferably produces 5 times, possibly 10 times, the respective phytochrome in comparison to the original plant or a part of the original plant.

“Phytochrome gene” means any gene that codes for a phytochrome as defined above.

By "hydrophobic" is meant an amino acid sequence that has more hydrophobic than polar amino acids, e.g. 10 alanines per 20 amino acids.

"Glycine-rich" means an amino acid sequence which has more than one glycine per four amino acids and is preferably at least 20 AA long, for example 12 Gly within 30 AA.

The invention describes that the stated goals can be achieved if phytochrome, in particular phytochrome B, is overexpressed in a plant. Surprisingly, the best results were obtained when the phytochrome was introduced into a plant that belongs to a different species than that Plant from which the gene came. This is shown in the examples in particular when the phytochrome B protein from Arabidopsis thaliana is synthesized in increased amounts in crop plants. This was explicitly shown using the example of the potato. The plants are characterized by a 30% higher phytosynthesis rate, which is also significantly more sensitive to light stress than the photosynthesis rate of conventional plants with normal phytochrome levels. Senescence is delayed. The underground shoot root system is at least tripled. Furthermore, when the number of tubers doubles, the plants show an increase in yield of 15-20%.

In contrast, the increased expression of phytochrome B from Solarium tuberosum showed less clearly the increased development of the root ball, the increased photosynthesis rate and the delayed senescence. An obvious difference between the phytochromes B from A. thaliana and S. tuberosum (amino acid identity: 76%) is in the different N-terminus. As shown below, the range between the amino acids VSGV and QAQSS is very different. The phytochrome B from A. thaliana codes for a glycine-rich region that is missing in the phytochrome B from Solanum tuberosum.

At. MVSGVGGSGGGRGGGRGGEEEPSSSHTPNNRRGGE QAQSS SeqlD Nr.1 S.t. MASGSRTKHSHHSS QAQSS SeqlD No.2

A sequence comparison of the N-termini of the B phytochromes of A. thaliana and S. tuberosum is shown above.

The objects of the present invention can be achieved with types of phytochrome B and other phytochromes especially when the phytochromes have a hydrophobic N-terminus. A glycine-rich N-terminus is particularly advantageous. The N-terminus described above has proven to be particularly advantageous. However, included are sequences which are derived from the N-termius of A. thaliana and show the same effects when introduced into plants as the N-terminus of PhyB from A. thaliana. Monocotyledonous, dicotyledonous useful plants or ornamental plants are particularly advantageous as starting plants for the present invention. This applies in particular to corn, wheat, millet, oat, rice, barley, sorghum, amaranth, onion, asparagus, sugar cane, alfalfa, soybean, petunias, cotton, sugar beet, sunflower, carrot, celery, cabbage, cucumber, pepper, canola, Tomato, potato, lentil, flax, broccoli, tobacco, bean, lettuce, rapeseed, cauliflower, spinach, Brussels sprouts, artichokes, peas, okra, pumpkin, kale, collard green, tea and coffee, geraniums, cloves, orchids, roses, impatiens , Petunias, begonias, fuchsias, yolk flowers, chrysanthemums, gladiolus, astromeria, sage, veronica, daisies and iris.

A particularly advantageous effect results when using a phytochrome B gene with the N-terminus given above for the production of a transgenic potato. In a particular embodiment of the invention, in order to produce a transgenic maize plant with delayed senescence, a phytochrome B gene coding for a protein with the N-terminus specified above is introduced into a maize plant. As a result, a “stay green” effect can be achieved in the transgenic plant.

The phytochrome B gene coding for a protein with the N-terminus indicated above can also be used to produce a transgenic sugar beet plant with advantageous properties. For example, by introducing this phytochrome B gene into a sugar beet plant, a delayed senescence and thus a reduction in nitrogen storage can be achieved.

The statements made above for the plants which overexpress phytochrome also apply to the plant parts, seeds and / or plant cells which overexpress phytochrome. In particular, the desired results can already be achieved, for example, by tissue-specific overexpression of phytochrome in the leaf. It is possible that one does not want to have the phytochrome overexpressed in certain parts of the plant. For example, the increased number of lines rather undesirable; the same number of tubers with increased yield would be advantageous. This can be achieved, for example, by not expressing the phytochrome in the whole plant, but only in the leaves. With a constitutive promoter, overexpression has so far been found in the shoot tip, and this part of the plant controls the number of tubers.

Phytochrome gene activation can also lead to the desired result. Plant-specific phytochrome genes are activated by introducing a strong promoter and / or enhancer.

The invention also relates to the use of one and / or more phytochrome genes for the production of the plants and / or parts of plants, seeds and / or cells described above, a method for producing transgenic plants, a construct which comprises a phytochrome as described above -Gen contains, as well as a plant containing a construct comprising a phytochrome gene and a glycine-rich N-terminus.

In the following, the invention will be explained in detail with reference to the examples and drawings, but it should be noted that these are only special embodiments, which are only exemplary and are not intended to limit the scope of the invention.

The following is shown in the figures:

Figures 1A and 1B: shoot habit of an untransformed control plant of potato (A) compared to a representative transgenic potato plant of the DARA12 line (B). The plants come from a sterile culture and had been grown in the greenhouse for 8 weeks.

Figure 2: Western analyzes with antibodies against phytochrome B from Arabidopsis thaliana with protein preparations from leaves of untransformed control potato plants (wt) in comparison to those of the transgenic lines DARA5 and DARA12. Figures 3A and 3B:

(A) Photosynthesis rate (μmol CO ₂ / m ² s, at 1, 2-1, 8 mmol photons / m ² s) of leaves of untransformed control plants of potato (WT) compared to that of leaves of transgenic plants of the DARA12 line.

(B) Decrease in the photosynthesis rate of the leaves from A after exposure for 5.5 hours under 1, 2-1, 8 mmol Ph./m ² s and 32-38 ° C. The data come from n = 6 measurements of 12 week old greenhouse plants.

Figure 4: Underground shoot-Λ / root system of representative potato plants of the untransformed control line (WT) and those of the transgenic lines DARA5 and DARA12. The plants were exposed to sterile culture and cultivated in the greenhouse from October 1996 to April 1997. At the time of harvest, they were 6 months old.

Figures 5A and 5B: Underground shoot / root system and tuber yield of representative potato plants of the untransformed control line (WT) compared to those of the transgenic plants of the lines DPOT1, DPOT9 (A) and DS13 and DS23 (B). The plants were exposed to sterile culture and cultivated in the greenhouse from October 1996 to April 1997. At the time of harvest, they were 6 months old.

Figure 6: Westem analyzes with first antibodies against phytochrome B from Arabidopsis thaliana with phytochrome preparations from leaves of untransformed control potato plants (Wt) compared to those of the transgenic lines DPOT1, DPOT9, DS13 and DS23.

METHODS

1. Transformation of potatoes (Rocha-Sosa et al .. 1985)

30 μl of an overnight culture of the A. tume faciens strain GV2260 carrying T-DNA plasmid, grown in selection medium (YEB + kanamycin (50 mg / l) and rifampicin (50 mg / l)), were mixed with 10 ml of liquid MS medium (MS medium: Murashige and Skoog, 1962). Leaflets from the sterile tissue culture were cut at the midrib and transferred to the inoculated MS medium and incubated dark at 22 ° C. for two days. Subsequently, the leaf disks for callus induction on MS medium with glucose (16 g / l), naphthylacetic acid (5 mg / l), 16-benzylaminopurine (0.1 mg / l), cephotaxime (250 mg / l), Bitec Incubate agar (6.3 g / l) and 1 mg / l hygromycin or 50 mg / l kanamycin for selection for seven days under tissue culture conditions. Subsequently, the leaf disks for the regeneration of the shoot on MS medium with glucose (16 g / l), zeatin ribose (1.4 mg / l), naphthylacetic acid (20 μg / l), gibberellic acid (GA ₃ ; 20 μg / l), cephotaxime ( 250 mg / l), Bitec agar (6.3 g / l) and 3 mg / l hygromycin or 50 mg / l kanamycin incubated under tissue culture conditions. The leaf disks were transferred to new regeneration medium after seven days. The resulting shoots were cultivated on MS medium with 2% sucrose, selective antibiotics (50 mg / l kanamycin or 3 mg / l hygromycin B) and 250 mg / l cephotaxime in tissue culture.

YEB medium: 2 g / l yeast extract

5 g / l beef extract 5 g / l peptone 5 g / l saccarose 2 mM MgSO ₄ pH 7.0

2. Plant culture

The plants were cultivated in the experimental greenhouse of the botanical garden at the University of Göttingen. They were grown from tubers or sterile culture (medium according to Murashige & Skoog, 1962). During cultivation, the temperature in the greenhouse was 15-25 ° C in the winter half-year (October - April) and 20-35 ° C in the summer half-year (May-September). Between day and night it fluctuated by 5 - 15 ° C. The relative air humidity was around 55% all year round. Depending on the age and therefore the height of the plant bodies, the light intensity was between 200 and 450 μmol photons (m ² s) ^"1 during the day, and a maximum of 500 μmol on clear days Ph. (M ² s) ^"1. The day lengths corresponded to the natural seasonal period. During the 4 to 6 month cultivation, the plants were transferred into larger pots at intervals of 2-6 weeks (up to a maximum of 20 cm pot diameter In order to obtain plants of comparable strength, all were cut back to the two strongest above-ground shoots 3 weeks after the bulbs had emerged or were exposed to sterile culture, due to their binding up and a pot spacing that corresponded at least to the respective pot diameter largely uniform light exposure guaranteed.

3. Preparation of phytochromes from plants (van Tuinen et al .. 1995)

0.3 g of fresh leaf material was pulverized with 0.03 g of polyvinylpyrrolidone under liquid nitrogen in an Eppendorf reaction vessel in a homogenizer and taken up in 300 μl of phytochrome extraction buffer. After 15 min centrifugation at 15,000xg and 4 ° C, 300 μl of the supernatant were removed, 30 μl of polyethyleneimine (10% w / v in H ₂ O) were added and 15 min at 4 ° C in an Eppendorf shaker incubated at maximum speed to precipitate nucleic acids and chlorophyll. After centrifugation at 12,000 × g and 4 ° C. for 10 minutes, the supernatant was mixed with 0.725 vol. Of an ammonium sulfate solution saturated in water and incubated for 15 minutes at 4 ° C. at full power in an Eppendorf shaker. After 15 min centrifugation at 12,000xg and 4 ° C, the sediment was taken up in 30 ul "Laemmli" buffer and incubated for 2 min at 100 ° C. For western blot analysis, 20 μl of the sample were loaded onto a 7% PAA gel (Laemmli, 1970).

Phytochrome extraction buffer:

Ethylene glycol 50%

EDTA 10 mM

Tris-HCl, pH 8.3 100 mM

add fresh: Na bisulfite 20 mM

(NH ₄ ) ₂ SO ₄ 140 mM β-mercaptoethanol 56 mM

Pefablock 4 mM

Iodoacetamide (IAA) 4 mM

Pepstatin A 1 μg / ml

Aprotinin 2 μg / ml

Leupeptin 2 μg / ml

4. Immunulogical detection of phytochrome B by Western analysis

For the immunological detection, the proteins separated in the PAA gel were transferred to nitrocellulose or PVDF membranes. For this purpose, the PAA separating gel was equilibrated in electroblot transfer buffer for 15 min. Three layers of Whatman paper soaked in transfer buffer, the membrane, the gel and three further layers of Whatman paper soaked in transfer buffer were placed on the anode plate of the BioRad electroblot apparatus without air bubbles. The cathode plate was placed under slight pressure and the electroblot was carried out at 15 V for 45-120 min. Nitrocellulose membranes were equilibrated in transfer buffer for 10 min, PVDF membranes were first soaked in methanol and then equilibrated in transfer buffer for 10 min. The transfer was checked with Ponceau red - a dye that can be removed with water.

Electroblot transfer buffer: PAG E-Lauf buffer with halved SDS concentration with 20% methanol

Ponceau staining solution: Ponceau red 10 drops

Acetic acid 10 drops

H ₂ O 100 ml

Immunological detection:

All steps were carried out at room temperature with gentle shaking. The filters were blocked as follows to saturate non-specific binding sites: TBSI 3 min

TBSII 10 min

TBSII 5 min.

The first antibody was then diluted 1: 400 (polyclonal antibody on rabbit) or 1: 5000 (monoclonal mouse antibody) in TBSIII with 1% skim milk powder (w / v) and incubated for 2 h. After washing 3 × 10 min with TBSIII, the filter was incubated for 90 min with the second, biotinylated antibody (1/1000 Amersham anti-mouse or anti-rabbit) in TBSIII + 1% (w / v) skim milk. After washing 3 × 10 min with TBSIII, the filter was equilibrated in TBSIII (without Tween). The filter was then incubated for 30 min with streptavidin-coupled horseradish peroxidase (1/500, Amersham) in TBSIII (without Tween). After washing 4 × 5 min in TBS, the proteins recognized by the first antibody can be stained on the filter or developed on X-ray film via chemiluminescence.

TBSI buffer: Tris-HCl, pH 8.3 20 mM NaCI 500 mM Tween 20 0.05% (w / v)

TBSII buffer: Tris-HCl, pH 8.3 20 mM NaCI 150 mM Tween 20 0.05% (w / v)

TBSIII buffer: Tris-HCl, pH 8.3 20 mM NaCI 150 mM Tween 20 0.05% (w / v)

Staining

The filter was developed in the following color reaction solution:

Diaminobenzidine 10 mg

TBS 20 ml

CoCI ₂ 0.01% (w / v)

Loosen for 20 min add fresh: H ₂ O ₂ (30%) 30 μl

The color reaction was stopped with water.

Chemiluminescence

For chemiluminescence detection, the filter (PVDF membrane) was incubated for 1 min in reaction solution (Amersham peroxidase chemiluminescence solutions 1 and 2 in the ratio 1: 1) and exposed to X-ray film in an exposure cassette for 2 to 10 min. The X-ray film was then developed

5. CQo assimilation using infrared spectroscopy

The activity of the dark photosynthetic reaction was determined by means of infrared spectroscopy via the rate of CO ₂ fixation. For this purpose, a portable gas exchange porometer from ADC (Hoddesdon, Great Britain) was used, consisting of the central LCA-3 analysis unit and a PLC-3 sheet chamber (diameter 6.2 cm ² ) with a thermal sensor and light sensor for simultaneous detection of the internal cell temperature and photon flux density. An integrated PC was used for data storage and data processing. This system made it possible to determine the assimilation rates achieved in different culture conditions (see 1.1. And 1.2.) (In μmol (m ² s) ^'1 ).

5.1 CO assimilation of selected leaf areas of unstressed control plants In order to ensure comparability of the data obtained, only leaves of approximately the same age and similar light exposure were used. All examinations were carried out with the end of the 3rd to 5th adult leaves (starting from the apex). The leaf leaflets, without separating them from the plant and keeping their orientation to the light, were clamped into the PLC-3 chamber and adapted immediately before each measurement for the purpose of equilibration of gas and temperature in the measuring system for 10 min. In the course of the following 10 minutes, the current rate of CO ₂ fixation, the temperature and light intensity were recorded every 2 minutes. 5.2. Light stress analysis

In order to analyze their sensitivity to light stress, the plants were exposed to photon flux densities of between 1.2 and 1.8 mmol (m ² s) ^"1 for 5.5 hours after determining the assimilation rates (2.1.) (Halogen lamp from Osram , Power Star HQI-T / N, 2000 W.) During this time, the temperature in the cuvette rose to 32-38 ° C. In the course of the following 10 minutes, the current rate of CO ₂ fixation was the temperature and light intensity registered.

δ.S. COo-assimilation of whole plants

The following procedure was used to estimate the overall assimilation performance of individual plants and thus their potential for starch synthesis and ultimately tuber production. The average total leaf surface per plant was determined for each line using the total mass of the leaves of representative individual plants (each n = 3) of a line and their average mass per leaf area (measurements with n = 18 leaf disks each). With the help of 3.1. determined CO ₂ assimilation rate per area, it was possible to infer the approximate assimilation rate (μmol s ^"1 ) of an entire plant. These determinations were carried out with 3-month-old, non-senescent plants.

6. Piomentanalvsen

6.1. Photometric determination of the chlorophyll-containing leaf disks (1.33 cm ² ) were pulverized in liquid nitrogen and the chlorophylls were extracted quantitatively with 80% ethanol. The extracts were centrifuged at maximum speed and the absorbances of the supernatants were determined photometrically at 645, 652 and 663 nm using a Uvikon 932 spectrophotometer (from Kontron, Neuzahrn). The chlorophyll contents were determined according to the following formulas.

Total chlorophyll (μg) = E ₆₅₂ x dilution factor x (34, 5) ^"1 chlorophyll a (mg / ml) = [E ₆₆₃ x 45.6 - E ^ x 9.25 x (3858 x ml aliquot) ^{" 1} ] x 1, 16 x ml vol Chlorophyll b (mg / ml) = [E ^ gx 82.04 - E ₆₆₃ x 16.75 x (3858 x ml aliquot) ^"1 ] x 1, 07x ml vol

6.2. HPLC analysis for the quantification of the photosynthetic pigments. The leaf disks (1.33 cm ² ) were stored in the dark in liquid nitrogen until they were worked up. The pigments were extracted according to Thiele et al. (1996). For the separation and detection of the carotenoids and chlorophylls, an HPLC device from Hitachi (Lichrograph; Merck, Darmstadt) was used, consisting of the gradient pump L-7100, the injection valve 7125 (Cotati, CA, USA), the UV detector L- 7400, the solvent degasser Gastorr 104 (Schambeck, Bad Honnef), the integrator L-7500 and a reversed phase LiChroCART column with LiChroCART guard column (250-4 or 4-4; both Merck). The column filling was formed by LiChrospher 100RP-18 (5 μm; Merck). (A) Acetonitrile: methanol: Tris / HCl (0.1 M, pH 8), 7: 10: 3, (B) methanol: hexane, 4: 1 were used as solvents. First, solvent A eluted, then via a linear gradient built up in 3.5 min (0-100%) with B. After 5.5 min isocratic elution with B, a linear gradient followed back to 100% A, whereupon the column up to the subsequent separation passage for was re-equilibrated in A for a further 4 min. The flow rate was 2 ml (min) ^"1. This method enabled the clean separation of neoxanthine, violaxanthin, antheraxanthin, lutein, zeaxanthin, α- and ß-carotene, and chlorophylls a and b. The quantification of these pigments was carried out according to Färber et al. (1997).

6.3. Photometric determination of UV-absorbing substances (e.g. Anthocvane)

The relative absorbance (% of the wild type) of ethanolic tissue extracts at 284 nm (see Garden, 1997), measured with the aid of a Uvikon 932 spectrophotometer (from Kontron, Neuzahm), served as a rough measure of the anthocyanin contents. At this wavelength, both the leaf extracts and the stem epidermis showed clear absorption maxima. The extraction was carried out as in 4.1. described. 7. Detection of dry matter and starch

The relative dry weights and starch contents of the tubers were determined indirectly as follows via the specific weights of the tubers in air (W _L ) and in water (W _w ) (Von Scheele et al., 1937).

Density δ = W _L (W _L -W _w ) ^"1

% Dry matter = (δ - 1, 0988) x (211, 04 + 24.182)

% Strength = (δ - 1, 0988) x (199.07 + 17.546)

8. Production and staining of leaf microtome sections for light microscopy

As for the studies on CO ₂ assimilation (see 3.1.), Only leaves of similar age and similar light exposure were used for direct anatomical comparability. The leaves were from 10 week old plants and showed no external signs of senescence. Central segments (1 cm distance from the main control line) were taken from the blade of 4 leaf leaflets from different plants per line. The production and staining of the sections followed the paraffin method (modified according to Gerlach 1969, personal communication, G. Weis, University of Göttingen), with Histosec (Merck) as an embedding medium and using a slide microtome (from Leitz, Wetzlar). Saffron red (chroma) primarily stained chloroplasts and cell nuclei, Astra blue (Merck) mainly stained cell walls and mucus.

EXAMPLES

1. Production of transgenic plants

The coding region of Arabidopsis thaliana phytochrome B (At phyB) was amplified by the polymerase chain reaction using the cDNA clone in IEMBL3 (Somers et al., 1991) as a template. The primers contained Kpnl interfaces. At phyB can thus be cloned as a Kpnl fragment into suitable expression vectors. In the example used here, it was brought under the control of the CaMV 35S promoter (Benfey et al., 1990), which is responsible for the expression of the Gene leads in all parts of the plant. The polyadenylation region of the nopaline synthase gene from Agrobacterium tumefaciens served to terminate the transcription. Genes in which the promoter, the coding region and the polyadenylation signal originate from different genes are referred to as chimeric genes. The construct is therefore referred to below as a chimeric At phyB gene. The techniques for producing such recombinant DNA are described in Sambrook et al. (1989). The chimeric At phyB gene was transformed via Agrobacterium tumefaciens-mediated gene transfer in potato plants (Rocha-Sosa et al., 1985). Transgenic plants that express phytochrome B from Arabidopsis thaliana are easily recognized by their dwarfism, reduced apical dominance, and darker green leaves (Figure 1, drawing). Two independent transformants, hereinafter abbreviated to DARA5 and DARA12, were characterized as follows.

Table 1

Shoot anatomy of the transgenic potato lines DARA5 and DARA12 compared to the untransformed control line (wild type). The shoot height indicates the distance between the soil and the apica bucket, the stem diameters were determined after harvesting the above-ground rung at the stem base (1 cm above the ground). The data come from 3.5-month-old greenhouse plants grown from tubers (July-October), n = number of individual measurements.

TABLE 1

2. Determination of the content of phytochrome B

In contrast to structural proteins, regulatory proteins are only expressed in small amounts in plants. The detection of phytochromes is often performed munologically using antibodies. Monoclonal antibodies against phytochrome B from A. thaliana are available (Somers et al., 1991). These antibodies also recognize the phytochrome B from S. tuberosum. The exact implementation of this proof is described in the method section. Figure 2 shows the result of the Western blot analysis. In comparison to the untransformed control (Wt) the lines DARA5 and DARA12 show a significantly stronger signal. The comparative quantification using dilution series showed that DARA5 synthesizes about five times more phytochrome B, DARA12 about 12 times more, compared to the untransformed control (wt).

3. Increase in the rate of photosynthesis

Cultivation conditions and the technique of measuring the phyotosynthesis rate are described in the method section. Figures 3A and 3B show the result of the measurements as an example for the DARA12 plant. The photosynthetic performance of greenhouse plants (transgenes and control plants) which had been grown in the greenhouse for 10 weeks was determined by measuring the CO ₂ assimilation rate. The plants were initially left at a light intensity of 0.3 mmol photons / m ² s for a few hours. Thereafter, photosynthetic performance under 1.5 to 1.8 mmol photons / m ² s was analyzed by measuring the CO ₂ uptake. In this measurement, the untransformed control plant showed an average CO ₂ assimilation rate of 17.5 mmol / m ² s, while the transgenic plant (DARA12) was increased by 28.5% with 22.5 mmol / m ² s. If these values are converted to the CO ₂ absorption capacity of whole plants, the performance per plant is increased by approx. 20%.

The diagram shown in Figure 3B shows that the performance of photosynthesis is increased by more than 30% in the transgenic plants compared to the conventional plants when light intensity is constantly high, as occurs in summer with a clear sky. The plants just described were kept at 1.2-1.8 mmol photons / m ² s for a further 5 hours, then the photosynthesis rate was measured again. This was in the transformed control plants reduced from 17.5 μmol CO ₂ / m ² s to 3 μmol CO ₂ / m ² s, while in the transgenic plants the rate dropped from 22.5 μmol CO ₂ / m ² s to 15 μmol CO ₂ / ms. If the areas under both curves are integrated, the photosynthesis rate of the DARA12 plants is doubled compared to the non-transformed control plants.

The increased photosynthetic performance is correlated with the following phenotypic changes that characterize the transgenic DARA plants.

• Anatomical changes

The anatomical changes in the leaves are summarized in Table 2. The leaves of the transgenic plants are smaller and about 15% thicker. The increased leaf thickness is due to an extension of the cells of the palisade parenchyma by 50%. Furthermore, they are characterized by a specific leaf weight (g / cm ² ) increased by 60%. With approximately the same total leaf area of all lines, the total leaf weight per plant is increased by 50%. In plants from which the leaves are harvested (alfalfa, lettuce, cabbage, tobacco), the overexpression of PhyB enables a yield increase of 50%.

Table 2

Anatomical characterization of the leaves of the transgenic potato lines DARA5 and DARA12 in comparison to the untransformed control line (wild type). The length of the palisade parenchyma cells and the total leaf thickness were determined with a light microscope using a scale eyepiece. The n samples for each parameter come from three different 6-12 week old greenhouse plants (April-July 1997) (with the exception of the last column: 2 different plants). kM = no measurements. TABLE 2

• Changes in the amount of pigment

Table 3 shows the absolute and relative amounts of the photosynthetic pigments in the leaves of the transgenic potato line DARA12 compared to the untransformed control line. All pigments measured are increased by the same amount (approx. 26%). It can be concluded that the photosynthetic centers of the transgenic lines do not differ qualitatively from those of the controls.

Table 3

Absolute and relative amounts of photosynthetic pigments in leaves of the transgenic potato line DARA12 compared to the untransformed control line (wild type). The data come from HPLC analyzes of leaf segments of 9 week old greenhouse plants. 6 segments of different plants were taken from each line and analyzed in 2 separations of 3 segments each. Carotenoids (*) are the sum of the amounts of neoxanthine, violaxanthin, antheraxanthin, lutein, zeaxanthin, α- and ß-carotene.

Chlorophyll Chlorophyll a / b Carotenoids ^* Car / Chl a + b a + b (nmol / cm ² ) (nmol / cm ² ) (mmol / mol)

DARA12 58.6 3.82 26.2 442.3 wild type 46.4 3.96 20.6 446.5 4. Delay in senescence

Senescence can be recognized phenotypically from the yellowing of the leaves. These symptoms occur in untransformed control plants at a time when the plants of the DARA12 lines are still dark green. Thus, the transgenic plants have the possibility to use the energy of the sunlight for assimilation of carbon dioxide over a longer period of time.

5. Enlargement of the root ball

An extensive underground shoot root system enables the plant to be well supplied with water and nutrients. Figure 4 shows the enlargement of the root ball of the transgenic DARA plants compared to untransformed control plants.

6. Increase in income

For potato plants, the tuber yield is crucial when assessing the performance of a line. Figure 4 and Table 4 show that DARA plants form up to three times more tubers. On average, the yield increase is between 10-40% if the plants are cultivated for at least 5 months. The starch content of the tubers is unchanged at 15% of the fresh weight.

Table 4

Bulb yield and number of individual bulbs per plant of the transgenic potato lines DARA5 and DARA12 compared to the untransformed control line (wild type). The plants were exposed to sterile culture and cultivated in a greenhouse for 6 months (November 1996 - April 1997). The tubers of 5 individual plants were evaluated per line. In the case of the number of tubers, only tubers over 1 g are taken into account. TABLE 4

Yield number of tubers (g / plant) (n / plant)

DARA5 196.7 ± 41, 5 21, 5 ± 6.6

DARA1 164.4 ± 18.7 28.3 ± 5.8 2

Wild type 138.2 ± 23.0 9.6 ± 1.9

7. Other properties of the DARA lines

The plants are also characterized by an up to 50% higher antocyanine content, which gives them better protection against UV radiation.

Description of the transgenic potato plants that express the type B phytochrome of Solanum tuberosum in increased amounts.

1. Production of transgenic plants

The coding region of phytochrome B from Solanum tuberosum (St phyB) was amplified by the polymerase chain reaction using a cDNA clone in IZAP2 (Heyer and Gatz, 1992b) as a template. The primers contained EcoRV interfaces. St phyB can thus be cloned as an EcoRV fragment into suitable expression vectors. Solanum tuberosum codes for two phyB alleles (phyB (c) and phyB (g)), which differ in 4 amino acids. The sequence information is stored in the database under the deposit number Y14572 S51538 (EMBL, Hinxton Hall, Hinxton, Cambridge, CD10 1SD, UK). In the example used here, the two phyB alleles were brought under the control of the CaMV 35S promoter (Benfey et al., 1990), which leads to the expression of the genes in all parts of the plant. The polyadenylation region of the octopine synthase gene from Agrobacterium tume faciens served to terminate the transcription. Genes in which the promoter, coding region and polyadenylly tion signal from different genes are referred to as chimeric genes. The constructs are therefore referred to below as chimeric St phyB genes. The techniques for producing such recombinant DNA molecules are described in Sambrook et al. (1989). The two chimeric St phyB gene was transformed via Agrobacterium tumefaciens-mediated gene transfer into potato plants (Rocha-Sosa et al., 1985). Transgenic plants that express increased amounts of phytochrome B from Solanum tuberosum can be identified phenotypically by the enlargement of the root ball and the increased number of tubers. (Figure 5, Table 5). Plants that overexpress the phyB (c) allele are referred to as DS plants, plants that overexpress the phyB (g) allele are referred to as DPOT plants. Two plants from each line were characterized more precisely (DS13 and DS23; DPOT1 and DPOT9).

2. Determination of the content of phytochrome B

In contrast to structural proteins, regulatory proteins are only expressed in small amounts in plants. The detection of phytochromes is often carried out immunologically using antibodies. Monoclonal antibodies against phytochrome B from A. thaliana are available (Somers et al., 1991). These antibodies also recognize the phytochrome B from S. tuberosum. A polyclonal serum is also available, which was obtained from S. tuberosum by immunizing rabbits with PhyB. The exact implementation of this proof is described in the method section. Figure 6 shows the result of the Western blot analysis. The comparative quantification using dilution series showed that the lines DS15 show a 12-fold increase in the amount of phytochrome compared to the untransformed control (Wt). The three other lines were increased by a factor of 5 in their phytochrome B content compared to the untransformed control plant. 3. Phenotypic characterization

The above-ground organs (stem, leaf) of the plants of the DS and the DPOT lines show no changes to untransformed control plants. Leaf anatomy, photosynthesis rate and senescence properties are unchanged.

However, the underground organs (shoot, root, tubers) are more developed (Figure 5). Table 5 documents that the number of tubers is increased by a factor of 2 to 3, the yield is increased by about 10-40% if the plants are cultivated for at least 5 months.

Table 5

Bulb yield and number of individual bulbs per plant of the transgenic potato lines DPOT1, DPOT9, DS13 and DS23 compared to the untransformed control line (wild type). The plants were exposed to sterile culture and cultivated in a greenhouse for 6 months (November 1996 - April 1997). The tubers of 5 individual plants (only DPOT9: 4 individual plants) were evaluated per line. In the case of the number of tubers, only tubers over 1 g are taken into account.

TABLE 5

Yield number of tubers (g / plant) (n / plant)

DPOT1 176.7 ± 47.3 19.0 ± 7.0

DPOT9 147.2 ± 75.9 17.2 ± 9.1

DS13 186.5 ± 32.0 27.0 ± 12.0

DS23 196.5 ± 46.6 20.3 ± 9.1

Wild type 138.2 ± 23.0 9.6 ± 1.9

These experiments show that the gene from Solanum tuberosum can also improve their properties after introduction into potatoes, although it improves a smaller number of traits than the gene from Arabidopsis thaliana. BIBLIOGRAPHY

Benfey PN, Ren L, Chua NH (1990) Combinatorial and synergistic properties of CaMV 35S enhancer subdomains. EMBO J 9, 1685-1696

Clack T, Mathews S, Sharrock RA (1993) The phytochrome apoprotein family in Arabidopsis is encoded by five genes: the sequences and expression of PHYD and PHYE. Plant Mol Biol 25, 413-427

Färber A, Young AJ, Ruban AV, Horton P, Jahns P (1997) Dynamics of xanthophyll-cycle activity in different antennae subcomplexes in photosynthetic membranes of higher plants. The relationship between zeaxanthin conversion and nonphotochemical fluorescence quencning. Plant Physiol (in press)

Halliday K.J., Thomas B., Whitelam G.C. (19 ..), Expression of heterologous phytochromes A, B or C in transgenic tobacco plants alters vegetative development and flowering time. Plant J. 12, 1089-1090.

Hershey HP et al., 1985

Analysis of cloned cDNA and genomic sequences for phytochrome: complete amino acid sequences for two gene products expressed in etiolated Avena. Nucleic Acids Res 13 (23), 8543-8559 (1985)

Heyer AG, Gatz C (1992) Isolation and characterization of a cDNA clone coding for potato type B phytochrome. Plant Mol Biol 20, 589-600

Kendrick, R.E., G.H.M. Kronenberg, ed., 1986, Photomorphogenesis in Plants. The Netherlands: Martinus Nijhoff, Dordrecht.

Laemmli UK (1970). Cleavage of structural proteins during the asssembly of the head of bacteriophage T4. Nature 227, 680-685

Murashige T, Skoog F. (1962) A revised for rapid growth and bio assays with tobacco tissue culture. Physiol Plant 15, 473-497

Pratt LH, Cordonnier-Pratt M-M, Kelmenson PM, Lazarova Gl, Kubota T, Alba (1997) The phytochrome family in tomato (Solanum lycopersicum L.) Plant Cell Environ 20: 6, 672-677

Rocha-Sosa M, Sonnewald U, Frommer W, Stratmann M, Schell J, Willmitzer L (1989) Both developmental and metabolic Signals activate promoter of the that I patatin gene. EMBO J 8, 23-29

Sambrook J, Fritsch EF, Maniatis T (1989) In: Molecular cloning: a laboratory manual. Cold Spring Harbor Laboratory, 2nd ed.Cold Spring Harbor, New York Sharrock RA, Quail PQ (1989) Novel phytochrome sequences in Arabidopsis thaliana: structure, evolution, and differential expression of a plant regulatory photoreceptor family. Genes Devel 3, 1745-1757

Somers DE, Quail PH (1991) Phytochrome-mediated light regulation of PHYA and PHYB-GUS transgenes in Arabidopsis thaliana. Plant Physiol 107, 523-534

Thiele A, Schirwitz K, Winter K, Krause GH (1996) Increased xanthophyll cycle activity and reduced D1 protein inactviation related to photoinhibifion in two plant Systems acclimated to excess light. Plant Science 115, 237-250

Van Tuinen A, Kerckhoffs LHJ, Nagatani A, Kendrick RE, Koorneef M (1995b) A temporarily red light-sensitive mutant of tomato lacks a light-stable, B-like phytochrome. Plant Physiol 108, 939-947

Von Scheele C, Swensson G, Rassmusson J (1937) Determining the starch content and dry matter of the potato using the specific weight. Agriculture! Verse station 127, 67-96

Wagner D, Tepperman JM, Quail PH (1991) Overexpression of phytochrome B induces a Short hypocotyl phenotype in transgenic Arabidopsis. Plant Cell 3, 1275-1288

SEQUENCE LOG

(1. GENERAL INFORMATION:

(i) APPLICANT:

(A) NAME: KWS Kleinwanzlebener Saatzucht AG

(B) ROAD: PO Box 1463

(C) LOCATION: Einbeck

(E) COUNTRY: Germany

(F) POSTAL NUMBER: 37555

(ii) DESCRIPTION OF THE INVENTION: Plants with increased phytochrome expression (iii) NUMBER OF SEQUENCES: 2

(iv) COMPUTER READABLE VERSION:

(A) DISK: Floppy disk

(B) COMPUTER: IBM PC compatible

(C) OPERATING SYSTEM: PC-DOS / MS-DOS

(D) SOFTWARE: Pateπtln Release # 1.0, Version # 1.30 (EPA)

(2) INFORMATION ON SEQ ID NO: 1:

(i) SEQUENCE LABEL:

(A) LENGTH: 40 amino acids

(B) TYPE: amino acid

(C) STRAND FORM:

(D) TOPOLOGY: linear

(ii) MOLECULE TYPE: Peptide

(xi) SEQUENCE DESCRIPTION: SEQ ID NO: 1:

Met Val Ser Gly Val Gly Gly Ser Gly Gly Gly Arg Gly Gly Gly Arg 1 5 10 15

Gly Gly Glu Glu Glu Pro Ser Ser Ser His Thr Pro Asn Asn Arg Arg 20 25 30

Gly Gly Glu Gin Ala Gin Ser Ser 35 40

(2) INFORMATION ON SEQ ID NO: 2:

(i) SEQUENCE LABEL:

(A) LENGTH: 19 amino acids

(B) TYPE: amino acid

(C) STRAND FORM:

(D) TOPOLOGY: linear

(ii) MOLECULE TYPE: Peptide

(xi) SEQUENCE DESCRIPTION: SEQ ID NO: 2: Met Ala Ser Gly Ser Arg Thr Lys His Ser His His Ser Ser Gin Ala 1 5 10 15

Gin Ser Ser

Claims

claims

1. Plant, characterized in that it overexpresses phytochrome.

2. Plant according to claim 1, characterized in that it overexpresses phytochrome-B.

3. Plant according to claim 1 or 2, characterized in that the overexpression is triggered by the introduction of a phytochrome gene into the plant and / or phytochromic gene activation.

4. Plant according to one of claims 1 to 3, characterized in that the overexpression is triggered by the introduction of the phytochrome B gene into the plant and / or its activation.

5. Plant according to claim 4, characterized in that the phytochrome B gene comes from the family of the Solanaceae or Cruciferae.

6. Plant according to claim 5, characterized in that the phytochrome B gene comes from Arabidopsis thaliana.

7. Plant according to one or more of the preceding claims, characterized in that it is a monocot plant.

8. Plant according to claim 7, characterized in that it is selected from the group consisting of corn, wheat, millet, oats, rice, barley, sorghum, amaranth, onion, asparagus and sugar cane.

9. Plant according to one or more of the preceding claims 1-5, characterized in that it is a dicot plant.

10. Plant according to claim 9, characterized in that it is selected from the group consisting of alfalfa, soybean, petunias, cotton, sugar beet, sunflower, carrot, celery, cabbage, cucumber, pepper, canola, tomato, potato, Lentil, flax, broccoli, tobacco, bean, lettuce, rapeseed, cauliflower, spinach, Brussels sprouts, artichokes, peas, okra, pumpkin, kale, collard green, tea and coffee.

11. Plant according to one or more of claims 1 to 5, characterized in that it is an ornamental plant selected from the group consisting of geraniums, carnations, orchids, roses, impatiens, petunias, begonias, fuchsias, yolk flowers, chrysanthemums, gladioli , Astromeria, sage, veronica, daisies and iris.

12. Plant according to claim 10, characterized in that the plant is the potato, a sugar beet or a corn plant.

13. Plant according to one or more of the preceding claims, characterized in that the N-terminus of the amino acid sequence encoded by the phytochrome gene is hydrophobic.

14. Plant according to claim 13, characterized in that the N-terminus of the amino acid sequence encoded by the phytochrome gene is rich in glycine and / or alanine.

15. Plant according to claim 13, characterized in that the N-terminus of the amino acid sequence encoded by the phytochrome gene comprises the following sequence:

MVSGVGGSGGGRGGGRGGEEEPSSSHTPNNRRGGEQAQSS (Seq.lD Nr.1)

16. Plant parts, seeds and / or cells of a plant according to one of claims 1 to 15.

17. Use of one or more phytochrome genes for the production of the plants according to one or more of claims 1 to 15 and / or the plant parts, seeds and / or cells according to claim 16.

18. Use of a plant according to one or more of claims 1 to 15 and / or the plant parts, seeds and / or cells according to claim 16 for the production of plants with an increased rate of synthesis.

19. Use of a plant according to one or more of claims 1 to 15 and / or the plant parts, seeds and / or cells according to claim 16 for the production of plants with increased resistance to solar radiation.

20. Use of a plant according to one or more of claims 1 to 15 and / or the plant parts, seeds and / or cells according to claim 16 for the production of plants with delayed senescence.

21. Use of a plant according to one or more of claims 1 to 15 and / or the plant parts, seeds and / or cells according to claim 16 for the production of plants with a more pronounced root ball.

22. Use of a plant according to one or more of claims 1 to 15 and / or the plant parts, seeds and / or cells according to claim 16 for the production of plants with increased tuber yield.

23. Use of a plant according to one or more of claims 1 to 15 and / or the plant parts, seeds and / or cells according to claim 16 for the production of plants with an improved harvest index.

24. Use of a plant according to one or more of the preceding claims 1 to 15 or a plant part, seed and / or a cell according to claim 16 for controlling the whitefly.

25. Use of a plant according to one or more of claims 1 to 15 or of a plant part, seed and / or a line according to claim 16 for the production of plants resistant to solar radiation.

26. A method for producing a plant according to one or more of claims 1 to 15 or of plant parts, seeds and / or cells according to claim 16, comprising the step:

- Introducing a phytochrome gene as specified in one of claims 1 to 15 into the plant and / or activating a phytochrome gene in the plant.

27. Construct containing a phytochrome gene which codes for an amino acid sequence whose N-terminus is hydrophobic.

28. Construct containing a phytochrome gene which codes for an amino acid sequence whose N-terminus is rich in glycine.

29. Construct according to claim 28, containing a phytochrome gene which codes for an amino acid sequence whose N-terminus comprises the following sequence:

MVSGVGGSGGGRGGGRGGEEEPSSSHTPNNRRGGEQAQSS (Seq.lD Nr.1)

30. Plant containing a construct according to one of claims 27 to 29.