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First in the world! Challenge for "blue roses"

Stories of development
Secret of the birth
suntory blue rose applause

Stories of development>

"blue roses"

The term "blue rose" in English signified "the impossible (a non-existent object)" because nobody could produce blue roses by hybridization breeding no matter how much they were desired. However, "the impossible" has become possible due to cutting-edge biotechnologies and the continuous efforts of scientists dedicated to producing blue roses. Senior General Manager, Yoshikazu Tanaka, Ph.D., who has participated in the blue rose project since the initiation of the project, Principal Researcher Yukihisa Katsumoto, Ph.D., who still engages in the study as the project leader, and Researcher Noriko Nakamura, Ph.D. talked about their passion and aspirations that led to the success of the development of blue roses.


1990 The Blue Rose Project started
Project members then: Australia (1990)

Producing blue roses---The challenge to the impossible started in 1990. Suntory decided to tackle this dreamful project in collaboration with an Australian venture company Florigene Ltd. (named Calgene Pacific Pty Ltd. then; hereinafter referred to as Florigene). Thanks to the rapid advancement of plant biotechnologies in the 1980s, it was expected that blue roses could be developed with these technologies. Thus, there were many research teams engaged in similar projects to produce blue roses in the world, and the competition had already started under cover.

Researchers at Suntory: Japan (back in those days)

There were two technical barriers that had to be solved to produce blue roses. One was to "isolate genes (blue genes) necessary to synthesize a blue pigment (delphinidin) from among tens of thousands of genes contained in blue flowers." The other was to "develop the methods to introduce these genes to cells of roses and produce genetically modified roses from these cells." It was especially necessary to solve the first issue of isolation of blue genes earlier than the rivals and apply for the patent of the gene because such genes were patentable.

The challenge to produce "blue roses," which signify the impossible, started. −-Senior General Manager Yoshikazu Tanaka Ph.D.

Researcher's aspiration

Blue genes were isolated from petunias, and the patent of the gene was filed

There are many plants that produce blue flowers in nature, but our research team first chose a petunia exhibiting a dark violet color in order to isolate blue genes. This was because the petunia had already served as a model plant for the research of flower pigments (such as anthocyanin) and their biosynthesis and the following knowledge had already accumulated:

Petunia from which blue genes were isolated

- Blue genes are cytochrome P450 type hydroxylase (enzymes involved in detoxification in the liver) genes.
- These genes work in petals but not in leaves.
- Red petunias that do not produce blue pigments do not have blue genes.
- Blue genes work most actively when petals are opening.
- Gene loci on the chromosomes were known.

We selected about 300 kinds of candidate genes from about 30,000 kinds of genes in petunias in order to identify the two blue genes of petunias. Our first plan was to introduce the candidate genes into petunias and observe flower color changes to determine whether they were actually blue genes. However, this method required several months until flowers bloomed to see color change. Therefore, to reduce the time required to obtain results, we decided to introduce the candidate genes of blue genes to yeast instead of plants and test their enzymatic activities in yeast. Thanks to this method, we obtained results in one week and were able to test the activities of many genes.

Finally, we succeeded in isolating blue genes on June 13, 1991.

A page of a laboratory notebook filled on June 13, 1991

Suntory and Florigene immediately applied for the patent of the blue genes. Because we applied for the patent earlier than any other competitors and monopolized the intellectual property right, we were able to engage in this research eliminating any competitors. We were also lucky to be granted with broad claims because our applications were the first ones for patents of genes with this kind of activity. Actually, introduction of these genes into petunias and tobacco resulted in the increase of blue pigments "delphinidin." Our paper describing these results was published in "Nature," the most prestigious scientific journal in the world.

The time when we discovered blue genes was the happiest moment of my research life. −-Senior General Manager Yoshikazu Tanaka Ph.D.

Researcher's aspiration

1994 Roses into which petunia blue genes were introduced flowered, but they showed no hint of blue

There are several methods to introduce genes into plants. In the blue rose project, we adopted a method of introducing them with using a soil bacterium called "Agrobacterium." Because this bacterium has the ability to carry its own genes into plant cells, it is used in the gene transfer of many plants. To select only cells that have received the genes after transfer and regenerate them into the plant body of roses, it is necessary to optimize plant hormones as well as kinds and concentrations of nutrients. The work to perform this procedure in sterilized conditions, using a plant in a test tube, is called "tissue culture."

Repeated tissue culture, learning through trial and error (back in those days)

In the case of roses, it takes about one year from the introduction of genes to the blooming. How effectively the introduced genes function differs from one genetically modified rose to another, so we have to produce as many genetically modified roses as possible. Efficiency of gene introduction also differs substantially by rose variety. Therefore, we continued tissue culturing, learning through trial and error

Finally, we were able to introduce genes to red roses. For the first time in 1994, roses to which two kinds of blue genes of petunia opened. However, the color of the flowers was still red and no blue pigment was detected although the genes were introduced without a doubt.

Color changes or delphinidin were not observed even after many promoters (DNA sequence regulation gene expression) were used to drive petunia blue genes. Thus, we decided to introduce blue genes other than from petunias to roses. We isolated blue genes from various plants with blue flowers, such as gentian, butterfly pea, and torenia, and introduced them into roses. However, no matter how many times we conducted experiments, only "roses with blue genes but without blue pigments" bloomed.

We even suspected that roses might decompose blue pigments.... −-Senior General Manager Yoshikazu Tanaka Ph.D.

Researcher's aspiration

We had many failures, but each of them taught us a useful lesson for the future −−Principal Researcher Yukihisa Katsumoto Ph.D.

Researcher's aspiration

1995 Succeeded in producing blue carnation
Moonseries

We could not harvest the fruits of our efforts, and hard times continued for the members involved in this project. What encouraged the researchers was the success of the development of genetically modified blue carnations. Blue genes of petunia did not work well in roses, but they worked as expected in carnations. The blue pigments "delphinidin" accumulated, and the color of flowers changed to blue.

These flowers, named "Moonseries," were put on sale in Japan in 1997, and the number of varieties has increased since then. These elegant and beautiful flowers, which signify "eternal happiness" in the language of flowers, have gained popularity.

Carnation production site (Ecuador)

Moonseries are the first genetically modified flowers in the world that were commercialized. Currently, these blue carnations are produced in Columbia and Ecuador, and are sold primarily in the USA, but in Europe and some countries as well. In Japan, six varieties with different dark and light color combinations are sold. Presently, not only blue genes of petunias but also those of pansies are used for some varieties.

"Blue carnations" were born using an additional gene to blue genes −-Senior General Manager Yoshikazu Tanaka Ph.D.

Researcher's aspiration

1996 Blue pigments finally accumulated in roses, too; flower color changed

Around that time, in addition to carnations, blue pigments started to be produced at last in roses by introducing blue genes isolated from a pansy. Clear color changes of roses were also finally observed. However, because we introduced these genes only into a red rose variety, these roses were far from something that could be called "blue rose," and their color was darkish red. Nevertheless, the production of blue pigments showed us a path to the birth of blue roses.

Even after blue pigments are produced, how blue the flower become depends greatly on the original characteristics of the roses to which genes are introduced. For example, if the pH is low (acidic) in the cell vacuoles in which blue pigments are accumulated, the color becomes red, and if it is neutral, the color becomes blue. The color also depends largely on whether components that can or cannot enhance blue exist in the vacuoles. In other words, the existence of blue pigments does not necessarily turn the flower color into blue.

We therefore chose, from among several hundred varieties, about 40 rose varieties which would be likely to result in the accumulation of a high percentage of blue pigments and a more bluish color, including those that were not commercialized, and continued experiments to introduce the blue genes in order to produce roses that really would look blue. At the same time, we continued our research to improve methods of tissue culture so that genes could be introduced into various rose varieties.

Pathway to synthesize rose flower pigments

We were excited about the prospect of blue roses coming! −-Senior General Manager Yoshikazu Tanaka Ph.D.

Researcher's aspiration

Our research was worthwhile though we had to cope with pressure −−Principal Researcher Yukihisa Katsumoto Ph.D.

Researcher's aspiration

1998-2002 The percentage of blue pigments increased to 100%; blue roses were finally born

To introduce genes into rose cells, we first need to induce undifferentiated cells (called "callus") whose functions or morphology has not been determined yet. In other words, blue genes are introduced into calluses for which no determination has been made as to whether they may become a part of a leaf or a stalk. We regenerate these cells to produce flowers. It takes as long as one year to produce calluses, so this experiment takes time and requires extreme patience. The researchers patiently devoted themselves to continue the work of introducing the blue genes of pansies into calluses. This was a method originally developed by Suntory. This technique forms the basis of the development of blue roses because it enables the introduction of genes into many rose varieties.

Around 1998-1999, slightly bluish roses started to bloom. We further continued gene transfer, and due to that effort, roses which accumulated almost 100% of the total blue pigments bloomed. In 2002, we selected transgenic lines of pure blue from them. Finally, the first blue roses in the world were born. Further, we succeeded in propagating these roses by grafting, and confirmed that roses of the same color were produced stably and grown normally.

Pursued blueness by trying various varieties −-Senior General Manager Yoshikazu Tanaka Ph.D.

Researcher's aspiration

Developmental process of blue roses
2004 Finally the success of the development was announced

In June 2004, we publicly announced the first success of the development of blue roses in the world, and showed them publicly in the press conference room. We received huge responses. The success was reported on the front pages of various newspapers, and covered by mass media overseas. The general consumer was also greatly interested, and warm responses were received from many people, including elementary school students and the elderly. The stories of the development of blue roses are included in scientific textbooks, exhibited at the National Museum of Nature and Science, and used in many teaching materials. We published some papers on the scientific aspects of the development, and received the PCP Award of the Japanese Society of Plant Physiologists in 2009.

However, we had to overcome another and possibly the highest hurdle before delivering blue roses to consumers. Because the blue roses developed by Suntory are genetically modified organisms, it was necessary to obtain legal permits from the Ministry of Agriculture, Forestry and Fisheries and the Ministry of Environment based on Act on the Conservation and Sustainable Use of Biological Diversity through Regulations on the Use of Living Modified Organisms (so-called Cartagena Protocol) in order to grow them commercially and sell them in Japan.

blue roses

For that purpose we had to conduct various experiments to prove that the production and sales of the developed roses in Japan would not affect Japanese biological diversity. For example, we spent as long as four years to conduct hybridization experiments including cross-pollinating the blue roses with wild rose species in order to prove that there was no risk of dispersal of the introduced genes among wild roses. We obtained the permits on January 31, 2008.

The news on the birth of blue roses made headlines around the world −-Senior General Manager Yoshikazu Tanaka Ph.D.

Researcher's aspiration

We traveled to the wilderness in Hokkaido to obtain permits --Researcher Noriko Nakamura Ph.D.

Researcher's aspiration

2009 Blue roses were marketed, and the pursuit to improve the blue color continued
夢かなう 佐治信忠

The first blue roses in the world were named "SUNTORY blue rose APPLAUSE," and marketed. The applause means cheers, and the blue roses signified "dreams come true" in the language of flowers. The flowers were given this name with the hope to send cheers to many people who have made efforts to make the dreams come true.

Blue roses were commercialized, but the Suntory's challenge to the impossible is not over yet. In the natural world, there are many flowers that are pure blue. These flowers are able to not only synthesize blue pigments but also produce pure blue by combining blue pigments with metal ions, such as aluminum, and various co-existing compounds with the blue pigments, including flavones. It is also known that the higher the pH in the cellular vacuoles in which blue pigments accumulated, the more pure blue the color is.

If such mechanisms to produce blue flowers are reproduced in roses, roses in more of a pure blue than the current APPLAUSE should be developed. Suntory's challenge to produce "pure blue roses" continues today.

The "challenge to the impossible" is not over yet −-Senior General Manager Yoshikazu Tanaka Ph.D.

Researcher's aspiration

Warm responses from consumers encourage me most −−Principal Researcher Yukihisa Katsumoto Ph.D.

Researcher's aspiration

Hope to sell APPLAUSE to overseas consumers --Researcher Noriko Nakamura Ph.D.

Researcher's aspiration

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