Invited Review

Genetic engineering of chrysanthemum (Chrysanthemum morifolium): Current progress and perspectives

Harue Shinoyama, Ryutaro Aida, Hiroaki Ichikawa, Yukio Nomura, Atsushi Mochizuki. . . . . . . 323 [PDF]

Chrysanthemum is one of the most popular ornamental flowers. Recently agronomically useful traits have been introduced through genetic transformation. We give an overview of the advances in the genetic engineering of chrysanthemums, clarifying their problems and solutions. Then we describe our improved transformation system and example of the introduction into chrysanthemum of insect-resistance to reduce agrochemicals and sterility to maintain biological diversity.

Original Papers

Evaluation of selected transgenic papaya (Carica papaya L.) lines for inheritance of resistance to papaya ringspot virus and horticultural traits

Abigail May O. Retuta, Pablito M. Magdalita, Evalour T. Aspuria, Rene Rafael C. Espino. . . . . . . 339 [PDF]

Evaluation for resistance to Papaya Ringspot Virus (PRSV) and selected horticultural traits of transgenic papaya of selected lines across T1 and T2 generations were conducted under Biological Containment Level 2 (BL2) greenhouse conditions. The main objective of this study is to determine the partial stability of phenotypic expression of the viral gene (CP) from a Philippine isolate of PRSV in transgenic papaya cv. ‘Kapoho Solo’ and its resistance to PRSV across two generations. Reaction of the transgenic papaya seedlings to mechanical inoculation of PRSV was examined. Phenotypic characterization was done at the flowering stage while evaluation of fruit traits was done on ripe fruits. Results of the evaluation revealed that the T1 lines showed higher percentage of resistance to PRSV compared to their T2 progenies. The T1 and T2 plants and their fruits exhibited typical morphology as the ‘Kapoho Solo’ control plants.

Isolation of polyphenol oxidase genes from Portulaca oleracea and evaluation of their ability to metabolize endocrine-disrupting chemicals

Hirokazu Kaneda, Takeshi Matsui, Ryosuke Tomiyasu, Yukako Kuroda, Yuka Higashimoto, Tomoka Oda, Hitoshi Miyasaka, Hiroshi Okuhata, Satoshi Tanaka, Kazuo Harada, Hideyuki Matsuura, Hideki Nakayama, Ko Kato, Kazumasa Hirata. . . . . . . 351 [PDF]


Portulaca (Portulaca oleracea cv.), a garden plant, efficiently removes endocrine-disrupting chemicals (EDCs) including bisphenol A (BPA) from hydroponic solution. We isolated five polyphenol oxidase (PPO) genes from portulaca root. PoPPO2, PoPPO4, PoPPO5 were introduced into cultured tobacco cells. We found that crude extracts from the cells expressing PoPPOs could metabolize BPA and other EDCs.

Rapid analysis of transgenic rice straw using near-infrared spectroscopy

Takefumi Hattori, Shinya Murakami, Mai Mukai, Tatsuhiko Yamada, Hirohiko Hirochika, Masakazu Ike, Ken Tokuyasu, Shiro Suzuki, Masahiro Sakamoto, Toshiaki Umezawa. . . . . . . 359 [PDF]

A rapid near-infrared (NIR) spectroscopy method was established to predict the lignin and starch contents and enzymatic saccharification efficiency of transgenic rice (Oryza satia cv. Nipponbare) straw wherein expression of genes encoding lignin synthetic enzymes are regulated. Strong correlations were obtained between laboratory wet chemistry values and the NIR-predicted values. This method is useful to develop transgenic rice where cell wall formation is engineered.

Arabidopsis ENHANCER OF SHOOT REGENERATION 2 and PINOID are involved in in vitro shoot regeneration

Naoki Matsuo, Hiroharu Banno. . . . . . . 367 [PDF]

We investigated the genetic interaction between ESR2 and PINOID (PID) during shoot regeneration in this study. Both esr2 and pid mutations markedly decreased the efficiency of shoot regeneration from root segments at comparable levels, while esr2-2 pid double mutants dramatically decreased the efficiency of shoot regeneration. Our results demonstrated an additive or synergistic effect of these two mutations on shoot regeneration.

Ectopic expression of the leucine-rich repeat-domain of cell wall extensin gene NpLRX1 disturbs morphogenesis of leaf cells by its association with cell membranes in tobacco

Katsumi Yazawa, Hiroaki Iwai, Haruna Sugiyama, Shu Ito, Seiichiro Hasezawa, Shinobu Satoh. . . . . . . 373 [PDF]

NpLRX1 encodes a chimeric leucine-rich repeat (LRR) /extensin protein and is thought to regulate cell morphogenesis. Abnormal cell morphogenesis was observed in the transgenic adventitious buds and BY-2 cultured cells, when truncated NpLRX1 (N/LRR) was expressed. NpLRX1-GFP fusion proteins were detected on the cell membrane. These results suggest that ectopic expression of N/LRR causes defects in cell morphogenesis of tobacco tissues by the association of the LRR-domain with cell membrane.

Transgenic tobacco plants expressing antimicrobial peptide bovine lactoferricin show enhanced resistance to phytopathogens

Shiro Fukuta, Kei-ichi Kawamoto, Yuko Mizukami, Yukie Yoshimura, Jun-ichi Ueda, Michio Kanbe. . . . . . . 383 [PDF]

Lactoferricin B is an antimicrobial peptide derived from bovine lactoferrin. We expressed the lactoferricin gene in tobacco plants to evaluate its antimicrobial activity. As a result, transgenic tobacco plants that overexpressed the lactoferricin gene, linked to the signal peptide of tobacco PR-1 protein under the control of a high expression constitutive promoter, showed enhanced resistance to bacterial (P. syringae pv. tabaci) and fungal diseases.

Short Communication

Silicon deficiency promotes lignin accumulation in rice

Shiro Suzuki, Jian Feng Ma, Naoki Yamamoto, Takefumi Hattori, Masahiro Sakamoto, Toshiaki Umezawa. . . . . . . 391 [PDF]

In this paper, we demonstrate a negative correlation between silicon accumulation and lignin deposition in rice aerial parts. Using rice mutants defecting silicon transporters and wild type cultivated on a low silicon medium, we demonstrate that large amount of lignin deposited in the aerial parts. Significant accumulation of guaiacyl lignin upon silicon deficiency was determined by nitrobenzene oxidation analysis.

Identification and properties of a small protein that interacts with a tobacco bZIP-type transcription factor TBZF

Seung Hwan Yang, Sang Hyon Kim, Thomas Berberich, Tomonobu Kusano. . . . . . . 395 [PDF]


A tobacco bZIP transcription factor, TBZF, functions in leaf senescence and floral development. Here we identify tobacco NtTIP1 as a TBZF-interaction partner using a yeast two-hybrid system. NtTIP1 is a nuclear protein belonging to the ASR/CI21 family. Both NtTIP1 and TBZF transcripts accumulated at high levels in flowers and senescing leaves, suggesting that NtTIP1 and TBZF form a complex in vivo, and they function in flower development and stress response.


Plant regeneration from embryogenic tissue of Pinus luchuensis Mayr, an endemic species in Ryukyu Island, Japan

Yoshihisa Hosoi, Tsuyoshi E. Maruyama. . . . . . . 401 [PDF]

Somatic embryogenesis in Ryukyumatsu (Pinus luchuensis Mayr.) was initiated from megagametophytes containing zygotic embryos. Embryogenic cultures were maintained and proliferated by subcultures at 2- to 3- week-intervals. The maturation of somatic embryos occurred on media containing maltose, activated charcoal, abscisic acid and polyethylene glycol. High frequencies of germination were obtained after the post-maturation treatment of somatic embryos, and around 90% of them were converted into plants.

The CAPRICE-LIKE MYB gene family cooperatively controls trichome branching and clustering in Arabidopsis

Rumi Tominaga-Wada, Yuka Nukumizu. . . . . . . 407 [PDF]

To understand the role of CPC-LIKE MYB genes in trichome development, we focused on the effect of these genes on trichome branching and clustering. The TRY and ETC1 genes cooperatively regulate trichome branch development. The ETC2 gene has little to no involvement in trichome branching. Double and triple mutant analyses revealed that the ETC1, ETC2 and CPL3 genes have some degree of functional redundancy with TRY in trichome cluster formation.

Characterization of cis-prenyltransferases from the rubber producing plant Hevea brasiliensis heterologously expressed in yeast and plant cells

Seiji Takahashi, Hye-Jin Lee, Satoshi Yamashita, Tanetoshi Koyama. . . . . . . 411 [PDF]

cis-Prenyltransferase (cPT) is thought to be a key enzyme in the natural rubber biosynthesis. The enzymatic characterization of HRT1 and HRT2, cPTs from latex of Hevea brasiliensis, was revealed that HRTs expressed in yeast and Arabidopsis T87 cultured cells showed distinct cPT activity, producing C80-100 polyisoprenoids, although failing to catalyze the formation of natural rubber. These results suggest that latex specific co-factor(s) may be required to enable HRT1/HRT2 rubber transferase activity.

Microscale thioacidolysis method for the rapid analysis of β-O-4 substructures in lignin

Masaomi Yamamura, Takefumi Hattori, Shiro Suzuki, Daisuke Shibata, Toshiaki Umezawa. . . . . . . 419 [PDF]

Thioacidolysis is a method to detect the β-O-4 substructures of lignin, and has been employed as a diagnostic test for presence of lignin. However, the conventional thioacidolysis protocol is low-throughput and is a bottleneck in the characterization of lignins in a large number of samples such as transgenic lines. Recently, rapid analysis protocol of thioacidolysis was reported (Robinson and Mansfield 2009). In this study, we modified mainly the work-up process. Our microscale protocol showed higher yields of thioacidolysis products than the conventional protocol.