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ABSTRACT: DNA markers are frequently used to analyze crop varieties, with the coded marker data summarized in a computer-generated table. Such summary tables often provide extraneous data about individual crop genotypes, needlessly complicating and prolonging DNA-based differentiation between crop varieties. At present, it is difficult to identify minimal marker sets - the smallest sets that can distinguish between all crop varieties listed in a marker-summary table - due to the absence of algorithms capable of such characterization. Here, we describe the development of just such an algorithm and MinimalMarker, its accompanying Perl-based computer program. MinimalMarker has been validated in variety identification of fruit trees using published datasets and is available for use with both dominant and co-dominant markers, regardless of the number of alleles, including SSR markers with numeric notation. We expect that this program will prove useful not only to genomics researchers but also to government agencies that use DNA markers to support a variety of food-inspection and -labeling regulations.
Journal of Bioinformatics and Computational Biology 04/2013; 11(2):1250022.
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ABSTRACT: Polyembryony, in which multiple somatic nucellar cell-derived embryos develop in addition to the zygotic embryo in a seed, is common in the genus Citrus. Previous genetic studies indicated polyembryony is mainly determined by a single locus, but the underlying molecular mechanism is still unclear. As a step towards identification and characterization of the gene or genes responsible for nucellar embryogenesis in Citrus, haplotype-specific physical maps around the polyembryony locus were constructed. By sequencing three BAC clones aligned on the polyembryony haplotype, a single contiguous draft sequence consisting of 380 kb containing 70 predicted open reading frames (ORFs) was reconstructed. Single nucleotide polymorphism genotypes detected in the sequenced genomic region showed strong association with embryo type in Citrus, indicating a common polyembryony locus is shared among widely diverse Citrus cultivars and species. The arrangement of the predicted ORFs in the characterized genomic region showed high collinearity to the genomic sequence of chromosome 4 of Vitis vinifera and linkage group VI of Populus trichocarpa, suggesting that the syntenic relationship among these species is conserved even though V. vinifera and P. trichocarpa are non-apomictic species. This is the first study to characterize in detail the genomic structure of an apomixis locus determining adventitious embryony.
Plant Science 02/2012; 183:131-42. · 2.94 Impact Factor
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Scientia Horticulturae 01/2012; 145:102. · 1.53 Impact Factor
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ABSTRACT: Ectopic expression of a Flowering Locus T (CiFT) from Citrus confers an early flowering phenotype on trifoliate orange (Poncirus trifoliata L. Raf.). To profile transcriptional effects of CiFT, mRNA extracted from the shoots of the transgenic trifoliate orange was subjected to genetic analysis using a 22-K oligo DNA microarray. Microarray results and reverse transcription polymerase chain reaction analysis indicated that genes relating to transcription, cell wall modification and defense responses were expressed at higher levels in the transgenic shoots than in the control wild-type shoots. Of the genes showing ectopic mRNA accumulation, two SEPALLATA (SEP) and one FRUITFULL (FUL) homologues (CuSEP1, CuSEP3 or CuFUL) were introduced to Arabidopsis thaliana. Constitutive expression of each gene caused early flowering in transgenic Arabidopsis, suggesting that these genes could function as regulators of flowering time.
Tree Physiology 03/2010; 30(3):431-9. · 2.88 Impact Factor
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Tomoko Endo,
Kuniyuki Kano,
Rie Motoki,
Kotaro Hama,
Shinichi Okudaira,
Mayuko Ishida,
Hideo Ogiso,
Masayuki Tanaka,
Norio Matsuki,
Ryo Taguchi,
Motomu Kanai,
Masakatsu Shibasaki,
Hiroyuki Arai,
Junken Aoki
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ABSTRACT: Lysophosphatidic acid (LPA) is a simple phospholipid but has numerous biological effects through a series of G-protein-coupled receptors specific to LPA. In general, LPA is short-lived when applied in vivo, which hinders most pharmacological experiments. In our continuing study to identify stable LPA analogues capable of in vivo applications, we identified here lysophosphatidylmethanol (LPM) as a stable and pan-LPA receptor agonist. A synthetic LPM activated all five LPA receptors (LPA(1-5)), and stimulates both cell proliferation and LPA-receptor-dependent cell motility. In addition, LPM showed a hypertensive effect in rodent when applied in vivo. We found that, when fetal calf serum was incubated in the presence of methanol, formation of LPM occurred rapidly, whereas it was completely blocked by depletion of autotaxin (ATX), a plasma enzyme that converts lysophosphatidylcholine (LPC) to LPA. When recombinant ATX was incubated with LPC in the presence of methanol, both LPM and LPA were produced with a ratio of 1:10, showing that ATX has transphosphatidylation activity in addition to its lysophospholipase D activity. Administration of methanol in mice resulted in the formation of several micromoles of LPM in plasma, which is much higher than that of LPA. The present study identified LPM as a novel and stable lysophospholipid mediator with LPA-like activities and ATX as a potential synthetic enzyme for LPM.
Journal of biochemistry 06/2009; 146(2):283-93. · 1.95 Impact Factor
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ABSTRACT: To determine differences in seasonal flowering between evergreen and deciduous woody perennials, endogenous expression of flowering-related genes was investigated in Satsuma mandarin (Citrus unshiu Marc.) and its close relative, trifoliate orange (Poncirus trifoliata (L.) Raf.), which are evergreen and deciduous, respectively, and show different seasonal flowering characteristics. In Satsuma mandarin, in which floral induction is triggered by low temperatures during fall and winter, mRNA levels of the citrus FLOWERING LOCUS T homologue CiFT increased during fall and winter, corresponding to the floral induction period, and mRNA levels of citrus LEAFY and SEPALLATA homologues (CsLFY and CuSEP) increased during early spring just before blooming. Citrus APETALA1 and FRUITFULL homologues (CsAP1 and CuFUL) did not show a significant association with seasonal flowering. In trifoliate orange, in which floral induction and flower bud development occur during early summer as in many deciduous trees, expression of CiFT, CsLFY, CsAP1, CuSEPs and CuFUL increased during early summer, corresponding to the period of floral induction and flower bud development. The CuSEPs expression peaked again during early spring just before blooming. In both species, the citrus TERMINAL FLOWER1 homologue (CsTFL), which acts as a floral repressor, showed low transcript levels during the period of floral induction and flower bud development. Thus, despite the difference in flowering season, in both species transcriptional changes in CiFT, CsLFY, CsTFL and CuSEPs were correlated with seasonal flowering. In contrast, the correspondence between CsAP1 and CuFUL expression and seasonal flowering differed between the species.
Tree Physiology 05/2009; 29(7):921-6. · 2.88 Impact Factor
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Budrul Ahsan,
Daisuke Kobayashi,
Tomoyuki Yamada,
Masahiro Kasahara,
Shin Sasaki,
Taro L Saito,
Yukinobu Nagayasu,
Koichiro Doi,
Yoichiro Nakatani,
Wei Qu, [......],
Shinobu Haga,
Fumiko Ohta,
Hisayo Nomoto,
Keiko Nogata,
Tomomi Morishita, Tomoko Endo,
Tadasu Shin-I,
Hiroyuki Takeda,
Yuji Kohara,
Shinichi Morishita
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ABSTRACT: Medaka (Oryzias latipes) is a small egg-laying freshwater teleost native to East Asia that has become an excellent model system for developmental genetics and evolutionary biology. The draft medaka genome sequence (700 Mb) was reported in June 2007, and its substantial genomic resources have been opened to the public through the University of Tokyo Genome Browser Medaka (UTGB/medaka) database. This database provides basic genomic information, such as predicted genes, expressed sequence tags (ESTs), guanine/cytosine (GC) content, repeats and comparative genomics, as well as unique data resources including (i) 2473 genetic markers and experimentally confirmed PCR primers that amplify these markers, (ii) 142,414 bacterial artificial chromosome (BAC) and 217,344 fosmid end sequences that amount to 15.0- and 11.1-fold clone coverage of the entire genome, respectively, and were used for draft genome assembly, (iii) 16,519,460 single nucleotide polymorphisms (SNPs), and 2 859 905 insertions/deletions detected between two medaka inbred strain genomes and (iv) 841 235 5'-end serial analyses of gene-expression (SAGE) tags that identified 344 266 transcription start sites on the genome. UTGB/medaka is available at: http://medaka.utgenome.org/.
Nucleic Acids Research 02/2008; 36(Database issue):D747-52. · 8.03 Impact Factor
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Kotaro Hama,
Junken Aoki,
Asuka Inoue, Tomoko Endo,
Tomokazu Amano,
Rie Motoki,
Motomu Kanai,
Xiaoqin Ye,
Jerold Chun,
Norio Matsuki,
Hiroshi Suzuki,
Masakatsu Shibasaki,
Hiroyuki Arai
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ABSTRACT: In polytocous animals, blastocysts are evenly distributed along each uterine horn and implant. The molecular mechanisms underlying these precise events remain elusive. We recently showed that lysophosphatidic acid (LPA) has critical roles in the establishment of early pregnancy by affecting embryo spacing and subsequent implantation through its receptor, LPA3. Targeted deletion of Lpa3 in mice resulted in delayed implantation and embryo crowding, which is associated with a dramatic decrease in the prostaglandins and prostaglandin-endoperoxide synthase 2 expression levels. Exogenous administration of prostaglandins rescued the delayed implantation but did not rescue the defects in embryo spacing, suggesting the role of prostaglandins in implantation downstream of LPA3 signaling. In the present study, to know how LPA3 signaling regulates the embryo spacing, we determined the time course distribution of blastocysts during the preimplantation period. In wild-type (WT) uteri, blastocysts were distributed evenly along the uterine horns at Embryonic Day 3.8 (E3.8), whereas in the Lpa3-deficient uteri, they were clustered in the vicinity of the cervix, suggesting that the mislocalization and resulting crowding of the embryos are the cause of the delayed implantation. However, embryos transferred singly into E2.5 pseudopregnant Lpa3-deficient uterine horns still showed delayed implantation but on-time implantation in WT uteri, indicating that embryo spacing and implantation timing are two segregated events. We also found that an LPA3-specific agonist induced rapid uterine contraction in WT mice but not in Lpa3-deficient mice. Because the uterine contraction is critical for embryo spacing, our results suggest that LPA3 signaling controls embryo spacing via uterine contraction around E3.5.
Biology of Reproduction 01/2008; 77(6):954-9. · 4.01 Impact Factor
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Masahiro Kasahara,
Kiyoshi Naruse,
Shin Sasaki,
Yoichiro Nakatani,
Wei Qu,
Budrul Ahsan,
Tomoyuki Yamada,
Yukinobu Nagayasu,
Koichiro Doi,
Yasuhiro Kasai, [......],
Shinobu Haga,
Fumiko Ohta,
Hisayo Nomoto,
Keiko Nogata,
Tomomi Morishita, Tomoko Endo,
Tadasu Shin-I,
Hiroyuki Takeda,
Shinichi Morishita,
Yuji Kohara
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ABSTRACT: Teleosts comprise more than half of all vertebrate species and have adapted to a variety of marine and freshwater habitats. Their genome evolution and diversification are important subjects for the understanding of vertebrate evolution. Although draft genome sequences of two pufferfishes have been published, analysis of more fish genomes is desirable. Here we report a high-quality draft genome sequence of a small egg-laying freshwater teleost, medaka (Oryzias latipes). Medaka is native to East Asia and an excellent model system for a wide range of biology, including ecotoxicology, carcinogenesis, sex determination and developmental genetics. In the assembled medaka genome (700 megabases), which is less than half of the zebrafish genome, we predicted 20,141 genes, including approximately 2,900 new genes, using 5'-end serial analysis of gene expression tag information. We found single nucleotide polymorphisms (SNPs) at an average rate of 3.42% between the two inbred strains derived from two regional populations; this is the highest SNP rate seen in any vertebrate species. Analyses based on the dense SNP information show a strict genetic separation of 4 million years (Myr) between the two populations, and suggest that differential selective pressures acted on specific gene categories. Four-way comparisons with the human, pufferfish (Tetraodon), zebrafish and medaka genomes revealed that eight major interchromosomal rearrangements took place in a remarkably short period of approximately 50 Myr after the whole-genome duplication event in the teleost ancestor and afterwards, intriguingly, the medaka genome preserved its ancestral karyotype for more than 300 Myr.
Nature 07/2007; 447(7145):714-9. · 36.28 Impact Factor
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Masahiro Kasahara,
Kiyoshi Naruse,
Shin Sasaki,
Yoichiro Nakatani,
Wei Qu,
Budrul Ahsan,
Tomoyuki Yamada,
Yukinobu Nagayasu,
Koichiro Doi,
Yasuhiro Kasai, [......],
Shinobu Haga,
Fumiko Ohta,
Hisayo Nomoto,
Keiko Nogata,
Tomomi Morishita, Tomoko Endo,
Tadasu Shin-I,
Hiroyuki Takeda,
Shinichi Morishita,
Yuji Kohara
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ABSTRACT: Teleosts comprise more than half of all vertebrate species and have adapted to a variety of marine and freshwater habitats
Nature 06/2007; 447(7145):714-719. · 36.28 Impact Factor
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ABSTRACT: After several years in the juvenile phase, adult citrus trees show seasonal periodicity of flowering. A prolonged exposure to low temperature is one of the most important environmental cues for floral induction in citrus. In the present study, the expression of flowering-related genes during the annual cycle of flowering and inductive low-temperature treatment in Satsuma mandarin (Citrus unshiu Marc.) trees is investigated. Simultaneously, floral induction, which occurs before the period of morphological flower development, was estimated as the number of flowers after the forcing of sprouting by defoliation at 25 degrees C. The expression of citrus FLOWERING LOCUS T homologues, CiFT, showed a seasonal increase during the floral induction period and was also induced by an artificial low-temperature treatment (15 degrees C) at which floral induction occurred. By contrast, the mRNA level of CiFT did not show any distinct changes following a warm-temperature treatment (25 degrees C) for 2.5 months, during which time floral induction was completely suppressed. Changes in the expression of the citrus homologues of TERMINAL FLOWER 1, LEAFY, and APETALA1 did not show any correlation with floral induction in the field or under artificial low-temperature conditions. In juvenile seedlings of Satsuma mandarin, which does not flower even under inductive low-temperature conditions, the mRNA levels of CiFT were not affected by the low-temperature treatment, unlike adult tissues. These results suggest that low temperature promotes floral induction via the activation of CiFT transcription in adult Satsuma mandarin trees and that, in the juvenile plant, CiFT transcription does not respond to low temperature.
Journal of Experimental Botany 02/2007; 58(14):3915-27. · 5.36 Impact Factor
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ABSTRACT: Reciprocal interactions between blastocysts and receptive uteri are essential for successful implantation. This process is regulated by the timely interplay of two ovarian hormones, progesterone and estrogen. However, the molecular targets of these hormones are largely unknown. We showed recently that a small bioactive lysophospholipid, lysophosphatidic acid, plays a pivotal role in the establishment of implantation via its cellular receptor, LPA(3). Here we demonstrate that LPA(3) expression is positively and negatively regulated by steroid hormones in mouse uteri. The LPA(3) mRNA level in the uteri increased during early pseudopregnancy, peaking around 3.5 days post coitus (3.5 d.p.c.), then, decreased to the basal level on 4.5 d.p.c. LPA(3) expression remained at a low level in ovariectomized mice, and administration of progesterone to ovariectomized mice up-regulated LPA(3) mRNA expression. In addition, simultaneous administration of estrogen counteracted the effect of progesterone. These results show that progesterone and estrogen cooperatively regulate LPA(3) expression, thereby contributing to the receptivity of uteri during early pregnancy.
Life Sciences 10/2006; 79(18):1736-40. · 2.53 Impact Factor
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ABSTRACT: Citrus FT (CiFT) cDNA, which promoted the transition from the vegetative to the reproductive phase in Arabidopsis thaliana, when constitutively expressed was introduced into trifoliate orange (Poncirus trifoliata L. Raf.). The transgenic plants in which CiFT was expressed constitutively showed early flowering, fruiting, and characteristic morphological changes. They started to flower as early as 12 weeks after transfer to a greenhouse, whereas wild-type plants usually have a long juvenile period of several years. Most of the transgenic flowers developed on leafy inflorescences, apparently in place of thorns; however, wild-type adult trifoliate orange usually develops solitary flowers in the axils of leaves. All of the transgenic lines accumulated CiFT mRNA in their shoots, but there were variations in the accumulation level. The transgenic lines showed variation in phenotypes, such as time to first flowering and tree shape. In F(1) progeny obtained by crossing 'Kiyomi' tangor (C. unshiu x sinensis) with the pollen of one transgenic line, extremely early flowering immediately after germination was observed. The transgene segregated in F(1) progeny in a Mendelian fashion, with complete co-segregation of the transgene and the early flowering phenotype. These results showed that constitutive expression of CiFT can reduce the generation time in trifoliate orange.
Transgenic Research 11/2005; 14(5):703-12. · 2.75 Impact Factor
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ABSTRACT: We isolated a cDNA clone (CitFLS) encoding flavonol synthase (FLS) from the satsuma mandarin (Citrus unshiu Marc.) fruit and investigated the steady state of CitFLS RNA expression during the fruit development. The CitFLS was 1274 bp long, encoded 335 amino acid residues, and belonged to a family of 2-oxoglutarate-dependent dioxygenases. The level of CitFLS transcript was higher in the young leaves than in the old leaves, and it was high at the early developmental stage and low at the mature stage in the juice sacs/segment epidermis (edible part). On the other hand, the CitFLS transcript increased in the peel during fruit maturation. These results indicated that the satsuma mandarin CitFLS was differentially regulated in the developmental stage and in a tissue-specific manner. Additionally, satsuma mandarin peel tissues produced rutin (a flavonol glycoside) from an exogenous dihydroquercetin (taxifolin), indicating the ability of these tissues to produce flavonols.
Physiologia Plantarum 03/2002; 114(2):251-258. · 3.11 Impact Factor
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ABSTRACT: The genomic region of a type 3 metallothionein (MT3)-like gene (CitMT45) with extensive expression in citrus fruit was isolated from Citrus unshiu Marc. and its promoter activity was analyzed using particle bombardment. A genomic library was screened with CitMT45 cDNA as a probe, and approximately 9.5 kb genomic fragments were obtained. The nucleotide sequence showed that the genomic CitMT45 consists of 3 exons and 2 introns, similar to other plant MT-like genes. Interestingly, the proximal 5′ flanking region had no canonical TATA-box or several cis-regulatory elements of G-box, MRE and ERE motifs common to plant MT-like genes. Lack of the MRE motif in the promoter region is in agreement with no response of CitMT45 to metal treatment, suggesting that CitMT45 might play other biological roles during fruit development rather than detoxification.Two promoters of proximal 190 bp region (PMT45C) and 4.2 kb full region (PMT45L) were fused to the uidA gene and directly incorporated into Citrus tissues using particle bombardment. The PMT45C promoter is sufficient to direct GUS expression in all tested organs and the upstream region from this proximal 190 bp sequence should have cis-elements that act as enhancers in citrus juice sacs. Moreover, a fluorometric GUS analysis of transgenic Arabidopsis indicated that the PMT45L promoter conferred quantitatively preferential expression in siliques. These results might indicate that the cis-elements required for preferential expression in citrus fruits are functionally conserved in heterologous Arabidopsis plants. Thus, it was demonstrated that the PMT45L promoter could be used for manipulation of fruit quality by genetic engineering in Citrus and Arabidopsis.
Scientia Horticulturae.
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ABSTRACT: We isolated and functionally characterized CitMTSL1 and CitMTSL4 cDNA clones from Satsuma mandarin (Citrus unshiu Marc.), which were identified as the 1,8 cineole and (E)-beta-ocimene synthase genes in Citrus species. Their predicted proteins possessed the general characteristics of plant monoterpene synthases, such as transit peptides in the N-terminus, RR sequences, and DDxxD motifs. Acyclic monoterpene synthase of CitMTSL4 had a unique structural feature that differentiated it from citrus cyclic monoterpene synthase genes. Phylogenetic tree analysis showed that the molecular evolutions of citrus monoterpene synthases might occur according to product specificity and enzymatic function before the divergence of Citrus species.In Satsuma mandarin, 1,8 cineole and (E)-beta-ocimene were comparatively abundant in flowers and then decreased toward fruit development. The transcript accumulation patterns of CitMTSL1 and CitMTSL4 were in good agreement with the biosynthesis of 1,8 cineole and (E)-beta-ocimene. We considered that the regulation of these gene expressions might be controlled according to some functional roles in plant development and that the different volatile profiles in tissues might depend on the gene expression.
Plant Science.
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ABSTRACT: We isolated four novel cDNA clones for monoterpene synthase genes, CitMTSE1, CitMTS3, CitMTS61, and CitMTS62, from Satsuma mandarin (Citrus unshiu Marc.). Functional tests of in vitro translation demonstrated that CitMTSE1, CitMTS3, CitMTS61, and CitMTS62 coded d-limonene synthase, gamma-terpinene synthase, gamma-terpinene synthase, and beta-pinene synthase, respectively. Their predicted proteins possessed typical characteristics for monoterpene synthases, such as transit peptides in the N-terminus, RR sequences, and DDXXD motifs. In citrus monoterpene synthase genes, the enzymatic product-specific interval length variations among conservative domains were conserved among interspecies, and they were conspicuous between limonene synthase genes and other cyclic monoterpene synthase genes, although the total amino acid length of each clone was similar. A phylogenetic tree indicated that differentiation in citrus monoterpene synthases might occur before the divergence of Citrus species. Besides, the mRNA expressions of most clones were active in the peel at an early stage of fruit development. CitMTSE1 and CitMTS3 were also powerfully expressed in the flower at anthesis, suggesting a different regulation mechanism.
Plant Science.
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ABSTRACT: To characterize the gene expression and regulation associated with anthesis stage of fruit development a total of 824 clones were randomly selected and sequenced from a cDNA library prepared from ovary tissue of Satsuma mandarin (Citrus unshiu Marc) at the anthesis stage. Clones derived from rDNA, mitochondrial and chloroplastic DNAs were removed and the remaining 544 cDNA clones were analyzed for homology with known sequences. From this analysis 455 clones were identified as homologues of a known gene or an expressed sequence tag (EST). We found 103 cDNA clones that were not detected in our previous analyses of Citrus ESTs including gene homologues related to transcription factors, molecular chaperones, and plant hormone biosynthesis/regulation. In addition, gene homologues related to amino acid biosynthesis, secondary metabolism and stress response were frequently detected, indicating that the ovary tissue may be metabolically active and have the intrinsic ability to react to biotic and abiotic stresses.
Plant Science. 165(1):167-168.
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ABSTRACT: We isolated a new d-limonene synthase gene (CitMTSE2) from Satsuma mandarin (Citrus unshiu Marc.) and compared its mRNA expression pattern with that of the previously isolated CitMTSE1. The predicted protein of CitMTSE2 showed 84.2% identity to that of CitMTSE1 at the amino acid level and possessed the typical structure and chemical features of general monoterpene synthases, such as transit peptides in the N-terminus, sequence motifs of RR and DDXXD, and cation-dependent reactions. A functional test demonstrated that CitMTSE2 is a d-limonene synthase gene.Under different expression regulations in response to the fruit developmental stage, the mRNA expression of CitMTSE2 indicated a specificity to fruit tissue and was different from that of CitMTSE1. Occurring mainly in peel at an early stage of fruit development, the mRNA expression profile of CitMTSE2 was similar to that of d-limonene biosynthesis in C. unshiu.
Scientia Horticulturae 105(4):507-512. · 1.53 Impact Factor
