Monoclonal Antibodies Specific for Non-Derivatized Gibberellins I. Preparation of Monoclonal Antibodies against GA4 and Their Use in Immunoaffinity Column Chromatography

Cell lines producing monoclonal antibodies specific for non-derivatizedgibberellins (GAs) were prepared from spleen cells of a mouseimmunized with an immunogen that carried 16-substituted GA₄as a hapten. The conditions for association and dissociationof antibody-hapten complexes were examined to determine themost effective procedure for immunoaffinity chromatography.The final procedure adopted was proven to be very effectiveby its application to the analysis of GAs in the anthers ofrice and immature seeds of Phaseolus vulgaris ⁴Present address: Department of Agricultural Chemistry, UtsunomiyaUniversity, Mine-machi 350, Utsunomiya, 321 Japan (Received August 8, 1990; Accepted March 7, 1991)     

Eu-Chlorella large subunit rDNA sequences and group I introns in ribosomal DNA of the paramecian symbiotic alga NC64A

Although the examination of large subunit ribosomal RNA genes (LSU rDNA) is advanced in phylogenetic studies, no corresponding sequence data from trebouxiophytes have been published, with the exception of ‘Chlorella’ellipsoidea Gerneck. We determined the LSU rDNA sequence of Chlorella vulgaris Beijerinck and of the symbiotic alga of green paramecium, Chlorella sp. NC64A. A total of 59 nucleotide substitutions were found in the LSU rDNA of the two species, which are disproportionately distributed. Primarily, 65% of the substitutions were encountered in the first 800 bp of the alignment. This segment apparently has evolved eight times faster than the complete SSU rDNA sequence, making it a good candidate for a phylogenetic marker and giving a resolution level intermediate between small subunit (SSU) rDNA and internal transcribed spacers. Green algae are known as a group I intron‐rich group along with rhodophytes and fungi. NC64A is particularly rich in the introns; five introns were newly identified from the LSU rDNA sequence, which we named Cnc.L200, Cnc.L1688, Cnc.L1926, Cnc.L2184 and Cnc.L2437, following the insertion positions. In the present study we analyzed these introns with three others (Cnc.S943, Cnc.S1367 and Cnc.S1512) that had already been found in NC64A SSU rDNA. Secondary structure modeling placed these introns in the group I intron family, with four introns belonging to subgroup C1 and the other four introns belonging to subgroup E. Five of the intron insertion positions are unique to the paramecian symbiont, which may indicate relatively recent events of intron infections that includes transpositions. Intron phylogeny showed unprecedented relationships; four Cnc. IC1 introns made a clade with some green algal introns with insertions at nine different positions, whereas four Cnc. IE introns made a clade with the S651 intron (Chlorella sp. AN 1–3), which lay as a sister to the S516 insertion position subfamily.     

Isolation of two new gibberellins from immature seeds of Canavalia

Summary Two new gibberellins, Canavalia gibberellin-I (CG-I) and Canavalia gebberellin-II (CG-II), were isolated from immature seeds of sword bean, Canavalia gladiata DC. The molecular formulas C₁₉H₂₂O₇ and C₁₉H₂₂O₆, were assigned to CG-I and CG-II, respectively, on the basis of high resolution mass, infrared and NMR spectra. The growth-promoting effects of CG-I and CG-II on dwarf maize mutants d ₁ and d ₅, rice seedlings and dwarf peas, Progress No. 5, are reported.     

Changes in Endogenous Ribosyl-trans-zeatin and IAA Levels in Relation to the Proliferation of Tobacco Crown Gall Cells

Changes in the contents of major endogenous plant hormones intobacco crown gall cells, namely IAA and ribosyl-trans-zeatin,during cell growth were examined using HPLC and ¹⁴C-labeledplant hormones. The content of IAA was high at the early logarithmicstage, while that of ribosyl-trans-zeatin was high at the middlelogarithmic stage. This suggests that cell growth is affectedfirst by IAA, then by ribosyl-trans-zeatin. ³ Present address: Department of Agricultural Chemistry, TottoriUniversity, Koyama, Tottori 680, Japan (Received July 13, 1981; Accepted September 11, 1981)     

The MIXTA-like Transcription factor MYB16 is a major regulator of cuticle formation in vegetative organs

Cuticle secreted on the surface of the epidermis of aerial organs protects plants from the external environment. We recently found that Arabidopsis MIXTA-like R2R3-MYB family members MYB16 and MYB106 regulate cuticle formation in reproductive organs and trichomes. However, the artificial miRNA (amiRNA)-mediated knockdown plants showed no clear phenotypic abnormality in vegetative tissues. In this study, we used RNA interference (RNAi) targeting MYB16 to produce plants with reduced expression of both MYB16 and MYB106. The rosette leaves of RNAi plants showed more severe permeable cuticle phenotypes than the myb106 mutants expressing the MYB16 amiRNA in the previous study. The RNAi plants also showed reduced expression of cuticle biosynthesis genes LACERATA and ECERIFERUM1. By contrast, expression of a gain-of-function MYB16 construct induced over-accumulation of waxy substances on leaves. These results suggest that MYB16 functions as a major regulator of cuticle formation in vegetative organs, in addition to its effect in reproductive organs and trichomes.     

Dry matter production, population structure and environmental conditions of the spring ephemeralErythronium japonicum growing in various habitats differing in sunlight exposure in cool temperate Japan

Phenology, dry matter production, population structure and environmental conditions were examined inErythronium japonicum Decne plants growing on the floor of a deciduous broad-leavedQuercus mongolica forest (Q.m. stand), an evergreen coniferousCryptomeria japonica plantation (C. j. stand) and bare ground left for 3 years after the clearing of a forest composed of youngQ. mongolica andPinus densiflora trees (bare stand) in the cool temperate zone of Japan. The average population density of the plants growing at theQ.m. stand was much higher than that observed at the bare stand, whereas the average number of flowering plants at the former stand was less than half of that at the latter. The population density and number of flowering plants growing at theC. j. stand were both less than 30% of those at theQ. m. stand. The number of seedlings at theQ. m. stand was much more than that at the bare andC. j. stands. Their survivorship rate over 1 year at the former stand also seems to be significantly higher than those at the other stands. Their aboveground and belowground parts at the bare stand were exposed to more severe heat and water stress than those at the other two stands. The net production per leaf area of the plants growing at theQ. m. stand was two and six times larger than those at the bare andC. j. stands, respectively. The plants at the bare stand did not use the available solar radiation as efficiently for dry matter production through photosynthesis as those growing at theQ. m. stand, whereas those at theC. j. stand are strongly restricted in their photosynthetic process by the significantly limited light condition on the floor of the evergreen coniferous plantation. The differences in the number of plants reaching sexual maturity, the density and structure of the population and the net production between their three habitats are discussed here from the standpoint of differences in environmental conditions.     

Radioimmunoassay for brassinosteroids and its use for comparative analysis of brassinosteroids in stems and seeds ofPhaseolus vulgaris

Antiserum against the brassinosteroid (BR), castasterone, was produced by immunizing a rabbit with castasterone-carboxymethoxylamine oxime conjugated with bovine serum albumin (BSA). In a radioimmunoassay (RIA), the antiserum recognized a range of naturally occurring BRs with varying specificities. Detection limits of castasterone and brassinolide were approximately 0.3 pmol. This RIA system was successfully used for analyzing endogenous BRs in seeds and stems ofPhaseolus vulgaris L., and showed that stems are quite different from seeds in terms of the species and quantity of the endogenous BRs.