An active proton pump of intact vacuoles isolated from Tulipa petals

Anthocyanin pigments within Tulipa petal vacuoles provide the means for real-time spectrophotometric monitoring of vacuolar sap pH and for studying ATP-dependent proton transport in isolated, intact vacuoles. Spectra of petal extracts were used to select empirically those wavelengths giving an approximately linear variation in anthocyanin absorbance with pH over a pH range of interest. A sensitive single-beam spectrophotometer with vertical optics was used to minitor absorbance changes of intact, settled vacuoles. Substrates and inhibitors of vacuolar ATPase (Lin, W., Wagner, G.J., Siegelman, H.W. and Hind, Q. (1977) Biochim. Biophys. Acta 465, 110–117) were added to probe proton transport. Acidification of the vacuole sap occurred following addition of MgATP, but not CaATP. Proton accumulation was inhibited by 10 μM Dio 9, an inhibitor of tonoplast ATPase in vitro, and the proton gradient established by addition of MgATP was dissipated after addition of 10 μM CCCP. No pumping response was observed with intact protoplasts. Potential differences across the tonoplast were directly measured by impaling vacuoles with glass microelectrodes. Potential differences of 10–20 mV (inside positive) were recorded when vacuoles were suspended in 0.7 M mannitol/10 mM Hepes buffer (adjusted to pH 8.0 with KOH), and 0.5 mM dithiothreitol. Addition of MgATP increased the potential difference by 2–5 mV.     

Rapid Purification and Crystallization of Thermostable Malate Dehydrogenase Isolated from a Thermophilic Bacterium,Thermus flavus AT 62

A structural study of the water-soluble dextran made by Leuconostoc mesenteroides strain C (NRRL B-1298) was conducted by enzymic degradation and subsequent ¹³C-NMR analysis of the native dextran and its limit dextrins. The α-l,2-debranching enzyme removed almost all of the branched D-glucose residues, and gave a limit dextrin having a much longer sequence of the internal chain length (degree of linearity: n = 24.5 compared with the value of n = 3.3 for the native dextran). The degree of hydrolysis with debranching enzyme corresponded to the content of α-1,2-linkages determined by chemical methods, which suggested that most of the α-l,2-linkages in the dextran B-1298 constituted branch points of a single D-glucose residue. A synergistic increase of susceptibility of the dextran B-1299 was observed by simultaneous use of debranching enzyme and endodex-tranase. ¹³C-NMR spectral analysis indicated the similarity of structure of dextran B-1298 to that of B-1396, rather than that of B-1299. Occurrence of α-l,3-linkages in the limit dextrin was supported by a newly visualized chemical shift at 83.7 ppm.     

Temporal rhythm of petal programmed cell death in Ipomoea purpurea

• Flowers are the main sexual reproductive organs in plants. The shapes, colours and scents of corolla of plant flowers are involved in attracting insect pollinators and increasing reproductive success. The process of corolla senescence was investigated in Ipomoea purpurea (Convolvulaceae) in this study.
• In the research methods of plant anatomy, cytology, cell chemistry and molecular biology were used.
• The results showed that at the flowering stage cells already began to show distortion, chromatin condensation, mitochondrial membrane degradation and tonoplast dissolution and rupture. At this stage genomic DNA underwent massive but gradual random degradation. However, judging from the shape and structure, aging characteristics did not appear until the early flower senescence stage. The senescence process was slow, and it was completed at the late stage of flower senescence with a withering corolla.
• We may safely arrive at the conclusion that corolla senescence of I. purpurea was mediated by programmed cell death (PCD) that occurred at the flowering stage. The corolla senescence exhibited an obvious temporal rhythm, which demonstrated a high degree of coordination with pollination and fertilization.

     

A petal‐specific InMYB1 promoter from Japanese morning glory: a useful tool for molecular breeding of floricultural crops

Production of novel transgenic floricultural crops with altered petal properties requires transgenes that confer a useful trait and petal‐specific promoters. Several promoters have been shown to control transgenes in petals. However, all suffer from inherent drawbacks such as low petal specificity and restricted activity during the flowering stage. In addition, the promoters were not examined for their ability to confer petal‐specific expression in a wide range of plant species. Here, we report the promoter of InMYB1 from Japanese morning glory as a novel petal‐specific promoter for molecular breeding of floricultural crops. First, we produced stable InMYB1_1kb::GUS transgenic Arabidopsis and Eustoma plants and characterized spatial and temporal expression patterns under the control of the InMYB1 promoter by histochemical β‐glucuronidase (GUS) staining. GUS staining patterns were observed only in petals. This result showed that the InMYB1 promoter functions as a petal‐specific promoter. Second, we transiently introduced the InMYB1_1 kb::GUS construct into Eustoma, chrysanthemum, carnation, Japanese gentian, stock, rose, dendrobium and lily petals by particle bombardment. GUS staining spots were observed in Eustoma, chrysanthemum, carnation, Japanese gentian and stock. These results showed that the InMYB1 promoter functions in most dicots. Third, to show the InMYB1 promoter utility in molecular breeding, a MIXTA‐like gene function was suppressed or enhanced under the control of InMYB1 promoter in Arabidopsis. The transgenic plant showed a conspicuous morphological change only in the form of wrinkled petals. Based on these results, the InMYB1 promoter can be used as a petal‐specific promoter in molecular breeding of floricultural crops.     

FRL1 is required for petal and sepal development in Arabidopsis

A novel flower mutant, frl1 (frill 1) was isolated in Arabidopsis thaliana. The frl1 mutant has serrated petals and sepals but the other floral and vegetative organs appear to be normal. To analyse the role of the FRL1 gene, morphological, cytological and double mutant analyses were carried out. The frl1 flower had broader petals and sepals as compared with the wild-type. The distal region of frl1 petals contained fewer epidermal cells but their size was variable and generally larger than that in the wild-type. However, no significant difference was found in the basal region. Observations of the early petal development revealed that the morphology of the developing frl1 petal was normal until the middle of stage 9, but the frl1 phenotype became apparent in stages later than 10. Furthermore, larger nuclei with varied sizes were observed in the distal region of frl1 petals, but not in this region in wild-type petals. This strongly suggests that abnormal endo-reduplication had occurred. These observations indicate that the frl1 mutation affects the number of cell divisions and the subsequent cell expansion during the late stage of petal lamina formation, and that FRL1 might be maintaining the mitotic state or suppressing the transition to the endo-reduplication cycle. Double mutants with the homeotic mutants apetala3-1 and agamous showed additive phenotypes. Ectopic petals in the third whorl of fr11 ag flowers were serrated, indicating that the FRL1 gene acts in petal and sepal development in an organ-specific manner.     

基于叶绿体全基因组的云南樱花品种‘红霞''的种系分析

云南樱花品种‘红霞’(Cerasus‘Hongxia’)广泛种植于滇中地区，3月开花，半重瓣至重瓣，粉红色，观赏价值较高。‘红霞’曾被认为是红花高盆樱(C.cerasoides var.rubea)的园艺品种，而其分子标记结果却聚在钟花樱(C.campanulata)种系，可见‘红霞’的种系归属存疑，对今后的育种及栽培极为不利。为探索‘红霞’的种系问题，该研究以‘红霞’的叶片DNA为材料，应用Illumina软件对叶绿体基因组特征进行测序，通过SPAdes 3.13.0、Gapcloser 1.12和CPGAVAS2 2.0软件进行组装、注释，之后使用MISA和Geneious 10.0软件对叶绿体基因组特征进行分析，并使用RAxML 8.0软件构建Cerasus分支系统发育树，分析‘红霞’在樱属植物的系统关系。结果表明：(1)‘红霞’的叶绿体全基因组长度为157 832 bp,鸟嘌呤(Guanine)、胞嘧啶(Cytosine)含量为36.73%,包含一对反向重复(inverted repeat sequence, IR, 26 381 bp)区，由小单拷贝(short single...