A biosensor for the direct visualization of auxin

One of the most important regulatory small molecules in plants is indole-3-acetic acid, also known as auxin. Its dynamic redistribution has an essential role in almost every aspect of plant life, ranging from cell shape and division to organogenesis and responses to light and gravity1,2. So far, it has not been possible to directly determine the spatial and temporal distribution of auxin at a cellular resolution. Instead it is inferred from the visualization of irreversible processes that involve the endogenous auxin-response machinery3–7; however, such a system cannot detect transient changes. Here we report a genetically encoded biosensor for the quantitative in vivo visualization of auxin distribution. The sensor is based on the Escherichia coli tryptophan repressor8, the binding pocket of which is engineered to be specific to auxin. Coupling of the auxin-binding moiety with selected fluorescent proteins enables the use of a fluorescence resonance energy transfer signal as a readout. Unlike previous systems, this sensor enables direct monitoring of the rapid uptake and clearance of auxin by individual cells and within cell compartments in planta. By responding to the graded spatial distribution along the root axis and its perturbation by transport inhibitors—as well as the rapid and reversible redistribution of endogenous auxin in response to changes in gravity vectors—our sensor enables real-time monitoring of auxin concentrations at a (sub)cellular resolution and their spatial and temporal changes during the lifespan of a plant.

A Role for Auxin in Triggering Lamina Outgrowth of Unifacial Leaves

A common morphological feature of typical angiosperms is the patterning of lateral organs along primary axes of asymmetry—a proximodistal, a mediolateral, and an adaxial–abaxial axis. Angiosperm leaves usually have distinct adaxial–abaxial identity, which is required for the development of a flat shape. By contrast, many unifacial leaves, consisting of only the abaxial side, show a flattened morphology. This implicates a unique mechanism that allows leaf flattening independent of adaxial–abaxial identity. In this study, we report a role for auxin in outgrowth of unifacial leaves. In two closely related unifacial-leaved species of Juncaceae, Juncus prismatocarpus with flattened leaves, and J. wallichianus with transversally radialized leaves, the auxin-responsive gene GLYCOSIDE HYDROLASE3 (GH3) displayed spatially different expression patterns within leaf primordia. Treatment of J. prismatocarpus seedlings with exogenous auxin or auxin transport inhibitors, which disturb endogenous auxin distribution, eliminated leaf flatness, resulting in a transversally radialized morphology. These treatments did not affect the radialized morphology of leaves of J. wallichianus. Moreover, elimination of leaf flatness by these treatments accompanied dysregulated expression of genetic factors needed to specify the leaf central-marginal polarity in J. prismatocarpus. The findings imply that lamina outgrowth of unifacial leaves relies on proper placement of auxin, which might induce initial leaf flattening and subsequently act to specify leaf polarity, promoting further flattening growth of leaves.

Multiplikasi in vitro stroberi kultivar Tochiotome dengan penambahan jenis dan konsentrasi sitokinin untuk perbanyakan bibit

Sari Perbanyakan tanaman stroberi secara konvensional dilakukan dengan menggunakan stolon, tetapi kurang efektif serta kualitas bibit yang dihasilkan kurang baik akibat adanya akumulasi penyakit. Budidaya stroberi memerlukan adanya perbanyakan bibit secara massal, tetapi tidak mengubah kualitasnya. Multiplikasi in vitro menjadi solusi untuk penyediaan bibit berkualitas dalam jumlah besar. Upaya untuk mendapatkan tunas in vitro dalam jumlah banyak yakni perlu adanya penambahan zat pengatur tumbuh golongan sitokinin seperti Benzylaminopurine (BAP) atau Thidiazuron (TDZ). Tujuan penelitian ini adalah mengetahui dan menetapkan jenis serta konsentrasi sitokinin dengan hasil terbaik dalam multiplikasi stroberi kultivar Tochiotome. Percobaan dilaksanakan di Laboratorium Kultur Jaringan Tanaman, Teknologi Benih, Fakultas Pertanian, Universitas Padjadjaran. Penelitian menggunakan Rancangan Acak Lengkap yang terdiri dari tujuh perlakuan yang diulang lima kali, yaitu: Kontrol (tanpa sitokinin); BAP (0,25 ppm; 0,50 ppm; 0,75 ppm), dan TDZ (0,25 ppm; 0,50 ppm; 0,75 ppm). Hasil dari percobaan menunjukkan bahwa penambahan sitokinin tidak berpengaruh nyata terhadap jumlah tunas dan bobot segar planlet. Media perlakuan kontrol dapat menghasilkan jumlah akar lebih banyak dibandingkan dengan media ditambah sitokinin. Penambahan BAP 0,50 ppm berpengaruh positif terhadap jumlah daun dan dapat menghasilkan runner secara in vitro. Pemberian BAP 0,50 ppm cenderung dapat meningkatkan dan mempercepat produksi bibit tanaman stroberi kultivar Tochiotome.Kata Kunci: Benzylaminopurine (BAP), Thidiazuron (TDZ), Stroberi, Kultur Jaringan AbstractStolon is used for conventional propagation of strawberry, but it is less effective and the quality of the seeds is not good due to the accumulation of disease. In vitro multiplication becomes a solution for the supply of quality seeds in a fast time. The addition of growth regulator cytokinin, such as Benzylaminopurine (BAP) or Thidiazuron (TDZ) can produced the large number of shoot. The objective of this study was to obtain the best type and concentration of cytokinin in the multiplication of strawberry ‘Tochiotome’. The study was conducted at the Plant Tissue Culture Laboratory, Seed Technology, Faculty of Agriculture, Universitas Padjadjaran. This study used a Completely Randomized Design (CRD) with seven treatments and five replications, that were: Control (without cytokinin); BAP (0.25 ppm; 0.50 ppm; 0.75 ppm), and TDZ (0.25 ppm; 0.50 ppm; 0.75 ppm). The results indicated that addition of cytokinin did not affected increasing number of shoots and fresh weightof plantlets. Control media can produce larger number of roots than those containing PGRs, this might be due to the endogenous auxin concentrations found in strawberry plants. Also, cytokinin inhibited root formations process. Plants treated with BAP 0.50 ppm increased for the number of leaves and produced runners in vitro. This study showed application of BAP with 0.50 ppm increased and accelerated the production of strawberry ‘Tochiotome’ seedlings.Keywords: Benzylaminopurine (BAP), Thidiazuron (TDZ), Strawberry, Tissue Culture

Endogenous Auxin Content Contributes to Larger Size of Apple Fruit

Fruit size is an important economic trait that is controlled by multiple genes. However, the regulatory mechanism for fruit size remains poorly understood. A bud sport variety of “Longfeng” (LF) apple (Malus domestica) was identified and named “Grand Longfeng” (GLF). The fruit size of GLF is larger than that of LF, and both varieties are diploid. We found that the cell size in GLF fruit was larger than that of LF. Then, we compared the fruit transcriptomes of the two varieties using RNA-Seq technology. A total of 1166 differentially expressed genes (DEGs) were detected between GLF and LF fruits. The KEGG analysis revealed that the phytohormone pathway was the most enriched, in which most of the DEGs were related to auxin signaling. Moreover, the endogenous auxin levels of GLF fruit were higher than those of LF. The expressions of auxin synthetic genes, including MdTAR1 and MdYUCCA6, were higher in GLF fruit than LF. Collectively, our findings suggest that auxin plays an important role in fruit size development.

Phloroglucinol Improves Direct Rooting of In Vitro Cultured Apple Rootstocks M9 and M26

Advances in micropropagation techniques have helped produce true-to-type clones of many horticulturally important plants. However, several cultivars of apple are difficult to root in vitro. In these cases, adventitious roots are induced together with undesirable formation of callus, which decrease the acclimatization rate of in vitro produced plantlets. In this study, two apple rootstocks, M9 and M26, were subjected to different concentrations of indole-3-butyric acid (IBA) to induce root formation. Although addition of IBA to the medium induced root formation, rhizogenesis was accompanied by the undesirable formation of callus in both cultivars. On the other hand, in gene expression analysis, the indole-3-acetic acid (IAA) synthase genes AAO1 and YUC1 were expressed more highly in M9 than in M26. This suggests that endogenous auxin levels may be higher in M9, which may explain why M9 plantlets are difficult to root and experience high levels of callus formation during propagation. In addition, rooting medium containing 0.1 mg·L−1 IBA was supplemented with different concentrations of phloroglucinol (0, 0.5, 1.0, and 2.0 mM) to examine whether direct rooting efficiency in the M9 could be improved. Addition of 1.0 mM phloroglucinol increased rooting percentage and decreased callus formation in the M9 rootstock. The rootstock M9 is a desirable cultivar but presents a problem with true-to-type direct rooting. Addition of phloroglucinol may improve direct rooting and eliminate callus formation during propagation.