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  • ReviewJune 30, 2017

    3 311 2239

    CRISPR/CAS9 as a Powerful Tool for Crop Improvement

    Jae-Young Song, Marjohn Niño, Franz Marielle Nogoy, Yu-Jin Jung, Kwon-Kyoo Kang, and Yong-Gu Cho

    J Plant Biotechnol 2017; 44(2): 107-114

    https://doi.org/10.5010/JPB.2017.44.2.107

    Abstract

    Abstract : Implementation of crop improvement programs relies on genetic diversity. To overcome the limited occurrence of natural mutations, researchers and breeders applied diverse methods, ranging from conventional crossing to classical biotechnologies. Earlier generations of knockout and gain-of- function technologies often result in incomplete gene disruption or random insertions of transgenes into plant genomes. The newly developed editing tool, CRISPR/Cas9 system, not only provides a powerful platform to efficiently modify target traits, but also broadens the scope and prospects of genome editing. Customized Cas9/guide RNA (gRNA) systems suitable for efficient genomic modification of mammalian cells or plants have been reported. Following successful demonstration of this technology in mammalian cells, CRISPR/Cas9 was successfully adapted in plants, and accumulating evidence of its feasibility has been reported in model plants and major crops. Recently, a modified version of CRISPR/Cas9 with added novel functions has been developed that enables programmable direct irreversible conversion of a target DNA base. In this review, we summarized the milestone applications of CRISPR/ Cas9 in plants with a focus on major crops. We also present the implications of an improved version of this technology in the current plant breeding programs.

  • Research ArticleJune 30, 2017

    4 224 912

    Chlorosis of Ogura-CMS Brassica rapa is due to down-regulation of genes for chloroplast proteins

    Seok-Won Jeong, Hankuil Yi, Hayoung Song, Soo-Seong Lee, Youn-Il Park, and Yoonkang Hur

    J Plant Biotechnol 2017; 44(2): 115-124

    https://doi.org/10.5010/JPB.2017.44.2.115

    Abstract

    Abstract : Cytoplasmic male sterility (CMS) is a maternally inherited trait leading to loss of the ability to produce fertile pollen and is extensively used in hybrid crop breeding. Ogura- CMS was originally generated by insertion of orf138 upstream of atp8 in the radish mitochondrial genome and transferred to Brassica crops for hybrid breeding. Gene expression changes by dysfunctional mitochondria in Ogura-CMS result in pollen developmental defects, but little is known about gene expression patterns in vegetative tissue. To examine the interaction between nuclear and organellar regulation of gene expression, microarray and subsequent gene expression experiments were conducted with leaves of F1 hybrid Chinese cabbage derived from self- incompatible (SI) or Ogura-CMS parents (Brassica rapa ssp. pekinensis). Out of 24,000 genes deposited on a KBGP24K microarray, 66 genes were up-regulated and 26 genes were down-regulated by over 2.5 fold in the CMS leaves. Up-regulated genes included stress-response genes and mitochondrial protein genes, while genes for ascorbic acid biosynthesis and thylakoid proteins were down-regulated. Most of the major component genes for light reactions of photosynthesis were highly expressed in leaves of both SI and CMS plants, but most of the corresponding proteins were found to be greatly reduced in leaves of CMS plants, indicating posttranscriptional regulation. Reduction in thylakoid proteins and chlorophylls led to reduction in photosynthetic efficiency and chlorosis of Ogura- CMS at low temperatures. This research provides a foundation for studying chloroplast function regulated by mitochondrial signal and for using organelle genome introgression in molecular breeding.

  • Research ArticleJune 30, 2017

    5 200 807
    Abstract

    Abstract : Enhanced disease susceptibility1 (EDS1) is a regulator of basal defense responses required for resistance mediated by TIR-NBS-LRR containing R proteins. We identified three transcripts of EDS1-like genes encompassing diverse/ separate expression patterns, based on the transcriptome analysis by Next Generation Sequencing (NGS) of V. flexuosa inoculated with Elsinoe ampelina. These genes were designated VfEDL1 (Vitis flexuosa Enhanced Disease Susceptibility1-like1), VfEDL2 and VfEDL3, and contained 2464, 1719 and 1599 bp, with 1791, 1227 and 1599 bp open reading frames (ORFs), encoding proteins of 596, 408 and 532 amino acids, respectively. The predicted amino acid sequences of all three genes showed the L-family lipase-like domain (class 3 lipase domain), and exhibited a potential lipase catalytic triad, aspartic acid, histidine and serine in the conserved G-X-S-X-G. All three VfEDL genes were upregulated at 1 hpi against the bacterial and fungal pathogens Rizhobiumvitis and E. ampelina, respectively, except VfEDL1, which was downregulated against E. ampelina at all time points. Against E. ampelina, VfEDL2 and VfEDL3 showed downregulated expression at later time points. When evaluated against R. vitis, VfEDL1 showed downregulated expression at all time points after 1 hpi, while VfEDL3 showed upregulation up to 24 hpi. Based on the expression response, all three genes may be involved in plant resistant responses against R. vitis, and VfEDL2 and VfEDL3 show additional resistant responses against E. ampelina infection.

  • Research ArticleJune 30, 2017

    3 227 994
    Abstract

    Abstract : Cudrania tricuspidata Bureau is a widely used medicinal perennial woody plant. For conservation and germplasm utilization of the plant, it is imperative to obtaining information regarding the genetic diversity of the plant populations. Although C. tricuspidata is an important medicinal plant registered in South Korea, no molecular markers are currently available to distinguish Korean-specific ecotypes from other ecotypes of different countries. In this study, we developed single nucleotide polymorphism (SNP) markers derived from chloroplast genomic sequences to identify distinct Korean-specific ecotypes of C. tricuspidata via the amplification refractory mutation system (ARMS)-PCR analyses. Molecular authentication of twelve C. tricuspidata ecotypes from different regions was performed, using DNA sequences in the trnL-F chloroplast intergenic region. The SNP markers developed in this study are useful for rapidly identifying specific C. tricuspidata ecotypes from different regions.

  • Research ArticleJune 30, 2017

    4 196 875

    Expression of NAC transcription factor is altered under intermittent drought stress and re-watered conditions in Hevea brasiliensis

    Lisha P. Luke, M.B. Mohamed Sathik, Molly Thomas, Linu Kuruvilla, and K.V. Sumesh

    J Plant Biotechnol 2017; 44(2): 142-148

    https://doi.org/10.5010/JPB.2017.44.2.142

    Abstract

    Abstract : Drought stress is one of the important factors that restrict the expansion of Hevea brasiliensis cultivation to non- traditional regions experiencing extreme weather conditions. Plants respond to drought stress by triggering expression of several drought responsive genes including transcription factors which in turn trigger expression of various downstream signalling pathways and adaptive networks. Expression of such drought responsive genes may revert back to their original level upon re-watering. However, no reports are available on such phenomenon in Hevea and hence, this study was initiated. For this purpose, NAC transcription factor (NAC tf) was chosen as candidate gene. Its expression levels were monitored under intermittent drought as well as irrigated conditions in two clones (RRII 105 and RRIM 600) of H. brasiliensis with contrasting tolerance level. Copy number of NAC tf was found similar in both the clones. Expression of NAC tf was found highly up-regulated in RRIM 600 (a relatively drought tolerant clone) than in RRII 105 (a relatively drought susceptible clone) throughout the drought incidences which upon re-watering, reached back to its original levels in both the clones. The study indicated the existence of an association between expression of NAC tf and drought tolerance trait exhibited by the tolerant clone RRIM 600. The study also proves the influence of drought and re-watering on the leaf photosynthesis and expression of NAC tf in H. brasiliensis.

  • Research ArticleJune 30, 2017

    13 366 1166

    Overexpression of rice NAC transcription factor OsNAC58 on increased resistance to bacterial leaf blight

    Sang Ryeol Park, Hye Seon Kim, Kyong Sil Lee, Duk-Ju Hwang, Shin-Chul Bae, Il-Pyung Ahn, Seo Hyun Lee, and Sun Tae Kim

    J Plant Biotechnol 2017; 44(2): 149-155

    https://doi.org/10.5010/JPB.2017.44.2.149

    Abstract

    Abstract : Bacterial blight in rice caused by Xanthomonas oryzae pv. oryzae (Xoo) greatly reduces the growth and productivity of this important food crop. Therefore, we sought to increase the resistance of rice to bacterial blight by using a NAC (NAM, ATAF, and CUC) transcription factor, one of the plant-specific transcription factors that is known to be involved in biotic/abiotic stress resistance. By isolating the OsNAC58 gene from rice and analyzing its biological functions related to Xoo resistance, phylogenetic analysis showed that OsNAC58 belongs to group III. To investigate the biological relationship between bacterial leaf blight (BLB) and OsNAC58 in rice, we constructed a vector for overexpression in rice and generated transgenic rice. The expression analysis resulting from use of RT-PCR showed that OsNAC58- overexpressed transgenic rice exhibited higher levels of OsNAC58 expression than wild types. Further, subcellular localization analysis using rice protoplasts showed that the 35S/OsNAC58-SmGFP fusion protein was localized in the nuclei. Thirteen OsNAC58-overexpressed transgenic rice lines, with high expression levels of OsNAC58, showed more resistant to Xoo than did the wild types. Together, these results suggest that the OsNAC58 gene of rice regulates the rice disease resistance mechanism in the nucleus upon invasion of the rice bacterial blight pathogen Xoo.

  • Research ArticleJune 30, 2017

    4 168 737
    Abstract

    Abstract : A greenhouse experiment was conducted for evaluation of ecological effects of transgenic melon plants in the rhizospheric soil in terms of soil properties, enzyme activities and microbial communities. Organic matter content of soil under transgenic melon plants was significantly higher than that of soil with non-transgenic melon plants. Significant variations were observed in organic matter, total P and K in soil cultivation with transgenic melon plants. There were also significant variations in the total numbers of colony forming units of fungi, actinomycetes and bacteria between soils treated with transgenic and non-transgenic melon plants. Transgenic and non-transgenic melon significantly enhanced several enzymes activities including urease, acid phosphatase, alkalin phosphatase, arysulphtase, β glucosidase, dehydrogenase, protease and catalase. Soil polyphenoloxidase activity of T1 transgenic melon was lower than that of T0 transgenic melon and a non-melon plant during the same period. The first generation transgenic melon plants (T0) showed significantly greater (p<0.05) effect on the activitiy of arylsulfatase, which increased from 2.540x106 CFU g-1 (control) to 19.860x106 CFU g-1 (T0). These results clearly indicated that transgenic melon might change microbial communities, enzyme activities and soil chemical properties.

  • Research ArticleJune 30, 2017

    5 242 1537

    Development of an efficient protocol for high-frequency regeneration system in Hibiscus syriacus L.

    Sang-Gyu Seo, Sun-Hee Ryu, Yang Zhou, and Sun-Hyung Kim

    J Plant Biotechnol 2017; 44(2): 164-170

    https://doi.org/10.5010/JPB.2017.44.2.164

    Abstract

    Abstract : A rapid and efficient in vitro regeneration system was established for Hibiscus syriacus L. The successful regeneration protocol employs induction of shoot organogenesis on leaf, petiole, and root explants. Among the various plant growth regulators evaluated, thidiazuron (TDZ) was the most effective for inducing rapid shoot formation. Most efficient shoot regeneration frequency was obtained from Murashige and Skoog (MS) media containing 0.01 mg/L TDZ. Regeneration efficiency was highest in the roots, and lowest in the leaves. A combination of 0.01 mg/L TDZ with benzyladenine (BAP) markedly improved the frequency of shoot differentiation from the root (up to 98%) and petiole (up to 88%) explants. Furthermore, leaf and petiole explants showed the highest frequency of shoot induction in half-strength MS media containing 0.01 mg/L TDZ and 1.0 mg/L BAP, while root explants formed the greatest number of shoots when 0.01 mg/L TDZ and 0.1 mg/L BAP were added to half-strength MS media. Although the frequency of shoot differentiation from leaf explants was only 50%, the leaf is considered the most efficient plant organ for use in tissue culture because leaves are easier to obtain than roots and petioles. Our findings show that various organs of H. syriacus can be used for plant regeneration, and the protocol developed in this study may be applicable in the horticulture industry.

  • Research ArticleJune 30, 2017

    0 214 916
    Abstract

    Abstract : A callus-mediated regeneration protocol for sea-milkwort, an endangered coastal plant species in South Korea, is reported here. The explants of in vitro-plantlets generated from a node culture revealed distinguishable responses in callus induction depending on genotype, explant source, light condition, and 2,4-D concentration. Especially, continuous darkness exclusively facilitated callus induction from explants prior to other treatments. The calli initiated on the media with 2,4-D ranging from 0.1 mg/L to 3.0 mg/L in the dark vigorously proliferated when sub- cultured on the same media in continuous darkness. Given 1.0 mg/L zeatin in addition to darkness to the calli of the ‘Pistachio’ genotype, normal adventitious shoots were only regenerated from nodular structures that formed earlier from the calli at the frequency of 24.4 percent. Regenerated shoots easily grew into plantlets with roots and green color on a phytohormone-free MS medium under lighted condition, that were used for node culture as plant materials. Node culture effectively multiplied plantlets in accordance with protocol by Bae et al. (2016). Acclimatized plantlet clusters developed mature plant clusters under inland environment, followed by flowering the following April. Results were merged with node culture protocol suggested by Bae et al. (2016), which, as an in vitro propagation system for sea-milkwort, may contribute to natural habitat restoration.

  • Research ArticleJune 30, 2017

    10 303 1024

    Sugar content analysis and expression profiling of sugar related genes in contrasting Strawberry (Fragaria × ananassa) cultivars

    Ashokraj Shanmugam, Mohammad Rashed Hossain, Sathishkumar Natarajan, Hee-Jeong Jung, Jae-Young Song, Hoy-Taek Kim, and Ill-Sup Nou

    J Plant Biotechnol 2017; 44(2): 178-190

    https://doi.org/10.5010/JPB.2017.44.2.178

    Abstract

    Abstract : Fragaria × ananassa, a strawberry evolved from hybridization between F. virginiana and F. chiloensis, is a globally cultivated and consumed fruit crop valued for its flavor and nutritional value. Flavor and quality of fruits are determined by factors such as sugars and organic acids present during fruit development. These characteristics are highly subjective in different genotypes and affected by various environmental factors. In this study, we analyzed contents of major sugar compounds including fructose, glucose and sucrose by HPLC analysis in four cultivars namely, Maehyang, Seolhyang, Festival and Sweet Charlie. We identified 55 genes related to fructose, glucose, sucrose and soluble sugar regulation whose expression were analyzed in four cultivars at three developmental stages of the fruit namely, green, white and ripened stages. Expression of these genes across these progressive fruit developmental stages varied among cultivars. Among the 55 genes, genes FaFru3, FaSuc11 and FaGlu8 revealed differential patterns of expression along developmental stages of the fruit in high and low sugar-containing genotypes, respectively and may be putative candidates for sugar content in strawberries. Expression of genes are discussed with regard to corresponding sugar content in these genotypes. Further analysis and application of these genes may be valuable in developing high sugar containing cultivars via marker-assisted breeding.

JPB
Vol 51. 2024

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Plant Biotechnology

pISSN 1229-2818
eISSN 2384-1397
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