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16. Comparative genomics of Ceriporiopsis subvermispora and Phanerochaete chrysosporium provide insight into selective ligninolysis

Author

Fernandez-Fueyo, Elena, Ruiz-Duenas, Francisco J., Ferreira, Patrica, Floudas, Dimitrios, HIbbett, David S., Canessa, Paulo, Larrondo, Luis F., James, Tim Y., Seelenfreund, Daniela, Lobos, Sergio, Polanco, Ruben, Tello, Mario, Honda, Yoichi, Watanabe, Takahito, Watanabe, Takashi, Ryu, Jae San, Kubicek, Christian P., Schmoll, Monika, Gaskell, Jill, Hammel, Kenneth E., John, Franz J. St., Wymelenberg, Amber Vanden, Sabat, Grzegorz, BonDurant, Sandra Splinter, Syed, Khajamohiddin, Yadav, Jagjit S., Doddapaneni, Harshavardhan, Subramanian, Venkataramanan, Lavin, Jose L., Oguiza, Jose A., Perez, Gumer, Pisabarro, Antonio G., Ramirez, Lucia, Santoyo, Francisco, Master, Emma, Coutinho, Pedro M., Henrissat, Bernard, Lombard, Vincent, Magnuson, Jon Karl, Kues, Ursula, Hori, Chiaki, Igarashi, Kiyohiko, Samejima, Masahiro, Held, Benjamin W., Barry, Kerrie W., LaButti, Kurt M., Lapidus, Alla, Lindquist, Erika A., Lucas, Susan M., Riley, Robert, Salamov, Asaf A., Hoffmeister, Dirk, Schwenk, Daniel, Hadar, Yitzhak, Yarden, Oded, Vries, Ronald P. de, Wiebenga, Ad, Stenlid, Jan, Eastwood, Daniel, Grigoriev, Igor V., Berka, Randy M., Blanchette, Robert A., Kersten, Phil, Martinez, Angel T., Vicuna, Rafael, and Cullen, Dan

Abstract

Efficient lignin depolymerization is unique to the wood decay basidiomycetes, collectively referred to as white rot fungi. Phanerochaete chrysosporium simultaneously degrades lignin and cellulose, whereas the closely related species, Ceriporiopsis subvermispora, also depolymerizes lignin but may do so with relatively little cellulose degradation. To investigate the basis for selective ligninolysis, we conducted comparative genome analysis of C. subvermispora and P. chrysosporium. Genes encoding manganese peroxidase numbered 13 and five in C. subvermispora and P. chrysosporium, respectively. In addition, the C. subvermispora genome contains at least seven genes predicted to encode laccases, whereas the P. chrysosporium genome contains none. We also observed expansion of the number of C. subvermispora desaturase-encoding genes putatively involved in lipid metabolism. Microarray-based transcriptome analysis showed substantial up-regulation of several desaturase and MnP genes in wood-containing medium. MS identified MnP proteins in C. subvermispora culture filtrates, but none in P. chrysosporium cultures. These results support the importance of MnP and a lignin degradation mechanism whereby cleavage of the dominant nonphenolic structures is mediated by lipid peroxidation products. Two C. subvermispora genes were predicted to encode peroxidases structurally similar to P. chrysosporium lignin peroxidase and, following heterologous expression in Escherichia coli, the enzymes were shown to oxidize high redox potential substrates, but not Mn2. Apart from oxidative lignin degradation, we also examined cellulolytic and hemicellulolytic systems in both fungi. In summary, the C. subvermispora genetic inventory and expression patterns exhibit increased oxidoreductase potential and diminished cellulolytic capability relative to P. chrysosporium.

Published
2012

17. Development of a stable SCAR marker for rapid identification of Ganoderma lucidum Hunong 5 cultivar using DNA pooling method and inter-simple sequence repeat markers

Author

Wen-zheng CHAO, Chuan-hong TANG, Jing-song ZHANG, Ling YU, and Honda Yoichi

Subjects
DNA pooling, Ganoderma lucidum, Hunong 5 cultivar, ISSR marker, SCAR marker, Agriculture (General), S1-972
Abstract

The cultivar Ganoderma lucidum Hunong 5 was obtained using cross-breeding. Hunong 5 has high commercial value due to its high polysaccharide and triterpene content. This is the first report of using a DNA pooling method to develop a stable sequence characterized amplified region (SCAR) marker for rapid identification of the G. lucidum Hunong 5 cultivar. The SCAR marker was developed by first generating and sequencing a distinctive inter simple sequence repeat (ISSR) fragment (882 bp) from G. lucidum Hunong 5 cultivar. A stable SCAR primer pair GLH5F/GLH5R were obtained to identify the cultivar and the SCAR marker is a DNA fragment of 773 bp.

Published
2018
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30. Double-gene targeting with preassembled Cas9 ribonucleoprotein for safe genome editing in the edible mushroom Pleurotus ostreatus.

Author

Boontawon, Tatpong, Nakazawa, Takehito, Choi, Yeon-Jae, Ro, Hyeon-Su, Oh, Minji, Kawauchi, Moriyuki, Sakamoto, Masahiro, and Honda, Yoichi

Subjects
EDIBLE mushrooms, PLEUROTUS ostreatus, CRISPRS, GENOME editing, DNA sequencing, GENE targeting
Abstract

CRISPR/Cas9 has potential for efficient molecular breeding. Recently, a foreign-DNA-free gene-targeting technology was established by introducing a preassembled Cas9 ribonucleoprotein (RNP) complex into the oyster mushroom Pleurotus ostreatus. However, the target gene was restricted to such a gene like pyrG , since screening of a genome-edited strain was indispensable and could be performed via examination of 5-fluoroorotic acid (5-FOA) resistance caused by the disruption of the target gene. In this study, we simultaneously introduced the Cas9 RNP complex targeting fcy1 , a mutation that conferred P. ostreatus resistance to 5-fluorocytosine (5-FC), together with that targeting pyrG. A total of 76 5-FOA resistant strains were isolated during the first screening. Subsequently, a 5-FC resistance examination was conducted, and three strains exhibited resistance. Genomic PCR experiments followed by DNA sequencing revealed that mutations were successfully introduced into fcy1 and pyrG in the three strains. The results indicated that double gene-edited mutants could be obtained in one experiment employing 5-FOA resistance screening for strains with Cas9 RNP incorporation. This work may pave the way for safe CRISPR/Cas9 technology to isolate mutant strains in any gene of interest without an ectopic marker gene. [ABSTRACT FROM AUTHOR]

Published
2023
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41. Additional file 1 of Efficient genome editing with CRISPR/Cas9 in Pleurotus ostreatus

Author

Tatpong Boontawon, Nakazawa, Takehito, Inoue, Chikako, Osakabe, Keishi, Moriyuki Kawauchi, Sakamoto, Masahiro, and Honda, Yoichi

Abstract

Additional file 1: Fig. S1. A plasmid map of the pCcPef3-126 plasmid shows the components of the construct. Fig. S2. Agarose gel electrophoresis. Genomic PCR experiments examining/verifying fcy1 (A) or pyrG (B) mutation. (C) Genomic PCR experiments confirming gene replacement in the 5-FC-resistant strains obtained after introducing the pCcPef3-126-fcy1sg2 plasmid with the donor DNA templates. Lane WT, the parental strain PC9 (A and B) and 20b (C) as a positive control; Lanes 1–10 (A and B), 5-FC- and 5-FOA- resistant strains, respectively; Lanes 1–17 (C), 5-FC-resistant strains; Lane M, a 1 kb DNA ladder plus (0.1–10.0 kb), or a 100-bp molecular weight marker (0.1–1.5 kb). For more details regarding the estimated lengths of the PCR products amplified from the genome, please see Table S2. Table S1. Primer pairs used in this study. Table S2. Estimated lengths of the PCR fragments that were amplified from each strain.

Published
2021
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