Although most eukaryotes have both MSH4 and MSH5 orthologs, Neurospora was initially thought to lack msh-4. We have deleted the most likely msh-4 candidate and observed a delay in the sexual cycle, disruption to meiosis and a reduction in fertility. Deletion is dominant, showing msh-4 is subject to MSUD. We conclude that Neurospora has a MSH4 ortholog and that it may have remained undetected because of an unusually high number of introns. Creative Commons License This work is licensed under a Creative Commons Attribution-Share Alike 4.0 License. This regular paper is available in Fungal Genetics Reports: http://newprairiepress.org/fgr/vol53/iss1/2
{"title":"Neurospora msh4 ortholog confirmed by split-marker deletion","authors":"Sue Conway, F. Bowring, J. Yeadon, D. Catcheside","doi":"10.4148/1941-4765.1105","DOIUrl":"https://doi.org/10.4148/1941-4765.1105","url":null,"abstract":"Although most eukaryotes have both MSH4 and MSH5 orthologs, Neurospora was initially thought to lack msh-4. We have deleted the most likely msh-4 candidate and observed a delay in the sexual cycle, disruption to meiosis and a reduction in fertility. Deletion is dominant, showing msh-4 is subject to MSUD. We conclude that Neurospora has a MSH4 ortholog and that it may have remained undetected because of an unusually high number of introns. Creative Commons License This work is licensed under a Creative Commons Attribution-Share Alike 4.0 License. This regular paper is available in Fungal Genetics Reports: http://newprairiepress.org/fgr/vol53/iss1/2","PeriodicalId":12490,"journal":{"name":"Fungal Genetics Reports","volume":"29 1","pages":"5-8"},"PeriodicalIF":0.0,"publicationDate":"2006-12-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"73708938","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
This bibliography attempts to cover genetical and biochemical publications on Aspergillus nidulans and also includes selected references to related species and topics. Entries have been checked as far as possible, but please tell me of any errors and omissions. Authors are kindly requested to send a copy of each article to the FGSC for its reprint collection. This bibliography is available in Fungal Genetics Reports: https://newprairiepress.org/fgr/vol53/iss1/13 ASPERGILLUS BIBLIOGRAPHY 2006 This bibliography attempts to cover genetical and biochemical publications on Aspergillus nidulans and also includes selected references to related species and topics. Entries have been checked as far as possible, but please tell me of any errors and omissions. Authors are kindly requested to send a copy of each article to the FGSC for its reprint collection. John Clutterbuck. Institute of Biomedical and Life Sciences, Anderson College, University of Glasgow, Glasgow G11 6NU, Scotland, UK. Email: j.clutterbuck@bio.gla.ac.uk
{"title":"Aspergillus Bibliography 2006","authors":"J. Clutterbuck","doi":"10.4148/1941-4765.1116","DOIUrl":"https://doi.org/10.4148/1941-4765.1116","url":null,"abstract":"This bibliography attempts to cover genetical and biochemical publications on Aspergillus nidulans and also includes selected references to related species and topics. Entries have been checked as far as possible, but please tell me of any errors and omissions. Authors are kindly requested to send a copy of each article to the FGSC for its reprint collection. This bibliography is available in Fungal Genetics Reports: https://newprairiepress.org/fgr/vol53/iss1/13 ASPERGILLUS BIBLIOGRAPHY 2006 This bibliography attempts to cover genetical and biochemical publications on Aspergillus nidulans and also includes selected references to related species and topics. Entries have been checked as far as possible, but please tell me of any errors and omissions. Authors are kindly requested to send a copy of each article to the FGSC for its reprint collection. John Clutterbuck. Institute of Biomedical and Life Sciences, Anderson College, University of Glasgow, Glasgow G11 6NU, Scotland, UK. Email: j.clutterbuck@bio.gla.ac.uk","PeriodicalId":12490,"journal":{"name":"Fungal Genetics Reports","volume":"78 1","pages":"13"},"PeriodicalIF":0.0,"publicationDate":"2006-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"88571992","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
David Perkins pointed out to me that research articles dealing with Neurospora do not always include the word Neurospora. There are instances where the fungus is named Chrysonilia sp. (usually C. sitophila), referring to the imperfect stage. Here follows a list of such citations, with a few caveats. In many cases, the journals in which these articles appeared are not available to me, so I generally have not confirmed these citations. Some may be only single-page abstracts. I have not attempted to find all citations using Monilia sitophila, a name in use before Chrysonilia. Creative Commons License This work is licensed under a Creative Commons Attribution-Share Alike 4.0 License. This bibliography is available in Fungal Genetics Reports: http://newprairiepress.org/fgr/vol52/iss1/10 Supplement to Neurospora bibliographies: Chrysonilia citations Craig Wilson – Northwest Fisheries Science Center, Seattle WA 98112 David Perkins pointed out to me that research articles dealing with Neurospora do not always include the word Neurospora. There are instances where the fungus is named Chrysonilia sp. (usually C. sitophila), referring to the imperfect stage. Here follows a list of such citations, with a few caveats. In many cases, the journals in which these articles appeared are not available to me, so I generally have not confirmed these citations. Some may be only single-page abstracts. I have not attempted to find all citations using Monilia sitophila, a name in use before Chrysonilia. 1. Abdel-Rahman, T. M. A., M. Salama A-A, M. I. A. Ali, and N. A. H. Tharwat. 1990. Fibrinolytic activity of some fungi isolated from self-heated composted fertilizer. Botanical Magazine Tokyo 103:313-324. 2. Ahmad, M. S., and M. A. Malik. 1997. Formulation of a synthetic medium for the production of antifungal antibiotic from Bacillus subtilis AECL 69. Pakistan Journal of Zoology 29:15-21. 3. Aidoo, K. E., A. Anderton, and K. A. Milligan. 1995. A 2-year survey of the airborne mycoflora in a hospital environment. International Journal of Environmental Health Research 5:223-228. 4. Alvarez-Rodriguez, M. L., L. Lopez-Ocana, J. M. Lopez-Coronado, E. Rodriguez, M. J. Martinez, G. Larriba, and J.-J. R. Coque. 2002. Cork taint of wines: Role of the filamentous fungi isolated from cork in the formation of 2,4,6-trichloroanisole by O methylation of 2,4,6-trichlorophenol. Applied and Environmental Microbiology 68:58605869. 5. Anaissie, E. J., S. L. Stratton, M. C. Dignani, C.-K. Lee, R. C. Summerbell, J. H. Rex, T. P. Monson, and T. J. Walsh. 2003. Pathogenic molds (including Aspergillus species) in hospital water distribution systems: A 3-year prospective study and clinical implications for patients with hematologic malignancies. Blood 101:2542-2546. 6. Birbir, M., O. Ozyaral, C. Johansson, and A. Ilgaz. 1994. Mold strains isolated from unfinished and finished leather goods and shoes. Journal of the American Leather Chemists Association 89:14-19. 7. Bollen, G. J., and M. M. Verf.
David Perkins向我指出,关于神经孢子虫的研究文章并不总是包含神经孢子虫这个词。有些情况下,真菌被命名为Chrysonilia sp.(通常是C. sitophila),指的是不完美的阶段。以下是这些引用的列表,并附有一些注意事项。在很多情况下,这些文章发表的期刊对我来说是不可用的,所以我通常没有确认这些引用。有些可能只有单页摘要。我并没有试图找到所有使用Monilia sitophila的引文,这个名字在Chrysonilia之前就已经使用了。本作品采用知识共享署名-相同方式共享4.0许可协议。这个参考书目可在真菌遗传学报告:http://newprairiepress.org/fgr/vol52/iss1/10补充神经孢子虫参考书目:金丝虫引文克雷格·威尔逊-西北渔业科学中心,西雅图华盛顿州98112大卫·珀金斯向我指出,研究文章处理神经孢子虫并不总是包括神经孢子虫这个词。有些情况下,真菌被命名为Chrysonilia sp.(通常是C. sitophila),指的是不完美的阶段。以下是这些引用的列表,并附有一些注意事项。在很多情况下,这些文章发表的期刊对我来说是不可用的,所以我通常没有确认这些引用。有些可能只有单页摘要。我并没有试图找到所有使用Monilia sitophila的引文,这个名字在Chrysonilia之前就已经使用了。Abdel-Rahman, t.m.a., M. Salama A-A, M. i.a. Ali和N. a.h. Tharwat, 1990。自热堆肥中分离真菌的纤溶活性。植物杂志东京103:313-324。2. Ahmad, m.s.和m.a. Malik. 1997。从枯草芽孢杆菌aecl69生产抗真菌抗生素的合成培养基的配方。巴基斯坦动物学杂志29:15-21。3.Aidoo, k.e., A. Anderton和K. A. Milligan, 1995。医院环境中空气传播菌群的2年调查。国际环境卫生研究杂志(5):223-228。4. 阿尔瓦雷斯-罗德里格斯,M. L.洛佩斯-奥卡纳,J. M.洛佩斯-科罗纳多,E.罗德里格斯,M. J.马丁内斯,G.拉里巴和J. J.。科克,2002。葡萄酒的软木污染:从软木中分离的丝状真菌在2,4,6-三氯苯酚O甲基化形成2,4,6-三氯苯甲醚中的作用。应用与环境微生物学68:58605869。5. 阿纳西,E. J., S. L. Stratton, M. C. Dignani, c - k。李,R. C. Summerbell, J. H. Rex, T. P. Monson和T. J. Walsh. 2003。医院配水系统中的病原菌(包括曲霉菌种类):一项为期3年的前瞻性研究及其对血液恶性肿瘤患者的临床意义血101:2542 - 2546。6. M. Birbir, O. Ozyaral, C. Johansson和A. Ilgaz. 1994。从未完成和完成的皮革制品和鞋子中分离出霉菌菌株。美国皮革化学家协会杂志89:14-19。7. 波伦,g.j.和m.m. Verf. 1978。温室空气中作为烟曲霉孢子来源的蒸盆栽土壤。荷兰植物学报27:152。8. 坎波斯,V., E. Salas, N. Duran, J. Rodriquez和J. Baeza. 1986。从铁三角菌中分离分离纤维素水解菌。Boletín micológico(智利)2:1619. P. D. Carranco, A. Hernandez, P. Rivera和I. Rosas, 1984。墨西哥州废物稳定池系统中的土壤和水生真菌。水、空气与土壤污染(23):249-256。新草原出版社2017年出版。卡瓦略,M. M. D., M. F. S.特谢拉,E.埃斯波西托,A.马丘卡,A.费拉兹和N.杜兰,1992。亚马逊木质纤维素材料。腐烂月桂树和雪松的真菌筛选。应用生物化学与生物技术37:33-41。11. ceneno, S.和M. A. Calvo. 2001。葡萄酒软木塞中分离微生物的酶活性。Microbios 106:69 - 73。12. 查普曼,J.,和H. Burge. 1987。Monilia和Candida不一样。过敏年鉴58:286。13. 查普曼,J.,和H. Burge. 1987。念珠菌和念珠菌是两种非常不同的真菌。变态反应与临床免疫学杂志79:207。14. 考克斯,L. J., B. Caicedo, V. Vanos, E. Heck, S. Hofstaetter, J. L. Cordier. 1987。一些厄瓜多尔发酵饮料的目录,附有其微生物区系的注释。中国生物医学工程学报(英文版)(3):143-154。15. P. Danesh, F. M. V. Caldas, J. J. F. Marques和M. V. S. Romao, 1997。葡萄牙“正常”软木塞和“绿色”软木塞生产过程中的真菌群。应用微生物学报82:689-694。16. de Lillo, E. 1997。膜马虱寄主偏好的观察(蜱螨亚纲:拟螨科)。昆虫学(巴里)31:7-12。17. Dezotti, M., L. H. Innocentini-Mei, N. Duran. 1995。二氧化硅固定化酶催化桉叶废液中氯木质素的去除。生物技术学报43(3):161-167。18. 多罗日金和费多罗夫。 1987. 挪威云杉溃疡病菌群的研究。医学病理学杂志,21:347-352。19. 德拉戈尼,I. 1979。嗜黑念珠菌、黑根霉和常青霉。工业食品,18:37 74-376。20.杜兰,n.n .布朗伯格和A.昆兹,2001。辣根过氧化物酶转化戊曲醇的动力学研究。无机生物化学学报84:279286。21. 杜兰,N., M. Dezotti, J. Rodriguez, 1991。生物质photochemistry.15。硫酸盐废水的光漂白和生物漂白。光化学与光生物学报[a](2):269-279。22. N.杜兰、I.费雷尔和J.罗德里格斯1987。嗜sitophila (Tfb-27441菌株)木质素酶。应用生物化学与生物技术(16):157-167。23. 杜兰,N., J. L. Reyes, J. Baeza和V. Campos, 1988。生物质光化学12稻壳化学光化学预处理及其真菌降解。生物技术与生物工程32:564-568。http://newprairiepress.org/fgr/vol52/iss1/10 DOI: 10.4148/1941-4765.1129杜兰,N., J. Rodriguez, V. Campos, A. Ferraz, J. L. Reyws, J. Amaya-Farfan, E. Esposito, F. Adao和J. Baeza. 1994。由木质纤维素材料生产的单细胞蛋白质量的子囊菌嗜sitonilia (TFB-27441菌株)。微生物学刊25:31-36。25. 杜兰,N., J. Rodriguez, E. Gomez, V. Campos和J. Ba
{"title":"Supplement to Neurospora bibliographies: Chrysonilia citations","authors":"Craig Wilson","doi":"10.4148/1941-4765.1129","DOIUrl":"https://doi.org/10.4148/1941-4765.1129","url":null,"abstract":"David Perkins pointed out to me that research articles dealing with Neurospora do not always include the word Neurospora. There are instances where the fungus is named Chrysonilia sp. (usually C. sitophila), referring to the imperfect stage. Here follows a list of such citations, with a few caveats. In many cases, the journals in which these articles appeared are not available to me, so I generally have not confirmed these citations. Some may be only single-page abstracts. I have not attempted to find all citations using Monilia sitophila, a name in use before Chrysonilia. Creative Commons License This work is licensed under a Creative Commons Attribution-Share Alike 4.0 License. This bibliography is available in Fungal Genetics Reports: http://newprairiepress.org/fgr/vol52/iss1/10 Supplement to Neurospora bibliographies: Chrysonilia citations Craig Wilson – Northwest Fisheries Science Center, Seattle WA 98112 David Perkins pointed out to me that research articles dealing with Neurospora do not always include the word Neurospora. There are instances where the fungus is named Chrysonilia sp. (usually C. sitophila), referring to the imperfect stage. Here follows a list of such citations, with a few caveats. In many cases, the journals in which these articles appeared are not available to me, so I generally have not confirmed these citations. Some may be only single-page abstracts. I have not attempted to find all citations using Monilia sitophila, a name in use before Chrysonilia. 1. Abdel-Rahman, T. M. A., M. Salama A-A, M. I. A. Ali, and N. A. H. Tharwat. 1990. Fibrinolytic activity of some fungi isolated from self-heated composted fertilizer. Botanical Magazine Tokyo 103:313-324. 2. Ahmad, M. S., and M. A. Malik. 1997. Formulation of a synthetic medium for the production of antifungal antibiotic from Bacillus subtilis AECL 69. Pakistan Journal of Zoology 29:15-21. 3. Aidoo, K. E., A. Anderton, and K. A. Milligan. 1995. A 2-year survey of the airborne mycoflora in a hospital environment. International Journal of Environmental Health Research 5:223-228. 4. Alvarez-Rodriguez, M. L., L. Lopez-Ocana, J. M. Lopez-Coronado, E. Rodriguez, M. J. Martinez, G. Larriba, and J.-J. R. Coque. 2002. Cork taint of wines: Role of the filamentous fungi isolated from cork in the formation of 2,4,6-trichloroanisole by O methylation of 2,4,6-trichlorophenol. Applied and Environmental Microbiology 68:58605869. 5. Anaissie, E. J., S. L. Stratton, M. C. Dignani, C.-K. Lee, R. C. Summerbell, J. H. Rex, T. P. Monson, and T. J. Walsh. 2003. Pathogenic molds (including Aspergillus species) in hospital water distribution systems: A 3-year prospective study and clinical implications for patients with hematologic malignancies. Blood 101:2542-2546. 6. Birbir, M., O. Ozyaral, C. Johansson, and A. Ilgaz. 1994. Mold strains isolated from unfinished and finished leather goods and shoes. Journal of the American Leather Chemists Association 89:14-19. 7. Bollen, G. J., and M. M. Verf.","PeriodicalId":12490,"journal":{"name":"Fungal Genetics Reports","volume":"15 1","pages":"10"},"PeriodicalIF":0.0,"publicationDate":"2005-12-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"85754162","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Boil-mediated lysis of Neurospora conidia (Boil-prep) is an extremely rapid, convenient and useful technique to obtain sufficient genomic DNA template for PCR amplification. We routinely use this technique for screening molecular markers, sequencing, and preliminary confirmation of transformants. Creative Commons License This work is licensed under a Creative Commons Attribution-Share Alike 4.0 License. This short communications is available in Fungal Genetics Reports: http://newprairiepress.org/fgr/vol52/iss1/7 24 Fungal Genetics Newsletter Short Communications Reliable PCR amplification from Neurospora crassa genomic DNA obtained from conidia Steven T. Henderson, Graham A. Eariss and David E. A. Catcheside – School of Biological Sciences, Flinders University, Bedford Park, SA 5042, Australia. Fungal Genetics Newsletter 52:24 Boil-mediated lysis of Neurospora conidia (Boil-prep) is an extremely rapid, convenient and useful technique to obtain sufficient genomic DNA template for PCR amplification. We routinely use this technique for screening molecular markers, sequencing, and preliminary confirmation of transformants. However, we have observed periods when successful PCR amplification from Boil-preps has been erratic, hampering the efficacy of this technique. As lysis of conidia results in the liberation of DNA and other cellular components, we reasoned the inconsistent results may have been due to inhibition and/or degradation of the Taq DNA polymerase by cellular material present in the lysis solution. We tried various DNA polymerases and found the reliability of PCR amplification of DNA from Boil-prep template is largely dependant on the specific enzyme used with some Taq polymerases producing <5% successful amplifications. In our hands, Red Hot DNA polymerase from ABgene (cat#AB-0406) facilitates consistent PCR ® amplification (>90%) from Boil-prep template DNA. We routinely use Red Hot DNA polymerase to amplify PCR products up ® to 1.35kb from Boil-prep DNA following the procedure described below (modified from Yeadon and Catcheside 1996). Boil-prep method: A wet loop of 3 7 day old conidia is transferred to 100ìl of sterile Tris-EDTA (pH 8.0) in a 1.5ml microfuge tube and vortexed briefly. The conidial suspension is placed in a boiling waterbath for 10 minutes and then on ice for 5 minutes. The cellular debris is pelleted by centrifugation at 13000 RPM in a benchtop centrifuge for 5 minutes. 70ul of supernatant is transferred to a new 1.5ml microfuge tube and stored at -20oC. 2-5ul of Boil-prep DNA solution is used as a template in a 50ul PCR reaction with 0.5U of Red Hot DNA polymerase. Figure 1. Comparison of PCR amplicons from Boilprep genomic DNA (BP), genomic DNA (G) (prepared by the method of Irelan et al. 1993), plasmid DNA (P) and no DNA (-ve) templates. Size marker (M) = 100bp DNA ladder (New England Biolabs) with a 500bp reference band that contains 97ng of DNA. Each sample lane contains 6ul of a 50ul PCR reaction. DNA yields fro
煮沸介导的分生神经孢子菌裂解(Boil-prep)是一种非常快速、方便和有用的技术,可以获得足够的基因组DNA模板进行PCR扩增。我们经常使用这种技术筛选分子标记,测序和初步确认转化子。本作品采用知识共享署名-相同方式共享4.0许可协议。这篇简短的通讯可在真菌遗传学报告中获得:http://newprairiepress.org/fgr/vol52/iss1/7 24真菌遗传学通讯简短通讯从分生孢子中获得的粗神经孢子虫基因组DNA进行可靠的PCR扩增。Steven T. Henderson, Graham A. Eariss和David E. A. Catcheside -弗林德斯大学生物科学学院,Bedford Park, SA 5042,澳大利亚。煮沸介导的分生神经孢子菌裂解(Boil-prep)是一种非常快速、方便和有用的技术,可以获得足够的基因组DNA模板进行PCR扩增。我们经常使用这种技术筛选分子标记,测序和初步确认转化子。然而,我们观察到,当成功的PCR扩增从沸腾准备已经不稳定的时期,阻碍了这项技术的效力。由于分生孢子的裂解会导致DNA和其他细胞成分的释放,我们推断,不一致的结果可能是由于裂解溶液中存在的细胞物质抑制和/或降解了Taq DNA聚合酶。我们尝试了各种DNA聚合酶,发现从煮沸准备模板中扩增DNA的可靠性在很大程度上取决于所使用的特定酶,一些Taq聚合酶从煮沸准备模板DNA中产生90%)。我们常规使用Red Hot DNA聚合酶根据以下程序(根据Yeadon和Catcheside 1996年修改)从煮沸准备的DNA中扩增PCR产物至1.35kb。煮沸准备法:将37日龄的分生孢子湿循环转移到1.5ml的无菌Tris-EDTA (pH 8.0) 100ìl中,并短暂旋转。将分生孢子悬浮液置于沸水浴中10分钟,然后放在冰上5分钟。细胞碎片在台式离心机中以13000 RPM离心5分钟制成球团。将70ul上清转移到新的1.5ml微管中,保存在-20℃。以2-5ul的煮沸DNA溶液为模板,用0.5U的Red Hot DNA聚合酶进行50ul的PCR反应。图1所示。Boilprep基因组DNA (BP)、基因组DNA (G) (Irelan et al. 1993方法制备)、质粒DNA (P)和无DNA (-ve)模板PCR扩增子的比较。尺寸标记(M) = 100bp DNA阶梯(New England Biolabs),参考带为500bp,包含97ng DNA。每个样品道含有6ul的50ul PCR反应。沸水制备PCR的DNA产量通常为100 - 10ng/ul。注:所有pcr均在一次实验中进行,循环条件优化为扩增1350bp产物。参考文献:Irelan, J., V. Miao和e.u. Selker, 1993。粗神经孢子虫的小尺度DNA准备。真菌麝猫。新闻40:24。Yeadon, p.j.和d.e.a. Catcheside, 1996。从神经孢子菌培养物中快速制备PCR模板的方法。真菌麝猫。新闻。43:71。新草原出版社2017年出版
{"title":"Reliable PCR amplification from Neurospora crassa genomic DNA obtained from conidia","authors":"S. Henderson, G. Eariss, D. Catcheside","doi":"10.4148/1941-4765.1126","DOIUrl":"https://doi.org/10.4148/1941-4765.1126","url":null,"abstract":"Boil-mediated lysis of Neurospora conidia (Boil-prep) is an extremely rapid, convenient and useful technique to obtain sufficient genomic DNA template for PCR amplification. We routinely use this technique for screening molecular markers, sequencing, and preliminary confirmation of transformants. Creative Commons License This work is licensed under a Creative Commons Attribution-Share Alike 4.0 License. This short communications is available in Fungal Genetics Reports: http://newprairiepress.org/fgr/vol52/iss1/7 24 Fungal Genetics Newsletter Short Communications Reliable PCR amplification from Neurospora crassa genomic DNA obtained from conidia Steven T. Henderson, Graham A. Eariss and David E. A. Catcheside – School of Biological Sciences, Flinders University, Bedford Park, SA 5042, Australia. Fungal Genetics Newsletter 52:24 Boil-mediated lysis of Neurospora conidia (Boil-prep) is an extremely rapid, convenient and useful technique to obtain sufficient genomic DNA template for PCR amplification. We routinely use this technique for screening molecular markers, sequencing, and preliminary confirmation of transformants. However, we have observed periods when successful PCR amplification from Boil-preps has been erratic, hampering the efficacy of this technique. As lysis of conidia results in the liberation of DNA and other cellular components, we reasoned the inconsistent results may have been due to inhibition and/or degradation of the Taq DNA polymerase by cellular material present in the lysis solution. We tried various DNA polymerases and found the reliability of PCR amplification of DNA from Boil-prep template is largely dependant on the specific enzyme used with some Taq polymerases producing <5% successful amplifications. In our hands, Red Hot DNA polymerase from ABgene (cat#AB-0406) facilitates consistent PCR ® amplification (>90%) from Boil-prep template DNA. We routinely use Red Hot DNA polymerase to amplify PCR products up ® to 1.35kb from Boil-prep DNA following the procedure described below (modified from Yeadon and Catcheside 1996). Boil-prep method: A wet loop of 3 7 day old conidia is transferred to 100ìl of sterile Tris-EDTA (pH 8.0) in a 1.5ml microfuge tube and vortexed briefly. The conidial suspension is placed in a boiling waterbath for 10 minutes and then on ice for 5 minutes. The cellular debris is pelleted by centrifugation at 13000 RPM in a benchtop centrifuge for 5 minutes. 70ul of supernatant is transferred to a new 1.5ml microfuge tube and stored at -20oC. 2-5ul of Boil-prep DNA solution is used as a template in a 50ul PCR reaction with 0.5U of Red Hot DNA polymerase. Figure 1. Comparison of PCR amplicons from Boilprep genomic DNA (BP), genomic DNA (G) (prepared by the method of Irelan et al. 1993), plasmid DNA (P) and no DNA (-ve) templates. Size marker (M) = 100bp DNA ladder (New England Biolabs) with a 500bp reference band that contains 97ng of DNA. Each sample lane contains 6ul of a 50ul PCR reaction. DNA yields fro","PeriodicalId":12490,"journal":{"name":"Fungal Genetics Reports","volume":"111 1","pages":"24"},"PeriodicalIF":0.0,"publicationDate":"2005-12-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"80632617","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
I have been developing strains which I think will make it possible to produce "minimally-sheltered knockouts" of essential genes routinely. They could equally well be called "self-adjusting knockdowns" of essential genes. Such minimally-sheltered knockouts could give results in microarray analysis that would be less subject to artifact than results with heterokaryons or with homokaryons sheltered with an inducible wild-type allele of the gene in question. The method addresses two questions about each cloned gene, henceforth and collectively called "your favorite gene" (yfg). yfg should be one which does not pass successfully through a cross as a hygromycin-resistant knockout. (1) Is yfg a dispensable gene? (2) If yfg is found to be essential, what is the "terminal phenotype" of the yfgΔ knockout mutation? By "terminal phenotype," I mean that of the heterokaryon between yfgΔ and a partial diploid, yfg+/Δ, in which the partial diploid is the minimum proportion that allows somewhat suboptimal growth. Creative Commons License This work is licensed under a Creative Commons Attribution-Share Alike 4.0 License. This regular paper is available in Fungal Genetics Reports: http://newprairiepress.org/fgr/vol52/iss1/4
{"title":"Construction of minimally-sheltered knockouts mutants of Neurospora crassa","authors":"R. L. Metzenberg","doi":"10.4148/1941-4765.1123","DOIUrl":"https://doi.org/10.4148/1941-4765.1123","url":null,"abstract":"I have been developing strains which I think will make it possible to produce \"minimally-sheltered knockouts\" of essential genes routinely. They could equally well be called \"self-adjusting knockdowns\" of essential genes. Such minimally-sheltered knockouts could give results in microarray analysis that would be less subject to artifact than results with heterokaryons or with homokaryons sheltered with an inducible wild-type allele of the gene in question. The method addresses two questions about each cloned gene, henceforth and collectively called \"your favorite gene\" (yfg). yfg should be one which does not pass successfully through a cross as a hygromycin-resistant knockout. (1) Is yfg a dispensable gene? (2) If yfg is found to be essential, what is the \"terminal phenotype\" of the yfgΔ knockout mutation? By \"terminal phenotype,\" I mean that of the heterokaryon between yfgΔ and a partial diploid, yfg+/Δ, in which the partial diploid is the minimum proportion that allows somewhat suboptimal growth. Creative Commons License This work is licensed under a Creative Commons Attribution-Share Alike 4.0 License. This regular paper is available in Fungal Genetics Reports: http://newprairiepress.org/fgr/vol52/iss1/4","PeriodicalId":12490,"journal":{"name":"Fungal Genetics Reports","volume":"29 1","pages":"11-13"},"PeriodicalIF":0.0,"publicationDate":"2005-12-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"84711463","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
If an organism is already well known to the public before scientists adopt it for research, the popular, vernacular name usually continues to be used. Examples are yeast for Saccharomyces, mouse for Mus, corn for Zea mays, and silkworm for Bombyx. These common n ames are we ll established, and us ers are g enerally comfortable wi th t hem. H owever, some wi dely used r esearch organisms happen to have acquired popular names that are clearly inaccurate or misleading. Such has been the fate of Drosophila and Neurospora. Melvin Green (2002) has protested the use of "fruit fly" for Drosophila, pointing out that the name does not apply for the many Drosophila species that use substrates other than fruit, and that it leads to confusion with the Mediterranean fruit fly, a serious agricultural pest. He urges that the popular name fruit fly be abandoned and that the scientific name Drosophila be used in all scientific literature and in textbooks. As with the name fruit fly for Drosophila, objection can be made to using red bread mold for Neurospora. The vernacular name is imprecise and misleading in two respects, regarding both color and substrate. As to color, homothallic Neurospora species are devoid of conidia and of visible carotenoid pigments, while N. crassa and other conidiating species, which do display carotenoids, are orange or yellow-orange rather than red. When dark-grown cultures of a conidiating Neurospora species are first brought into the light, they are colorless. Then, within an hour, they become pigmented. The initial blush of color is pink or red, but this is quickly transformed to orange. The ephemeral red stage, which is rarely seen and is probably unknown to most observers, seems a poor choice for naming the organism. As to substrate, N eurospora exi sted for mi llions of years on nat ural substrates, in t he ab sence of human art ifacts. Calling Neurospora a bread mold might be considered an example of anthropocentric arrogance. Although Neurospora is able to grow profusely on bread, and was first recorded as a nuisance in bakeries (Payen 1843; see Perkins 1991), its occurrence is rare compared to other contaminating molds, especially since antifungal agents were introduced and bakery sanitation was improved. I have myself seen many examples of moldy bread that was black or green, but never one that was orange or red.
{"title":"Why \"Red bread mold\" is an inappropriate name for Neurospora","authors":"D. D. Perkins","doi":"10.4148/1941-4765.1121","DOIUrl":"https://doi.org/10.4148/1941-4765.1121","url":null,"abstract":"If an organism is already well known to the public before scientists adopt it for research, the popular, vernacular name usually continues to be used. Examples are yeast for Saccharomyces, mouse for Mus, corn for Zea mays, and silkworm for Bombyx. These common n ames are we ll established, and us ers are g enerally comfortable wi th t hem. H owever, some wi dely used r esearch organisms happen to have acquired popular names that are clearly inaccurate or misleading. Such has been the fate of Drosophila and Neurospora. Melvin Green (2002) has protested the use of \"fruit fly\" for Drosophila, pointing out that the name does not apply for the many Drosophila species that use substrates other than fruit, and that it leads to confusion with the Mediterranean fruit fly, a serious agricultural pest. He urges that the popular name fruit fly be abandoned and that the scientific name Drosophila be used in all scientific literature and in textbooks. As with the name fruit fly for Drosophila, objection can be made to using red bread mold for Neurospora. The vernacular name is imprecise and misleading in two respects, regarding both color and substrate. As to color, homothallic Neurospora species are devoid of conidia and of visible carotenoid pigments, while N. crassa and other conidiating species, which do display carotenoids, are orange or yellow-orange rather than red. When dark-grown cultures of a conidiating Neurospora species are first brought into the light, they are colorless. Then, within an hour, they become pigmented. The initial blush of color is pink or red, but this is quickly transformed to orange. The ephemeral red stage, which is rarely seen and is probably unknown to most observers, seems a poor choice for naming the organism. As to substrate, N eurospora exi sted for mi llions of years on nat ural substrates, in t he ab sence of human art ifacts. Calling Neurospora a bread mold might be considered an example of anthropocentric arrogance. Although Neurospora is able to grow profusely on bread, and was first recorded as a nuisance in bakeries (Payen 1843; see Perkins 1991), its occurrence is rare compared to other contaminating molds, especially since antifungal agents were introduced and bakery sanitation was improved. I have myself seen many examples of moldy bread that was black or green, but never one that was orange or red.","PeriodicalId":12490,"journal":{"name":"Fungal Genetics Reports","volume":"2 1","pages":"7-8"},"PeriodicalIF":0.0,"publicationDate":"2005-12-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"74388365","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
We compared promoters, coding sequences, introns and terminators of glyceraldehyde 3-phosphate dehydrogenase genes (gpd) from various basidiomycetes. Coding regions of these housekeeping genes are highly conserved (between 60 to 99% DNA identity) whilst non-coding regions have DNA identities of around 40%. Amongst all homobasidiomycete promoters, the TATA region and a CT-rich region with the potential transcription start sites are highest conserved. Surprisingly, there are no other conserved motifs common to all promoters. Up to five introns are clustered at the far 5 ́ ends of the genes, hinting to a potential function in efficient gene expression. Creative Commons License This work is licensed under a Creative Commons Attribution-Share Alike 4.0 License. This regular paper is available in Fungal Genetics Reports: http://newprairiepress.org/fgr/vol52/iss1/6 18 Fungal Genetics Newsletter Comparison of gpd genes and their protein products in basidiomycetes Sreedhar Kilaru and Ursula Kües Molecular Wood Biotechnology, Institute of Forest Botany, Georg-August-University Göttingen, 37077 Göttingen, Germany Fungal Genetics Newsletter 52:18-23 We compared promoters, coding sequences, introns and terminators of glyceraldehyde 3-phosphate dehydrogenase genes (gpd) from various basidiomycetes. Coding regions of these housekeeping genes are highly conserved (between 60 to 99% DNA identity) whilst non-coding regions have DNA identities of around 40%. Amongst all homobasidiomycete promoters, the TATA region and a CT-rich region with the potential transcription start sites are highest conserved. Surprisingly, there are no other conserved motifs common to all promoters. Up to five introns are clustered at the far 5 ́ ends of the genes, hinting to a potential function in efficient gene expression. Little is known about promoters in higher basidiomycetes. Constitutive promoter activities have been described for some homologous and heterologous promoters in Coprinopsis cinerea with the Agaricus bisporus gpdII (glyceraldehyde 3-phosphate dehydrogenase gene 2) promoter being highest in activity (Kilaru et al., 2005). Use of gpd promoters from A. bisporus, Flammulina velutipes, Lentinula edodes, Phanerochaete chrysosporium, Schizophyllum commune and Trametes versicolor has by now been made in different species either for laccase and peroxidase production or for expression of gfp (green fluorescent protein gene) or the bacterial hygromycin resistance gene hph (for references see Kilaru et al., 2005). Surprisingly, homology among these promoter sequences is relatively low (Kilaru et al., 2005). In contrast, the two known gpd genes from A. bisporus (termed gpdI and gpdII), the single gpd gene from P. chrysosporium and an isolated gpd gene from S. commune have been described as highly conserved in intron positions as well as in sequence of their products (Harmsen et al., 1992). Analysis of all gpd genes from basidiomycetes currently present in the NCBI database and of t
{"title":"Comparison of gpd genes and their protein products in basidiomycetes","authors":"Sreedhar Kilaru, U. Kües","doi":"10.4148/1941-4765.1125","DOIUrl":"https://doi.org/10.4148/1941-4765.1125","url":null,"abstract":"We compared promoters, coding sequences, introns and terminators of glyceraldehyde 3-phosphate dehydrogenase genes (gpd) from various basidiomycetes. Coding regions of these housekeeping genes are highly conserved (between 60 to 99% DNA identity) whilst non-coding regions have DNA identities of around 40%. Amongst all homobasidiomycete promoters, the TATA region and a CT-rich region with the potential transcription start sites are highest conserved. Surprisingly, there are no other conserved motifs common to all promoters. Up to five introns are clustered at the far 5 ́ ends of the genes, hinting to a potential function in efficient gene expression. Creative Commons License This work is licensed under a Creative Commons Attribution-Share Alike 4.0 License. This regular paper is available in Fungal Genetics Reports: http://newprairiepress.org/fgr/vol52/iss1/6 18 Fungal Genetics Newsletter Comparison of gpd genes and their protein products in basidiomycetes Sreedhar Kilaru and Ursula Kües Molecular Wood Biotechnology, Institute of Forest Botany, Georg-August-University Göttingen, 37077 Göttingen, Germany Fungal Genetics Newsletter 52:18-23 We compared promoters, coding sequences, introns and terminators of glyceraldehyde 3-phosphate dehydrogenase genes (gpd) from various basidiomycetes. Coding regions of these housekeeping genes are highly conserved (between 60 to 99% DNA identity) whilst non-coding regions have DNA identities of around 40%. Amongst all homobasidiomycete promoters, the TATA region and a CT-rich region with the potential transcription start sites are highest conserved. Surprisingly, there are no other conserved motifs common to all promoters. Up to five introns are clustered at the far 5 ́ ends of the genes, hinting to a potential function in efficient gene expression. Little is known about promoters in higher basidiomycetes. Constitutive promoter activities have been described for some homologous and heterologous promoters in Coprinopsis cinerea with the Agaricus bisporus gpdII (glyceraldehyde 3-phosphate dehydrogenase gene 2) promoter being highest in activity (Kilaru et al., 2005). Use of gpd promoters from A. bisporus, Flammulina velutipes, Lentinula edodes, Phanerochaete chrysosporium, Schizophyllum commune and Trametes versicolor has by now been made in different species either for laccase and peroxidase production or for expression of gfp (green fluorescent protein gene) or the bacterial hygromycin resistance gene hph (for references see Kilaru et al., 2005). Surprisingly, homology among these promoter sequences is relatively low (Kilaru et al., 2005). In contrast, the two known gpd genes from A. bisporus (termed gpdI and gpdII), the single gpd gene from P. chrysosporium and an isolated gpd gene from S. commune have been described as highly conserved in intron positions as well as in sequence of their products (Harmsen et al., 1992). Analysis of all gpd genes from basidiomycetes currently present in the NCBI database and of t","PeriodicalId":12490,"journal":{"name":"Fungal Genetics Reports","volume":"6 1","pages":"18-23"},"PeriodicalIF":0.0,"publicationDate":"2005-12-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"74385843","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
We report the expression of Discosoma red fluorescent protein (RFP) and RFP fusion proteins in Neurospora crassa. RFP was expressed under the control of the Neurospora ccg-1 promoter in transformants with single copies integrated at the his-3 locus by gene targeting. Because this RFP gene, tdimer2(12), contains a 677 bp direct tandem repeat of dsRed, RFP constructs underwent RIP at high frequency in rid strains. Fusion proteins of RFP to the amino terminus of Neurospora heterochromatin protein 1 (HP1) +
{"title":"Expression and Visualization of Red Fluorescent Protein (RFP) in Neurospora crassa","authors":"M. Freitag, E. Selker","doi":"10.4148/1941-4765.1124","DOIUrl":"https://doi.org/10.4148/1941-4765.1124","url":null,"abstract":"We report the expression of Discosoma red fluorescent protein (RFP) and RFP fusion proteins in Neurospora crassa. RFP was expressed under the control of the Neurospora ccg-1 promoter in transformants with single copies integrated at the his-3 locus by gene targeting. Because this RFP gene, tdimer2(12), contains a 677 bp direct tandem repeat of dsRed, RFP constructs underwent RIP at high frequency in rid strains. Fusion proteins of RFP to the amino terminus of Neurospora heterochromatin protein 1 (HP1) +","PeriodicalId":12490,"journal":{"name":"Fungal Genetics Reports","volume":"25 1","pages":"14-17"},"PeriodicalIF":0.0,"publicationDate":"2005-12-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"76813655","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
{"title":"Using Neurospora to demonstrate the unidirectional nature of fungal mating.","authors":"K. McCluskey","doi":"10.4148/1941-4765.1122","DOIUrl":"https://doi.org/10.4148/1941-4765.1122","url":null,"abstract":"","PeriodicalId":12490,"journal":{"name":"Fungal Genetics Reports","volume":"76 1","pages":"9-10"},"PeriodicalIF":0.0,"publicationDate":"2005-12-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"83367969","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}