香菇菌丝体对四种抗生素的敏感性
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摘要
为探究4种常用抗生素对香菇菌丝体生长的影响,筛选适用于香菇遗传转化的抗生素。本研究以香菇[Lentinula edodes(Berk.)Pegler]沪香F2双核菌丝以及其原生质体单核菌丝为材料,通过测试两者在含有不同种类、不同浓度下抗生素培养皿中的生长速率以及抑制率,探究香菇单双核菌丝对氨苄青霉素、卡那霉素、头孢霉素、潮霉素4种抗生素的敏感性。结果表明:氨苄青霉素、卡那霉素、头孢霉素对香菇单双核菌丝生长以及菌落形态基本没有影响,潮霉素对香菇单双核菌丝生长有强烈的抑制作用,12mg·L-1的潮霉素能够完全抑制香菇双核菌丝的生长,14mg·L-1的潮霉素能够完全抑制香菇单核菌丝的生长。卡那霉素不能作为香菇遗传转化的筛选抗生素使用;潮霉素可以用作香菇遗传转化的筛选抗生素;氨苄青霉素和头孢霉素均可以用于对农杆菌的抑制。
In order to investigate the effects of four antibiotics on the mycelium growth of lentinus edodes,screen out suitable antibiotics for genetic transformation of lentinus edodes,we used the mycelium of Lentinula edodes(Berk.)Pegler-huxiang F2 as material,growth rates and inhibition rates were measured in petri dishes containing different types and concentration,the research aimed to investigate that resistance of mycelia of Lentinula edodes against four kind antibiotics,including ampicillin,kanamycin,cefotaxime and hygromycin.The results showed that ampicillin,kanamycin,cefotaximehad no effect on mycelia growth and colony morphology of Lentinula edodes,hygromycin had a strong inhibitory effect on the growth of mushroom mycelium,hygromycin at 12 mg·L-1 could completely inhibit the growth of Lentinula edodes dikaryon mycelia,hygromycin at14 mg·L-1 could completely inhibit the growth of Lentinula edodes monokaryon mycelia.Kanamycin cannot be used as a screening antibiotic for genetic transformation of Lentinula edodes.Hydramycin can be used as a screening antibiotic for genetic transformation of Lentinula edodes.Ampicillin and cefotaxime could be used to inhibit Agrobacterium.
In order to investigate the effects of four antibiotics on the mycelium growth of lentinus edodes,screen out suitable antibiotics for genetic transformation of lentinus edodes,we used the mycelium of Lentinula edodes(Berk.)Pegler-huxiang F2 as material,growth rates and inhibition rates were measured in petri dishes containing different types and concentration,the research aimed to investigate that resistance of mycelia of Lentinula edodes against four kind antibiotics,including ampicillin,kanamycin,cefotaxime and hygromycin.The results showed that ampicillin,kanamycin,cefotaximehad no effect on mycelia growth and colony morphology of Lentinula edodes,hygromycin had a strong inhibitory effect on the growth of mushroom mycelium,hygromycin at 12 mg·L-1 could completely inhibit the growth of Lentinula edodes dikaryon mycelia,hygromycin at14 mg·L-1 could completely inhibit the growth of Lentinula edodes monokaryon mycelia.Kanamycin cannot be used as a screening antibiotic for genetic transformation of Lentinula edodes.Hydramycin can be used as a screening antibiotic for genetic transformation of Lentinula edodes.Ampicillin and cefotaxime could be used to inhibit Agrobacterium.
引文
[1]聂林林.香菇热泵除湿干燥技术的研究[D].郑州:河南工业大学,2015.
[2]刘春如.香菇的分布概况及生物学特性[J].中国林副特产,2001(4):32-33.
[3]Chen L,Gong Y,Cai Y,et al.Genome sequence of the edible cultivated mushroom Lentinula edodes(Shiitake)reveals insights into lignocellulose degradation[J].Plos One,2016,11(8).
[4]Shim D,Park S G,Kim K,et al.Whole genome de novo,sequencing and genome annotation of the world popular cultivated edible mushroom,Lentinula edodes[J].Journal of Biotechnology,2016,223:24-25.
[5]胡乐琴,潘迎捷.食用菌分子转化研究状况[J].中国食用菌,2006,25(4):7-10.
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[7]盛立柱,宋冰,戴月婷,等.农杆菌介导的刺芹侧耳遗传转化体系的建立[J].菌物学报,2015,34(4):653-661.
[8]康林芝,林俊芳,黄秀琴,等.紫杉烯合酶基因遗传转化金针菇的研究[J].食品工业科技,2013,34(2):190-193.
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[11]Yin Y,Liu Y,Jin H,et al.Polyethylene glycol-mediated transformation of fused egfp-hph gene under the control of gpd promoter in Pleurotus eryngii[J].Biotechnology Letters,2012,34(10):1895-1900.
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[14]徐茂军.转基因植物中卡那霉素抗性(KANR)标记基因的生物安全性[J].生物学通报,2001,36(2):18-19.
[15]王紫萱,易自力.卡那霉素在植物转基因中的应用及其抗性基因的生物安全性评价[J].中国生物工程杂志,2003,23(6):9-13.
[16]杨桥,蔺自敏,侯详文,等.小麦遗传转化中潮霉素筛选体系的建立及应用[J].麦类作物学报,2014,34(3):304-310.
[17]张学炜,王笑梅,李明春,等.以潮霉素B抗性为选择标记的深黄被孢霉原生质体转化[J].生物工程学报,2007,23(3):462-466.
[18]Michielse C B,Hooykaas P J,Ca V D H,et al.Agrobacterium-mediated transformation of the filamentous fungus Aspergillus awamori[J].Nature Protocols,2008,3(10):1671-8.
[19]Hanhineva K J,Krenlampi S O.Production of transgenic strawberries by temporary immersion bioreactor system and verification by TAIL-PCR[J].Bmc Biotechnology,2007,7(1):11.
[20]范春节.尾赤桉再生体系及农杆菌介导的转基因体系研究[D].北京:中国林业科学研究院,2008.
[21]Lin J J,Assad-garcia N,Kuo J.Plant hormone effect of antibiotics on the transformation efficiency of plant tissues by Agrobacterium tumefaciens,cells[J].Plant Science,1995,109(2):171-177.
[22]Yepes L M,Aldwinekle H S.Factors that effect leaf regeneration efficiency in apple,and effect of antibiotics in morphogenesis[J].Plant Cell Tissue&Organ Culture,1994,37(3):257-269.
[2]刘春如.香菇的分布概况及生物学特性[J].中国林副特产,2001(4):32-33.
[3]Chen L,Gong Y,Cai Y,et al.Genome sequence of the edible cultivated mushroom Lentinula edodes(Shiitake)reveals insights into lignocellulose degradation[J].Plos One,2016,11(8).
[4]Shim D,Park S G,Kim K,et al.Whole genome de novo,sequencing and genome annotation of the world popular cultivated edible mushroom,Lentinula edodes[J].Journal of Biotechnology,2016,223:24-25.
[5]胡乐琴,潘迎捷.食用菌分子转化研究状况[J].中国食用菌,2006,25(4):7-10.
[6]黄小丹,柳建良.食用菌分子生物学研究进展[J].仲恺农业工程学院学报,2009,22(3):53-58.
[7]盛立柱,宋冰,戴月婷,等.农杆菌介导的刺芹侧耳遗传转化体系的建立[J].菌物学报,2015,34(4):653-661.
[8]康林芝,林俊芳,黄秀琴,等.紫杉烯合酶基因遗传转化金针菇的研究[J].食品工业科技,2013,34(2):190-193.
[9]姜宁,宋春艳,刘建雨,等.农杆菌介导香菇菌丝遗传转化体系的研究[J].菌物学报,2017,36(11):1514-1523.
[10]Rhee M D V D,Gra9a P M A,Huizing H J,et al.Transformation of the cultivated mushroom,Agaricus bisporus,to hygromycin B resistance[J].Molecular&General Genetics Mgg,1996,250(3):252-258.
[11]Yin Y,Liu Y,Jin H,et al.Polyethylene glycol-mediated transformation of fused egfp-hph gene under the control of gpd promoter in Pleurotus eryngii[J].Biotechnology Letters,2012,34(10):1895-1900.
[12]Sunagawa M,Magae Y.Transformation of the edible mushroom Pleurotus ostreatus by particle bombardment[J].Fems Microbiology Letters,2002,211(2):143-146.
[13]Cummings W J,Celerin M,Crodian J,et al.Insertional mutagenesis in Coprinus cinereus:Use of a dominant selectable marker to generate tagged,sporulation-defective mutants[J].Current Genetics,1999,36(6):371-382.
[14]徐茂军.转基因植物中卡那霉素抗性(KANR)标记基因的生物安全性[J].生物学通报,2001,36(2):18-19.
[15]王紫萱,易自力.卡那霉素在植物转基因中的应用及其抗性基因的生物安全性评价[J].中国生物工程杂志,2003,23(6):9-13.
[16]杨桥,蔺自敏,侯详文,等.小麦遗传转化中潮霉素筛选体系的建立及应用[J].麦类作物学报,2014,34(3):304-310.
[17]张学炜,王笑梅,李明春,等.以潮霉素B抗性为选择标记的深黄被孢霉原生质体转化[J].生物工程学报,2007,23(3):462-466.
[18]Michielse C B,Hooykaas P J,Ca V D H,et al.Agrobacterium-mediated transformation of the filamentous fungus Aspergillus awamori[J].Nature Protocols,2008,3(10):1671-8.
[19]Hanhineva K J,Krenlampi S O.Production of transgenic strawberries by temporary immersion bioreactor system and verification by TAIL-PCR[J].Bmc Biotechnology,2007,7(1):11.
[20]范春节.尾赤桉再生体系及农杆菌介导的转基因体系研究[D].北京:中国林业科学研究院,2008.
[21]Lin J J,Assad-garcia N,Kuo J.Plant hormone effect of antibiotics on the transformation efficiency of plant tissues by Agrobacterium tumefaciens,cells[J].Plant Science,1995,109(2):171-177.
[22]Yepes L M,Aldwinekle H S.Factors that effect leaf regeneration efficiency in apple,and effect of antibiotics in morphogenesis[J].Plant Cell Tissue&Organ Culture,1994,37(3):257-269.