Labeling of biocontrol agents ZJY-1 and ZJY-116 gfp gene and its ecological adaptability in cucumber rhizosphere

Abstract

Abstract: The recombined plasmid pRP22-GFP contained with gfp gene and chloramphenicol resistant was successfully introduced into two biocontrol agents Brevibacillus brevis ZJY-1 and Bacillus subtilis ZJY-116.After seed inoculation,the survival and colonization of the two strains were studied by periodically retrieving the GFP-tagged strains in the cucumber rhizosphere based on the selective markers.The results showed that both the strains could successfully colonize in the rhizosphere during the whole life of cucumber,and a higher colonization level was observed during anthesis and fruition stages.In pot trials,they could migrate to the nearby non-inoculated spontaneous weed plants,and reestablish in the rhizosphere of plants subsequently grown in the same pot.

Key words: Plasmid pRP22-GFP, Gfp gene, Brevibacillus brevis, Bacillus subtili, Cucumber rhizosphere, Colonization

CLC Number:

S336

ZHANG Xin, ZHANG Bingxin, YU Jingquan, ZHANG Zhen, SHEN Weifeng, CHEN Zhenyu, SHI Jiang . Labeling of biocontrol agents ZJY-1 and ZJY-116 gfp gene and its ecological adaptability in cucumber rhizosphere[J]. Chinese Journal of Applied Ecology, 2005, 16(11): 2144-2148.

References

[1] Bonkowski M, Cheng W, Griffiths BS, et al. 2000. Microbial-faunal interactions in the rhizosphere and effects on plant growth. Eur J Soil Biol, 36:135～ 147
[2] Bloemberg GV, Lugtenberg BJ. 2001. Molecular basis of plant growth promotion and biocontrol by rhizobacteria. Curr Opin Plant Biol, 4:343 ～ 350
[3] Bennett AJ, Leifert C, Whipps JM. 2003. Survival of the biological agents Coniothyriurn minitans and Bacillus subtilis MBI600 introduced into pasteurized, sterilized and non-sterile soils. Soil Biol Biochem, 35:1565 ～ 1573
[4] Bloemberg GV, O' toole GA, Lugtenberg BJJ, et al. 1997. Green fluorescent protein as a marker for Pseudomonas spp. Appl Environ Microbiol, 63:4543～4551
[5] Chao WL, Nelson EB, Harman GE, et al. 1986. Colonization of the rhizosphere by biological control agents applied to seeds. Phytopathology, 76:60 ～ 65
[6] Chen X-B(陈晓斌),Zhang B-X(张炳欣),Lou B-G(楼兵干),et al.1999.Effect of plant growth-promoting rhizobacteria on disease control of cucumber seedlings.J Zhejiang Univ Sci(Agric Life Sci)(浙江大学学报·农业与生命科学版),25(6):578～582(in Chinese)
[7] Chen X-B(陈晓斌),Zhang B-X(张炳欣),Lou B-G(楼兵干),et al.2001.Introduction of the chromogenic gene to the plant Growth-oromoting rhizobacteria of cucumber.Acta Microbiol Sin (微生物学报),41(3):287～292(in Chinese)
[8] Dong Y-M(董越梅),An Q-L(安千里),Li J-D(李久蒂),et al.2000.Use of GFP and antibiotic resistance as selective markers to monitor colonization of associated nitrogen-fixing bacteria in maize rhizosphere.Chin J Appl Ecol(应用生态学报),6(1):61～65(in Chinese)
[9] Guo X-H(郭兴华),Jia S-F(贾士芳),Chen N-Y(陈乃用).1982.Bacillus protoplasts as recipients for plasmid DNA transfonnation.Acta Microbiol Sin(微生物学报),22(3):263～268(in Chinese)
[10] He L-Y(何琳燕),Huang W-Y(黄为一).2004.Introduction of bioluminescence genes into silicate-dissolving bacteria strain NBT.Chin J Appl Ecol(应用生态学报),15(7):1241～1244(in Chinese)
[11] Jetiyanon K, Kloepper JW. 2002. Mixtures of plant growth-promoting rhizobacteria for induction of systemic resistance against multiple plant diseases. Biol Control, 24:285～291
[12] Leff LG, Leff AA. 1996. Use of green fluorescent protein to monitor survival of genetically engineered bacteria in aquatic environments. Appl Environ Microbiol,62:3486 ～ 3488
[13] Lü Z-X(吕泽勋),Li J-D(李久蒂),Zhu Z-Q(朱至清).2001.Klebsiella oxytoca SG-11 labeling by green fluorescent protein gene (gfp)and its colonization in rice seeding roots.J Agric Biotechnol (农业生物技术学报),9(1):13～18(in Chinese)
[14] Luo Q-S(罗启仕),Wang H(王慧),Zhang X-H(张锡辉),et al.2004.Using of gfp geneto monitor environmental microorganisms.Chin J Water Supply Sewerage(中国给水排水),20(6):32～34(in Chinese)
[15] Li Y-G(李友国),Zhou J-C(周俊初).2003.Root colonization and nodulation of Sinorhizobium fredii HN01DL in Glycine max rhizosphere.Chin J Appl Ecol(应用生态学报),14(8):1283～1286(in Chinese)
[16] Lin F-C(林福呈),Zhang B-X(张炳欣),Ge Q-X(葛起新).1990.Effects of antagonistic substances produced by three isolates of Bacillus subtilis on conidia of Fusarium oxysporum f.sp niveum.J Zhejiang Agric Univ Sci(浙江农业大学学报),16(2):235～240 (in Chinese)
[17] Nautiyal CS, Johri JK, Singh HB. 2002. Survival of the rhizospherecompetent biocontrol strains Pseudomonas fluorescences NBR12650in the soil and phytosphere. Can Microbiol, 48(7):588 ～ 601
[18] Ramos HJO, Roncato-Maccari LDB, Souza EM, et al. 2002. Monitoring Azospirillum-wheat interaction using the gfp and gusA genes constitutively expressed from a new broad-host range vector.J Biotechnol, 97:243 ～252
[19] Ramamoorthy V, Viswanathan R, Raguchander T, et al.2001. Induction of systemic resistance by plant growth promoting rhizobacteria in crop plants against pests and diseases. Crop Prot, 20:1～11
[20] Sheng X-F(盛下放).2003.Colonization of silicate bacterium strain NBT in wheat roots.Chin J Appl Ecol(应用生态学报),14(11):1914～1946 (in Chinese)
[21] Scott KP, Mercer DK, Glover LA, et al. 1998. The green fluorescent protein as a visible marker for lactic acid bacteria in complex ecosystem. FEMS Microbiol Ecol, 26:219～230
[22] Waid JS. 1999. Does soil biodiversity depend upon metabiotic activity and influences. Appl Soil Ecol, 13:151 ～ 158
[23] Wiehe W, Hoflich G. 1995. Survival of plant growth rhizosphere bacteria in the rhizosphere of different crops and migration to noninoculated plants under field conditions in north-east Germany. Microbiol Res, 150:201 ～206
[24] Wiehe W, Hoflich G. 1995. Establishment of plant growth promoting bacteria in the rhizosphere of subsequent plants after harvest of the inoculated precrops. Microbiol Res, 150:331 ～ 335
[25] Winstanley C, Morgan JAW, Pickup RW, et al. 1991. Use of a xylE marker gene to monitor survival of recombinant Pseudomonas putida population in lake water by culture on nonselective media.Appl Environ Microbiol,57(7):1905～1913
[26] Wang G-H(王光华), Raaijmakers JM. 2004. Antibiotics production by bacterial agents and its role in biological control. Chin J Appl Ecol(应用生态学报),15(6):1100～1104(in Chinese)
[27] Wu L-J(吴连举),Yang Y-J(杨依军),Wu X(武侠),et al.1999. Studies on the control of Cylindrocarpon destructans of Ginseng by soil antagonistic microbes. Chin J Biol Contr (中国生物防治), 15(4):166 ～ 168(in Chinese)
[28] Xi CW, Lambrecht M, Vanderleyden J, et al. 1999. Bi-functional gfp-and gusA-containing mini-Tn5 transposon derivatives for combined gene expression and bacterial localization studies. J Microbiol Meth, 35:85 ～ 92
[29] Zhang S, Reddy MS, Kloepper JW. 2002. Development of assay for assessing induced systemic resistance by plant growth-promoting Rhizobacteria against blue mold of tobacco. Biol Contr, 23:79 ～86