Two cellulase genes, Cel15 and Cel73, were amplified from Bacillus subtilis genome DNA in a previous study. Two integrative vectors, pLEM4153 and pLEM4154, containing the genes Cel15 and Cel73, respectively, were constructed and successfully electroporated into the wild-type Lactobacillus reuteri which was isolated from chick guts through an optimized procedure. Two recombinant L. reuteri were selected from a Man, Rogosa, and Sharp (MRS) plate with 10 µg/ml erythromycin, and named L. reuteri XNY-Cel15 and L. reuteri XNY-Cel73, respectively. To verify the transcription and expression of the two cellulase genes in the recombinant L. reuteri strains, the mRNA relative quantity (RQ) and the cellulase activity were determined. The mRNA RQ of Cel15 in L. reuteri XNY-Cel15 is 1,8849.5, and that of Cel73 in L. reuteri XNY-Cel73 is 1,388, and the cellulase activity of the modified MRS broth cultured with L. reuteri XNY-Cel15 was 0.158 U/ml, whereas that with L. reuteri XNY-Cel73 was 0.15 U/ml.

Bayer EA, Chanzy H, Lamed R, Shoham Y: Cellulose, cellulases and cellulosomes. Curr Opin Struct Biol 1998;8:548–557.
Chaurasia AK, Parasnis A, Apte SK: An integrative expression vector for strain improvement and environmental applications of the nitrogen fixing cyanobacterium, Anabaena sp. strain pcc7120. J Microbiol Methods 2008;73:133–141.
Ferenczi S, Ganyu A, Blaha B, Semsey S, Nagy T, Csiszovszki Z, Orosz L, Papp PP: Integrative plasmid vector for constructing single-copy reporter systems to study gene regulation in Rhizobium meliloti and related species. Plasmid 2004;52:57–62.
Graminha EBN, Goncalves AZL, Pirota RDPB, Balsalobre MAA, Da Silva R, Gomes E: Enzyme production by solid-state fermentation: application to animal nutrition. Anim Feed Sci Technol 2008;144:1–22.
Heng NC, Jenkinson HF, Tannock GW: Cloning and expression of an endo-1,3–1,4-β-glucanase gene from Bacillus macerans in Lactobacillus reuteri. Appl Environ Microbiol 1997;63:3336–3340.
Hernandez M, Esteve T, Prat S, Pla M: Development of real-time PCR systems based on SYBR green I, AmplifluorTM and TaqMan technologies for specific quantitative detection of the transgenic maize event GA21. J Cereal Sci 2004;39:99–107.
Hormann S, Vogel RF, Ehrmann M: Construction of a new reporter system to study the NaCl-dependent dnaK promoter activity of Lactobacillus sanfranciscensis. Appl Microbiol Biotechnol 2006;70:690–697.
Hubalek Z: Protectants used in the cryopreservation of microorganisms. Cryobiology 2003; 46:205–229.
Inoue T, Moriya S, Ohkuma M, Kudo T: Molecular cloning and characterization of a cellulase gene from a symbiotic protist of the lower termite, Coptotermes formosanus. Gene 2005;349:67–75.
Lee Y-J, Kim B-K, Lee B-H, Jo K-I, Lee N-K, Chung C-H, Lee Y-C, Lee J-W: Purification and characterization of cellulase produced by Bacillus amyoliquefaciens DL-3 utilizing rice hull. Bioresour Technol 2008;99:378–386.
Li W, Huan X, Zhou Y, Ma Q, Chen Y: Simultaneous cloning and expression of two cellulase genes from Bacillus subtilis newly isolated from golden takin (Budorcas taxicolor bedfordi). Biochem Biophys Res Commun 2009;383:397–400.
Li W, Zhang W-W, Yang M-M, Chen Y-L: Cloning of the thermostable cellulase gene from newly isolated Bacillus subtilis and its expression in Escherichia coli. Mol Biotechnol 2008;40:195–201.
Liu JR, Yu B, Lin SH, Cheng KJ, Chen YC: Direct cloning of a xylanase gene from the mixed genomic DNA of rumen fungi and its expression in intestinal Lactobacillus reuteri. FEMS Microbiol Lett 2005a;251:233–241.
Liu JR, Yu B, Liu FH, Cheng KJ, Zhao X: Expression of rumen microbial fibrolytic enzyme genes in probiotic Lactobacillus reuteri. Appl Environ Microbiol 2005b;71:6769–6775.
Liu JR, Yu B, Zhao X, Cheng KJ: Coexpression of rumen microbial β-glucanase and xylanase genes in Lactobacillus reuteri. Appl Microbiol Biotechnol 2007;77:117–124.
Mason CK, Collins MA, Thompson K: Modified electroporation protocol for lactobacilli isolated from the chicken crop facilitates transformation and the use of a genetic tool. J Microbiol Methods 2005;60:353–363.
Miller GL: Use of dinitrosalicylic as reagent for the determination of reducing sugars. Anal Chem 1959;31:426–428.
Morita H, Toh H, Fukuda S, Horikawa H, Oshima K, Suzuki T, Murakami M, Hisamatsu S, Kato Y, Takizawa T, Fukuoka H, Yoshimura T, Itoh K, O’Sullivan DJ, McKay LL, Ohno H, Kikuchi J, Masaoka T, Hattori M: Comparative genome analysis of Lactobacillus reuteri and Lactobacillus fermentum reveal a genomic island for reuterin and cobalamin production. DNA Res 2008;15:151–161.
Naumenko VS, Osipova DV, Kostina EV, Kulikov AV: Utilization of a two-standard system in real-time PCR for quantification of gene expression in the brain. J Neurosci Methods 2008;170:197–203.
O’Connor-Robison CI, Nielsen BD, Morris R: Cellulase supplementation does not improve the digestibility of a high-forage diet in horses. J Equine Vet Sci 2007;27:535–538.
Rizzatti AC, Jorge JA, Terenzi HF, Rechia CG, Polizeli ML: Purification and properties of a thermostable extracellular β-d-xylosidase produced by a thermotolerant Aspergillus phoenicis. J Ind Microbiol Biotechnol 2001;26:156–160.
Sambrook J, Fritsch EF, Maniatis T: Molecular Cloning: A Laboratory Manual, ed 2. New York, Cold Spring Harbor Laboratory Press, 1989.
Schwab C, Walter J, Tannock GW, Vogel RF, Ganzle MG: Sucrose utilization and impact of sucrose on glycosyltransferase expression in Lactobacillus reuteri. Syst Appl Microbiol 2007;30:433–443.
Serror P, Sasaki T, Ehrlich SD, Maguin E: Electrotransformation of Lactobacillus delbrueckii subsp. bulgaricus and l. Delbrueckii subsp. lactis with various plasmids. Appl Environ Microbiol 2002;68:46–52.
Teresa Alegre M, Carmen Rodriguez M, Mesas JM: Transformation of Lactobacillus plantarum by electroporation with in vitro modified plasmid DNA. FEMS Microbiol Lett 2004;241:73–77.
Yang C-Y, Ho Y-C, Pang J-C, Huang S-S, Tschen JS-M: Cloning and expression of an antifungal chitinase gene of a novel Bacillus subtilis isolate from Taiwan potato field. Bioresour Technol 2009;100:1454–1458.
Zhao F-Y, Lin J-F, Zeng X-L, Guo L-Q, Wang Y-H, You L-R: Improvement in fruiting body yield by introduction of the Ampullaria crossean multi-functional cellulase gene into Volvariella volvacea. Bioresour Technol 2010; 101:6482–6486.
You do not currently have access to this content.