Detailed information of the strains used in this study is provided in Supplementary Data 4 . All anaerobes were cultured in an anaerobic chamber with an atmosphere of 83% N2, 2% H2 and 15% CO2 at 37 °C. For most experiments, Bacteroides strains, AC, CC were grown in ABB media, ER was grown in ABB media with the addition of 3.3 mM sodium acetate (Sigma) and RI was grown in Brain Heart Infusion Broth (BHI, Sigma). We used E. coli pir2 (Invitrogen) for cloning and maintenance of plasmids with R6K origin (pNBU2-ermGb derivatives and pFW1000 derivatives, Supplementary Data 4 ). E. coli DH5α (Thermo Fisher Scientific) was used for cloning and maintenance of plasmids with p15A, pSC101ts and ColE1 origins (pFW2000 derivatives, pFW3000 and pFW4000). We used E. coli BW29427 (E. coli Genetic Stock Center, CGSC) for E. coli-Bacteroides conjugations. All E. coli strains were grown aerobically in Luria Bertoni (LB, Sigma) media. To support the growth of E. coli BW29427, we supplemented LB media with 25 μM of 2,6-Diaminopimelic acid (DAP, Sigma). We used the following antibiotics when required including 100 μg mL−1 carbenicillin (Carb, IBI Scientific), 25 μg mL−1 erythromycin (Erm, Sigma) and 200 μg mL−1 gentamicin (Gm, Sigma). We used 1 mM of Isopropyl β-D-1-thiogalactopyranoside (IPTG, Gold Biotechnology) for the induction of FnCpf1.
Carbenicillin carb
Manufactured by IBI Scientific
Carbenicillin is a semi-synthetic penicillin antibiotic. Its core function is to inhibit the synthesis of the bacterial cell wall, thereby preventing the growth and reproduction of certain bacteria.
Automatically generated - may contain errors
Lab products found in correlation
E coli dh5α, by Thermo Fisher Scientific (2 mentions)
E coli pir2, by Thermo Fisher Scientific (2 mentions)
Sodium acetate, by Merck Group (2 mentions)
Isopropyl β d 1 thiogalactopyranoside iptg, by GoldBio (1 mentions)
Erythromycin erm, by Merck Group (1 mentions)
Gentamicin gm, by Merck Group (1 mentions)
Erythromycin em, by Merck Group (1 mentions)
2 protocols using carbenicillin carb
1
Anaerobic Bacteroides and E. coli Culturing
2
Anaerobic Culturing and Genetic Manipulation
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Detailed information of the strains used in this study is provided in Table S1. All anaerobes were cultured in an anaerobic chamber with an atmosphere of 83% N2, 2% H2 and 15% CO2 at 37 ºC.
For most experiments, Bacteroides strains, AC, CC were grown in Anaerobe Basal Broth (ABB, Oxoid), ER was grown in ABB media with the addition of 3.3 mM sodium acetate (Sigma) and RI was grown in Brain Heart Infusion Broth (BHI, Sigma). We used E. coli pir2 (Invitrogen) for cloning and maintenance of plasmids with R6K origin (pNBU2-ermGb derivatives and pFW1000 derivatives, Table S2). E. coli DH5α (Thermo Fisher Scientific) was used for cloning and maintenance of plasmids with p15A, pSC101ts and ColE1 origins (pFW2000 derivatives, pFW3000 and pFW4000). We used E. coli BW29427 (E. coli Genetic Stock Center, CGSC) for E. coli-Bacteroides conjugations. All E. coli strains were grown aerobically in Luria Bertoni (LB) media. To support the growth of E. coli BW29427, we supplemented LB media with 25 nM of 2,6-Diaminopimelic acid (DAP, Sigma). We used the following antibiotics when required including 100 µg mL -1 carbenicillin (Carb, IBI Scientific), 25 µg mL -1 erythromycin (Em, Sigma) and 200 µg mL -1 gentamicin (Gm, Sigma). We used 1 mM of Isopropyl β-D-1-thiogalactopyranoside (IPTG, Gold Biotechnology) for the induction of FnCpf1.
For most experiments, Bacteroides strains, AC, CC were grown in Anaerobe Basal Broth (ABB, Oxoid), ER was grown in ABB media with the addition of 3.3 mM sodium acetate (Sigma) and RI was grown in Brain Heart Infusion Broth (BHI, Sigma). We used E. coli pir2 (Invitrogen) for cloning and maintenance of plasmids with R6K origin (pNBU2-ermGb derivatives and pFW1000 derivatives, Table S2). E. coli DH5α (Thermo Fisher Scientific) was used for cloning and maintenance of plasmids with p15A, pSC101ts and ColE1 origins (pFW2000 derivatives, pFW3000 and pFW4000). We used E. coli BW29427 (E. coli Genetic Stock Center, CGSC) for E. coli-Bacteroides conjugations. All E. coli strains were grown aerobically in Luria Bertoni (LB) media. To support the growth of E. coli BW29427, we supplemented LB media with 25 nM of 2,6-Diaminopimelic acid (DAP, Sigma). We used the following antibiotics when required including 100 µg mL -1 carbenicillin (Carb, IBI Scientific), 25 µg mL -1 erythromycin (Em, Sigma) and 200 µg mL -1 gentamicin (Gm, Sigma). We used 1 mM of Isopropyl β-D-1-thiogalactopyranoside (IPTG, Gold Biotechnology) for the induction of FnCpf1.
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