TABLE 2.
Plasmids and strains used in this study
| Plasmid or strain | Description and relevant characteristicse | Source or reference |
|---|---|---|
| Plasmids | ||
| pUC19 | General purpose cloning vector, Ap | NEBa |
| pMU102 | pMU104, region between the FokI and EcoRI sites has been deleted, Cm | This study |
| pMU104 | pNW33N, E. coli-C. thermocellum shuttle vector, Cm | BGSCb |
| pMU110 | pMQ87, S. cerevisiae-E. coli shuttle vector, Ura+, Gm | This study |
| pMU111 | pMU110 with aac1 (Gm) replaced by cat from pMU104, Ura+, Cm | This study |
| pMU113 | pMU111 with C. thermocellum gapDHp driving cat, Ura+, Cm | This study |
| pMU245 | E. coli-C. thermocellum cloning vector, Ap | This study |
| pMU357 | S. cerevisiae-E. coli-C. thermocellum shuttle vector for expressing genes in C. thermocellum, C. thermocellum gapDHp, Ura+, Cm | This study |
| pMU440 | S. cerevisiae-E. coli-C. thermocellum shuttle vector, C. thermocellum ΔpyrF cassette, Ura+, Cm | This study |
| pMU482 | E. coli-C. thermocellum shuttle vector, C. thermocellum ΔpyrF cassette, Cm | This study |
| pMU582 | pMU110, C. thermocellum cbp promoter, cbp gene, T1T2 terminator, Ura+, Gm | This study |
| pMU597 | pMU582, C. thermocellum cbp gene replaced by C. thermocellum pyrF gene, creating cbpp-pyrF cassette, Ura+, Gm | This study |
| pMU612 | pMU102 containing cbpp-pyrF cassette, Cm | This study |
| pMU749 | pMU245, CEN6/ARSH4, ura3, S. cerevisiae-E. coli-C. thermocellum vector, Ura+, Ap | This study |
| pMU769 | pMU749 with ΔpyrF::gapDHp-cat cassette, Ura+, Ap, Cm | This study |
| pMU1016 | pMU749 with Δpta::gapDHp-cat cassette, Ura+, Ap, Cm | This study |
| pMU1162 | pMU1016 with cbpp-pyrF cassette, Ura+, Ap, Cm | This study |
| Strains | ||
| E. coli Top10 | Cloning strain | Invitrogenc |
| S. cerevisiae InvSC1 | Ura3− for gap repair cloning | Invitrogenc |
| C. thermocellum M0003 | Wild-type C. thermocellum DSM 1313 | DSMZd |
| C. thermocellum M0970 | DSM 1313 ΔpyrF | This study |
| C. thermocellum M0971 | DSM 1313 ΔpyrF Δpta::gapDHp-cat | This study |
| C. thermocellum M1061 | DSM 1313 ΔpyrF pMU612 | This study |
| C. thermocellum M1062 | DSM 1313 ΔpyrF pMU102 | This study |
a
New England Biolabs, Ipswich, MA.
b
Bacillus Genetic Stock Center; http://www.bgsc.org/.
c
Carlsbad, CA.
d
Deutsche Sammlung von Mikroorganismen und Zellkulturen GmbH, Germany.
e
Ap, ampicillin; Cm, chloramphenicol; Gm, gentamicin.