TABLE 1.
Strains, plasmids, and oligonucleotides used in this study
| Strain, plasmid, or oligonucleotide | Characteristic, description, or sequence (5′–3′)a | Reference, source, or restriction site |
|---|---|---|
| Strains | ||
| N. nova SH22a | GP- and rubber-degrading wild type | 2 |
| N. nova SH22a SE3-4-18 | Transposon-induced mutant, GP- and rubber-degradation defective, Aprr | This study |
| N. nova SH22a SE13-5-5 | Transposon-induced mutant, GP- and rubber-degradation defective, Aprr | This study |
| N. nova SH22a SE16-4-35 | Transposon-induced mutant, GP- and rubber-degradation defective, Aprr | This study |
| N. nova SH22a SE18-1-3 | Transposon-induced mutant, GP- and rubber-degradation defective, Aprr | This study |
| N. nova SH22a SE18-8-1 | Transposon-induced mutant, GP- and rubber-degradation defective, Aprr | This study |
| N. nova SH22a OC11-7-29 | Transposon-induced mutant, GP- and rubber-degradation defective, Aprr | This study |
| N. nova SH22a OC20-2-42 | Transposon-induced mutant, GP- and rubber-degradation defective, Aprr | This study |
| N. nova SH22a SE10-4-21 | Transposon-induced mutant, GP- and rubber-degradation defective, Aprr | This study |
| N. nova SH22a OC14-14-5 | Transposon-induced mutant, GP-degradation defective, Aprr | This study |
| N. nova SH22a SE9-8-16 | Transposon-induced mutant, GP- and rubber-degradation defective, Aprr | This study |
| N. nova SH22a OC19-4-17 | Transposon-induced mutant, GP- and rubber-degradation defective, Aprr | This study |
| N. nova SH22a OC22-3-17 | Transposon-induced mutant, GP- and rubber-degradation defective, Aprr | This study |
| N. nova SH22a OC26-3-28 | Transposon-induced mutant, GP- and rubber-degradation defective, Aprr | This study |
| N. nova SH22a ΔfadD32SH22a | fadD32SH22a deletion mutant, GP- and rubber-degradation defective, Kmr | This study |
| N. nova SH22a Δpck | pck deletion mutant, GP- and rubber-degradation defective, Kmr | This study |
| E. coli XL10-Gold | Tetr Δ(mcrA)183 Δ(mcrCB-hsdSMR-mrr)173 endA1 supE44 thi-1 recA1 gyrA96 relA1 lac Hte [F′ proAB lacIqZΔM15 Tn10 (Tetr) Amy Camr] | Agilent Technologies |
| Plasmids | ||
| pMA5096 | Contains transposon Tn5096, Ampr Aprr | 12 |
| pSKΩKm | Kanamycin resistance cassette-containing plasmid | Strain SK07729b |
| pJET1.2/blunt | Blunt-end cloning vector, Ampr | Thermo Fisher Scientific |
| pJET::FAALKN | Used for fadD32SH22a deletion experiment, Ampr | This study |
| pJET::FAALKN::Km | Used for fadD32SH22a deletion experiment, Ampr Kmr | This study |
| pJET::PCKKN | Used for pck deletion experiment, Ampr | This study |
| pJET::PCKKN::Km | Used for pck deletion experiment, Ampr Kmr | This study |
| pNV18.1 | E. coli-Nocardia shuttle vector, Kmr Neor | 96 |
| pNV18.1::fadD32SH22a | Used for complementation experiment, Kmr Neor | This study |
| pNV18.1::pck | Used for complementation experiment, Kmr Neor | This study |
| Oligonucleotides | ||
| FAAL-kn1 | TGAGGGCAACCAGCAGGAG | |
| FAAL-kn2 | CAGAACTCGCCGGGCGCACGATCCCGGGGGCCGCTGATACCTGTGAG | SmaI |
| FAAL-kn3 | CTCACAGGTATCAGCGGCCCCCGGGATCGTGCGCCCGGCGAGTTCTG | SmaI |
| FAAL-kn4 | GATGAAGACGCCGACCGCCTC | |
| PCK-kn1 | AAACATATGGTGGATCTGACGCACGAGG | NdeI |
| PCK-kn2 | CGTCCGAAAAAGCCTATCCCAGGTCGACGAAAACTCTCCTGAGATGAGCG | HincII |
| PCK-kn3 | CGCTCATCTCAGGAGAGTTTTCGTCGACCTGGGATAGGCTTTTTCGGACG | HincII |
| PCK-kn4 | AAACATATGGAACCGCACCCACACCTC | NdeI |
| FAAL-exp1 | GAAGACCATCGAGAAGGTG | |
| FAAL-exp2 | GATGCTGATACACGATGGTC | |
| PCK-exp2 | AGTCCGGTCATGCCCACAC |
a
Ampr, ampicillin resistance; Aprr, apramycin resistance; Camr, chloramphenicol resistance; Gmr, gentamicin resistance; Kmr, kanamycin resistance; Neor, neomycin resistance; Tetr, tetracycline resistance. Underlining in the sequences indicates the restriction endonuclease sites for DNA cloning.
b
From the strain collection of our lab.