Table 1.
Different options of restriction enzyme sites for designing MuST transposon variants.
| Restriction enzyme site at the 5′end | Restriction enzyme site at the 3′end | Nucleotides encoding Inter-domain linker | Inter-domain linker composition | ||||||||||||||
|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
| T+TfiI | 4 | 5 | T | G | A | A | T | C | N | X-Glu-Ser | |||||||
| 4 | 5 | A | C | T | T | A | G | N | |||||||||
| BsrGI | 4 | 5 | T | G | T | A | C | A | N | X-Val-His/Gln | |||||||
| 4 | 5 | A | C | A | T | G | T | N | |||||||||
| TG+Hind III | 4 | 5 | T | G | A | A | G | C | T | T | N | N | X-Glu-Ala-Y | ||||
| 4 | 5 | A | C | T | T | C | G | A | A | N | N | ||||||
| TGAAG +EagI | 4 | 5 | T | G | A | A | G | C | G | G | C | C | G | N | N | X-Glu-Ala-Ala-Z | |
| 4 | 5 | A | C | T | T | C | G | C | C | G | G | C | N | N | |||
| BssHII +CTTCA | N | G | C | G | C | G | C | C | T | T | C | A | Gly/Ser/Arg/Cys-Ala-Pro-Ser | ||||
| N | C | G | C | G | C | G | G | A | A | G | T | ||||||
| BtsI+TCA | G | C | A | G | T | G | T | C | A | Ala-Val-Ser | |||||||
| C | G | T | C | A | C | A | G | T | |||||||||
| NgoMIV +TCA | G | C | C | G | G | C | T | C | A | Ala-Gly-Ser | |||||||
| C | G | G | C | C | G | A | G | T | |||||||||
Note. The numbers represent nucleotide sequences derived from a host DNA. N represents an additional nucleotide placed either upstream of the 5′ end or downstream of the 3′end of the guest DNA to ensure inter-domain linker in second and first reading frames, at the N and C terminus of the guest protein, respectively. The mutated nucleotides introduced in the MuST transposon variants compared to the wild-type Mu transposon are shown with underlines. X represents one of 15 amino acids (i.e., Ala, Arg, Asn, Asp, Cys, Gly, His, Ile, Leu, Phe, Pro, Ser (twice), Thr, Tyr, and Val) encoded by codons containing 3′ thymine. Y represents one of 6 possible amino acids (i.e. Phe, Leu, Ser, Tyr, Cys, Trp) encoded by codons containing 5′ thymine. Z represents one of 5 possible amino acids (i.e., Val, Ala, Asp, Glu, Gly) encoded by codons containing 5′ guanine.