LOCUS       sequence-485669-        6549 bp    DNA     circular SYN 10-SEP-2025
DEFINITION  This plasmid will be used to replace the promoter of a gene with the
            PenotetSW1 promoter at its endogenous locus.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6549)
  TITLE     Tetracyclin Promoter swap system
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 6549)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Sep 10, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: 
            "Unpublished"
COMMENT     Sequence Label: pFS552
FEATURES             Location/Qualifiers
     source          1..6549
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      1..589
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     490..509
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     743..760
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        847..865
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(990..1010)
                     /label=GFP-F
                     /note="GFP, forward primer. Does NOT anneal to EGFP"
     primer_bind     1605..1633
                     /label=GFP-R
                     /note="GFP, reverse primer. Does NOT anneal to EGFP"
     protein_bind    1724..1742
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     enhancer        2050..2429
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        2430..2633
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     2584..2604
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     2630..2654
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             2735..3358
                     /codon_start=1
                     /gene="tetR from transposon Tn10"
                     /product="tetracycline repressor TetR"
                     /label=TetR
                     /note="TetR binds to the tetracycline operator tetO to 
                     inhibit transcription. This inhibition can be relieved by 
                     adding tetracycline or doxycycline."
                     /translation="MSRLDKSKVINSALELLNEVGIEGLTTRKLAQKLGVEQPTLYWHV
                     KNKRALLDALAIEMLDRHHTHFCPLEGESWQDFLRNNAKSFRCALLSHRDGAKVHLGTR
                     PTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEDQEHQVAKEERETPTT
                     DSMPPLLRQAIELFDHQGAEPAFLFGLELIICGLEKQLKCESGS"
     primer_bind     complement(2808..2827)
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     promoter        complement(3368..3386)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(3396..3413)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(3396..3412)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3405..3427)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     gene            complement(3581..4937)
                     /label=kanMX
                     /note="yeast selectable marker conferring kanamycin 
                     resistance (Wach et al., 1994)"
     terminator      complement(3581..3778)
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     CDS             complement(3784..4593)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin"
                     /translation="MGKEKTHVSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYGKP
                     DAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGKTA
                     FQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDASD
                     FDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGIAD
                     RYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     4507..4526
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     promoter        complement(4594..4937)
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     promoter        complement(5050..5068)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(5051..5068)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(5149..5168)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5268..5290
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5328..5346)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5414..5518
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5519..5587,5588..6379)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(5737..5756)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
       61 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
      121 cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt
      181 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
      241 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
      301 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
      361 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg
      421 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
      481 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
      541 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa
      601 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc
      661 gttatcccct gattctgtgg ataaccgtat taccgccttt gagtgagctg ataccgctcg
      721 ccgcagccga acgaccgagc gcagcgagtc agtgagcgag gaagcggaag agcgcccaat
      781 acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcaggttaa cctggcttat
      841 cgaaattaat acgactcact atagggagac cggcagatcc gcggccgcat aggccactag
      901 tggatctgat atcatcgatg aattcgagct cgtttaaact gtaagatctt ttgtatagtt
      961 catccatgcc atgtgtaatc ccagcagctg ttacaaactc aagaaggacc atgtggtctc
     1021 tcttttcgtt gggatctttc gaaagcttag attgatagga caggtaatgg ttgtctggta
     1081 aaaggacagg gccatcgcca attggagtat tttgttgata atggtctgct agttgaacgc
     1141 ttccatcttc aatgttgtgt ctaattttga agttaacttt gattccattc ttttgtttgt
     1201 ctgccatgat gtatacattg tgtgagttat agttgtattc caatttgtgt ccaagaatgt
     1261 ttccatcttc tttaaaatca atacctttta actcgattct attaacaagg gtatcacctt
     1321 caaacttgac ttcagcacgt gtcttgtagt tcccgtcatc tttgaaaaat atagttcttt
     1381 cctgtacata accttcgggc atggcactct tgaaaaagtc atgccgtttc atatgatctg
     1441 ggtatctagc aaagcacatt agaccataac ccaaagtagt gacaagtgtt ggccatggaa
     1501 caggtagttt tccagtagtg caaataaatt taagggtaag ttttccgtat gttgcatcac
     1561 cttcaccctc tccactgaca gaaaatttgt gcccattaac atcaccatct aattcaacaa
     1621 gaattgggac aactccagtg aaaagttctt ctcctttact catgctagcg cccgagcggc
     1681 cgatgtttac tgtagaatac taattgattc ttactgaaac aagtctctat cactgatagg
     1741 gacactctat atatacctgg aggaagcgcc gatcctctca atcttccatg acatgcccct
     1801 gagcccacag aagccggtgg aacaaattgg tcgcttcttt aagaatcgtt aaccggaacg
     1861 ttagggtacg ttgaaggagc gaccaagcga gcgacgcgag ccaccgtagc caatgtgcga
     1921 gtccagcgag cttagaaggg gcaagagatg gggcccgggg ggtcgaggag cttggcccat
     1981 tgcatacgtt gtatccatat cataatatgt acatttatat tggctcatgt ccaacattac
     2041 cgccatgttg acattgatta ttgactagtt attaatagta atcaattacg gggtcattag
     2101 ttcatagccc atatatggag ttccgcgtta cataacttac ggtaaatggc ccgcctggct
     2161 gaccgcccaa cgacccccgc ccattgacgt caataatgac gtatgttccc atagtaacgc
     2221 caatagggac tttccattga cgtcaatggg tggagtattt acggtaaact gcccacttgg
     2281 cagtacatca agtgtatcat atgccaagta cgccccctat tgacgtcaat gacggtaaat
     2341 ggcccgcctg gcattatgcc cagtacatga ccttatggga ctttcctact tggcagtaca
     2401 tctacgtatt agtcatcgct attaccatgg tgatgcggtt ttggcagtac atcaatgggc
     2461 gtggatagcg gtttgactca cggggatttc caagtctcca ccccattgac gtcaatggga
     2521 gtttgttttg gcaccaaaat caacgggact ttccaaaatg tcgtaacaac tccgccccat
     2581 tgacgcaaat gggcggtagg cgtgtacggt gggaggtcta tataagcaga gctcgtttag
     2641 tgaaccgtca gatcgcctgg agacgccatc cacgctgttt tgacctccat agaagacacc
     2701 gggaccgatc cagcctccgc ggccccgaat tcatatgtct agattagata aaagtaaagt
     2761 gattaacagc gcattagagc tgcttaatga ggtcggaatc gaaggtttaa caacccgtaa
     2821 actcgcccag aagctaggtg tagagcagcc tacattgtat tggcatgtaa aaaataagcg
     2881 ggctttgctc gacgccttag ccattgagat gttagatagg caccatactc acttttgccc
     2941 tttagaaggg gaaagctggc aagatttttt acgtaataac gctaaaagtt ttagatgtgc
     3001 tttactaagt catcgcgatg gagcaaaagt acatttaggt acacggccta cagaaaaaca
     3061 gtatgaaact ctcgaaaatc aattagcctt tttatgccaa caaggttttt cactagagaa
     3121 tgcattatat gcactcagcg ctgtggggca ttttacttta ggttgcgtat tggaagatca
     3181 agagcatcaa gtcgctaaag aagaaaggga aacacctact actgatagta tgccgccatt
     3241 attacgacaa gctatcgaat tatttgatca ccaaggtgca gagccagcct tcttattcgg
     3301 ccttgaattg atcatatgcg gattagaaaa acaacttaaa tgtgaaagtg gatcttaacc
     3361 aattcgccct atagtgagtc gtattacgcg cgctcactgg ccgtcgtttt acaacgtcgt
     3421 gactgggaaa accctggcgt tacccaactt aatcgccttg cagcacatcc ccctttcgcc
     3481 agctggcgta atagcgaaga ggcccgcacc gatcgccctt cccaacagtt gcgcagcctg
     3541 aatggcgaat gggatggcgg cgttagtatc gaatcgacag cagtatagcg accagcattc
     3601 acatacgatt gacgcatgat attactttct gcgcacttaa cttcgcatct gggcagatga
     3661 tgtcgaggcg aaaaaaaata taaatcacgc taacatttga ttaaaataga acaactacaa
     3721 tataaaaaaa ctatacaaat gacaagttct tgaaaacaag aatcttttta ttgtcagtac
     3781 tgattagaaa aactcatcga gcatcaaatg aaactgcaat ttattcatat caggattatc
     3841 aataccatat ttttgaaaaa gccgtttctg taatgaagga gaaaactcac cgaggcagtt
     3901 ccataggatg gcaagatcct ggtatcggtc tgcgattccg actcgtccaa catcaataca
     3961 acctattaat ttcccctcgt caaaaataag gttatcaagt gagaaatcac catgagtgac
     4021 gactgaatcc ggtgagaatg gcaaaagctt atgcatttct ttccagactt gttcaacagg
     4081 ccagccatta cgctcgtcat caaaatcact cgcatcaacc aaaccgttat tcattcgtga
     4141 ttgcgcctga gcgagacgaa atacgcgatc gctgttaaaa ggacaattac aaacaggaat
     4201 cgaatgcaac cggcgcagga acactgccag cgcatcaaca atattttcac ctgaatcagg
     4261 atattcttct aatacctgga atgctgtttt gccggggatc gcagtggtga gtaaccatgc
     4321 atcatcagga gtacggataa aatgcttgat ggtcggaaga ggcataaatt ccgtcagcca
     4381 gtttagtctg accatctcat ctgtaacatc attggcaacg ctacctttgc catgtttcag
     4441 aaacaactct ggcgcatcgg gcttcccata caatcgatag attgtcgcac ctgattgccc
     4501 gacattatcg cgagcccatt tatacccata taaatcagca tccatgttgg aatttaatcg
     4561 cggcctcgaa acgtgagtct tttccttacc catggttgtt tatgttcgga tgtgatgtga
     4621 gaactgtatc ctagcaagat tttaaaagga agtatatgaa agaagaacct cagtggcaaa
     4681 tcctaacctt ttatatttct ctacaggggc gcggcgtggg gacaattcaa cgcgtctgtg
     4741 aggggagcgt ttccctgctc gcaggtctgc agcgaggagc cgtaattttt gcttcgcgcc
     4801 gtgcggccat caaaatgtat ggatgcaaat gattatacat ggggatgtat gggctaaatg
     4861 tacgggcgac agtcacatca tgcccctgag ctgcgcacgt caagactgtc aaggagggta
     4921 ttctgggcct ccatgtcgct ggccgggtga cccggcgggg acgaggcaag ctaaacagat
     4981 ctggcgcgcc ttaattaacc cggggatccg tcgacctgca gcgtacgaag cttcagctgg
     5041 cggccgcgtt ctatagtgtc acctaaatcg tatgtgtatg atacataagg ttatgtatta
     5101 attgtagccg cgttctaacg acaatatgtc catatggtgc actctcagta caatctgctc
     5161 tgatgccgca tagttaagcc agccccgaca cccgccaaca cccgctgacg cgccctgacg
     5221 ggcttgtctg ctcccggcat ccgcttacag acaagctgtg accgtctccg ggagctgcat
     5281 gtgtcagagg ttttcaccgt catcaccgaa acgcgcgaga cgaaagggcc tcgtgatacg
     5341 cctattttta taggttaatg tcatgataat aatggtttct tagacgtcag gtggcacttt
     5401 tcggggaaat gtgcgcggaa cccctatttg tttatttttc taaatacatt caaatatgta
     5461 tccgctcatg agacaataac cctgataaat gcttcaataa tattgaaaaa ggaagagtat
     5521 gagtattcaa catttccgtg tcgcccttat tccctttttt gcggcatttt gccttcctgt
     5581 ttttgctcac ccagaaacgc tggtgaaagt aaaagatgct gaagatcagt tgggtgcacg
     5641 agtgggttac atcgaactgg atctcaacag cggtaagatc cttgagagtt ttcgccccga
     5701 agaacgtttt ccaatgatga gcacttttaa agttctgcta tgtggcgcgg tattatcccg
     5761 tattgacgcc gggcaagagc aactcggtcg ccgcatacac tattctcaga atgacttggt
     5821 tgagtactca ccagtcacag aaaagcatct tacggatggc atgacagtaa gagaattatg
     5881 cagtgctgcc ataaccatga gtgataacac tgcggccaac ttacttctga caacgatcgg
     5941 aggaccgaag gagctaaccg cttttttgca caacatgggg gatcatgtaa ctcgccttga
     6001 tcgttgggaa ccggagctga atgaagccat accaaacgac gagcgtgaca ccacgatgcc
     6061 tgtagcaatg gcaacaacgt tgcgcaaact attaactggc gaactactta ctctagcttc
     6121 ccggcaacaa ttaatagact ggatggaggc ggataaagtt gcaggaccac ttctgcgctc
     6181 ggcccttccg gctggctggt ttattgctga taaatctgga gccggtgagc gtgggtctcg
     6241 cggtatcatt gcagcactgg ggccagatgg taagccctcc cgtatcgtag ttatctacac
     6301 gacggggagt caggcaacta tggatgaacg aaatagacag atcgctgaga taggtgcctc
     6361 actgattaag cattggtaac tgtcagacca agtttactca tatatacttt agattgattt
     6421 aaaacttcat ttttaattta aaaggatcta ggtgaagatc ctttttgata atctcatgac
     6481 caaaatccct taacgtgagt tttcgttcca ctgagcgtca gaccccgtag aaaagatcaa
     6541 aggatcttc
//
