LOCUS       sequence-252225-        5388 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Tagging plasmid for WTC 846: Encodes the PtetO7.1 promoter and Hyg 
            resistance gene.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5388)
  AUTHORS   Azizoglu A, Brent R, Rudolf F
  TITLE     A precisely adjustable, variation-suppressed eukaryotic 
            transcriptional controller to enable genetic discovery.
  JOURNAL   Elife. 2021 Aug 3;10. pii: 69549. doi: 10.7554/eLife.69549.
  PUBMED    34342575
REFERENCE   2  (bases 1 to 5388)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Elife."; 
            date: "2021-08-3"; volume: "10. pii: 69549. doi"; pages: " 
            10.7554/eLife.69549"
COMMENT     Sequence Label: FRP2350
FEATURES             Location/Qualifiers
     source          1..5388
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     1..17
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     24..43
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        24..42
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     gene            102..1677
                     /label=hphMX6
                     /note="yeast selectable marker conferring hygromycin 
                     resistance"
     promoter        102..445
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     CDS             446..1474
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MGKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGR
                     GYVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDL
                     PETELPAVLQPVAGAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHW
                     QTVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAM
                     FGDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVD
                     GNFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRA
                     KE"
     terminator      1480..1677
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     CDS             2467..2538
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags"
                     /label=3xFLAG
                     /translation="DYKDDDDKDYKDDDDKDYKDDDDK"
     promoter        complement(2601..2619)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(2640..2656)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2640..2656)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2653..2675)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2664..2680
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(2688..2694,2695..2712,2713..2718))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2733..2754
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2871..2888)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3042..3630)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3122..3141)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3801..4592,4593..4661))
                     /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     4424..4443
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4662..4766)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(4792..5247)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4929..4950)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     5141..5160
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     join(5374..5388,1..8)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     join(5388,1..17)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
ORIGIN
        1 gtaaaacgac ggccagtgaa ttgtaatacg actcactata gggcgaattg gagctcgatc
       61 ctcgagagct tgccttgtcc ccgccgggtc acccggccag cgacatggag gcccagaata
      121 ccctccttga cagtcttgac gtgcgcagct caggggcatg atgtgactgt cgcccgtaca
      181 tttagcccat acatccccat gtataatcat ttgcatccat acattttgat ggccgcacgg
      241 cgcgaagcaa aaattacggc tcctcgctgc ggacctgcga gcagggaaac gctcccctca
      301 cagacgcgtt gaattgtccc cacgccgcgc ccctgtagag aaatataaaa ggttaggatt
      361 tgccactgag gttcttcttt catatacttc cttttaaaat cttgctagga tacagttctc
      421 acatcacatc cgaacataaa caaccatggg taaaaagcct gaactcaccg cgacgtctgt
      481 cgagaagttt ctgatcgaaa agttcgacag cgtctccgac ctgatgcagc tctcggaggg
      541 cgaagaatct cgtgctttca gcttcgatgt aggagggcgt ggatatgtcc tgcgggtaaa
      601 tagctgcgcc gatggtttct acaaagatcg ttatgtttat cggcactttg catcggccgc
      661 gctcccgatt ccggaagtgc ttgacattgg ggagttcagc gagagcctga cctattgcat
      721 ctcccgccgt gcacagggtg tcacgttgca agacctgcct gaaaccgaac tgcccgctgt
      781 tctgcagccg gtcgcggggg ccatggatgc gatcgctgcg gccgatctta gccagacgag
      841 cgggttcggc ccattcggac cgcaaggaat cggtcaatac actacatggc gtgatttcat
      901 atgcgcgatt gctgatcccc acgtgtatca ctggcaaact gtgatggacg acaccgtcag
      961 tgcgtccgtc gcgcaggctc tcgatgagct gatgctttgg gccgaggact gccccgaagt
     1021 ccggcacctc gtgcacgcgg atttcggctc caacaatgtc ctgacggaca atggccgcat
     1081 aacagcggtc attgactgga gcgaggcgat gttcggggat tcccaatacg aggtcgccaa
     1141 catcttcttc tggaggccgt ggttggcttg tatggagcag cagacgcgct acttcgagcg
     1201 gaggcatccg gagcttgcag gatcgccgcg gctccgggcg tatatgctcc gcattggtct
     1261 tgaccaactc tatcagagct tggttgacgg caatttcgat gatgcagctt gggcgcaggg
     1321 tcgatgcgac gcaatcgtcc gatccggagc cgggactgtc gggcgtacac aaatcgcccg
     1381 cagaagcgcg gccgtctgga ccgatggctg tgtagaagta ctcgccgata gtggaaaccg
     1441 acgccccagc actcgtccga gggcaaagga ataatcagta ctgacaataa aaagattctt
     1501 gttttcaaga acttgtcatt tgtatagttt ttttatattg tagttgttct attttaatca
     1561 aatgttagcg tgatttatat tttttttcgc ctcgacatca tctgcccaga tgcgaagtta
     1621 agtgcgcaga aagtaatatc atgcgtcaat cgtatgtgaa tgctggtcgc tatactgctg
     1681 tcgattcgat actaacgccg ccatccagtg tttaaacgaa ctagtgcggc cgccagttcg
     1741 agtttatcat tatcaatact gccatttcaa agaatacgta aataattaat agtagtgatt
     1801 ttcctaactt tatttagtca aaaaattagc ctctatcatt gatagagtgt ctgggtgatc
     1861 tatcattgat agagcatcca ctaattttaa ttctgctgta acccgtacat gcccaaaata
     1921 gggggcgggt tacacagaat ctatcattga tagagtgtct gggtgatcta tcattgatag
     1981 agcatccact aaatataatg gagctctatc attgatagag catccactaa aaaaaagaat
     2041 cccagcacca aaatattgtt ttcttcacca accatcagtt cataggtcca ttctcttagc
     2101 gcaactacag agaacagggg cacaaacagg caaaaaacgg gcacaacctc aatggagtga
     2161 tgcaacctgc ctggagtaaa tgatgacaca aggcaattga cccacgcatg tatctatctc
     2221 attttcttac accttctatt accttctgct ctctctgatt tggaaaaagc tgaaaaaaaa
     2281 ggttgaaacc agttccctga aattattccc ctatctatca ttgatagata taaatatcta
     2341 tcattgatag agtaattctg taaatctatt tcttaaactt cttaaattct acttttatag
     2401 ttagtctttt ttttagtttt aaaacaccaa gaacttagtt tcgaataaac acacataaac
     2461 aaaatggatt ataaagatga cgatgacaag gattataaag atgacgatga caaggattat
     2521 aaagatgacg atgacaaggg aggtggtggt ggtggaggcg gtccgacaga gaatcatcga
     2581 tgggtaccca gcttttgttc cctttagtga gggttaattt cgagcttggc gtaatcatgg
     2641 tcatagctgt ttcctgtgtg aaattgttat ccgctcacaa ttccacacaa catacgagcc
     2701 ggaagcataa agtgtaaagc ctggggtgcc taatgagtga gctaactcac attaattgcg
     2761 ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt gccagctgca ttaatgaatc
     2821 ggccaacgcg cggggagagg cggtttgcgt attgggcgct cttccgcttc ctcgctcact
     2881 gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat cagctcactc aaaggcggta
     2941 atacggttat ccacagaatc aggggataac gcaggaaaga acatgtgagc aaaaggccag
     3001 caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag gctccgcccc
     3061 cctgacgagc atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc gacaggacta
     3121 taaagatacc aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt tccgaccctg
     3181 ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc
     3241 tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac
     3301 gaaccccccg ttcagcccga ccgctgcgcc ttatccggta actatcgtct tgagtccaac
     3361 ccggtaagac acgacttatc gccactggca gcagccactg gtaacaggat tagcagagcg
     3421 aggtatgtag gcggtgctac agagttcttg aagtggtggc ctaactacgg ctacactaga
     3481 agaacagtat ttggtatctg cgctctgctg aagccagtta ccttcggaaa aagagttggt
     3541 agctcttgat ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag
     3601 cagattacgc gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc tacggggtct
     3661 gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt atcaaaaagg
     3721 atcttcacct agatcctttt aaattaaaaa tgaagtttta aatcaatcta aagtatatat
     3781 gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc
     3841 tgtctatttc gttcatccat agttgcctga ctccccgtcg tgtagataac tacgatacgg
     3901 gagggcttac catctggccc cagtgctgca atgataccgc gagacccacg ctcaccggct
     3961 ccagatttat cagcaataaa ccagccagcc ggaagggccg agcgcagaag tggtcctgca
     4021 actttatccg cctccatcca gtctattaat tgttgccggg aagctagagt aagtagttcg
     4081 ccagttaata gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt gtcacgctcg
     4141 tcgtttggta tggcttcatt cagctccggt tcccaacgat caaggcgagt tacatgatcc
     4201 cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag
     4261 ttggccgcag tgttatcact catggttatg gcagcactgc ataattctct tactgtcatg
     4321 ccatccgtaa gatgcttttc tgtgactggt gagtactcaa ccaagtcatt ctgagaatag
     4381 tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac gggataatac cgcgccacat
     4441 agcagaactt taaaagtgct catcattgga aaacgttctt cggggcgaaa actctcaagg
     4501 atcttaccgc tgttgagatc cagttcgatg taacccactc gtgcacccaa ctgatcttca
     4561 gcatctttta ctttcaccag cgtttctggg tgagcaaaaa caggaaggca aaatgccgca
     4621 aaaaagggaa taagggcgac acggaaatgt tgaatactca tactcttcct ttttcaatat
     4681 tattgaagca tttatcaggg ttattgtctc atgagcggat acatatttga atgtatttag
     4741 aaaaataaac aaataggggt tccgcgcaca tttccccgaa aagtgccacc taaattgtaa
     4801 gcgttaatat tttgttaaaa ttcgcgttaa atttttgtta aatcagctca ttttttaacc
     4861 aataggccga aatcggcaaa atcccttata aatcaaaaga atagaccgag atagggttga
     4921 gtgttgttcc agtttggaac aagagtccac tattaaagaa cgtggactcc aacgtcaaag
     4981 ggcgaaaaac cgtctatcag ggcgatggcc cactacgtga accatcaccc taatcaagtt
     5041 ttttggggtc gaggtgccgt aaagcactaa atcggaaccc taaagggagc ccccgattta
     5101 gagcttgacg gggaaagccg gcgaacgtgg cgagaaagga agggaagaaa gcgaaaggag
     5161 cgggcgctag ggcgctggca agtgtagcgg tcacgctgcg cgtaaccacc acacccgccg
     5221 cgcttaatgc gccgctacag ggcgcgtccc attcgccatt caggctgcgc aactgttggg
     5281 aagggcgatc ggtgcgggcc tcttcgctat tacgccagct ggcgaaaggg ggatgtgctg
     5341 caaggcgatt aagttgggta acgccagggt tttcccagtc acgacgtt
//
