LOCUS       sequence-329937-        8725 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Punc-4::rtetR-QFAD::P2A::mKate::T2A::tetR-pie1.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8725)
  AUTHORS   Mao S, Qi Y, Zhu H, Huang X, Zou Y, Chi T
  TITLE     A Tet/Q Hybrid System for Robust and Versatile Control of Transgene 
            Expression in C. elegans.
  JOURNAL   iScience. 2019 Jan 25;11:224-237. doi: 10.1016/j.isci.2018.12.023. 
            Epub 2018 Dec 27.
  PUBMED    30634168
REFERENCE   2  (bases 1 to 8725)
  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: "iScience.";
            date: "2019-01-25"; volume: "11:224-237. doi"; pages: " 
            10.1016/j.isci.2018.12.023. Epub 2018 Dec 27"
COMMENT     Sequence Label: TC550
FEATURES             Location/Qualifiers
     source          1..8725
                     /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"
     CDS             4466..4522
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     CDS             5381..5434
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     primer_bind     complement(7285..7307)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     7407..7426
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(7504..7522)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        7590..7694
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(7695..7763,7764..8555)
                     /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(7913..7932)
                     /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 taccgcggcc gccaacaata taggatgctc
      721 aggagactat aattcaaaaa acccaaccca aattagcata agcttcccca aattggaaca
      781 gtgaaatact caatcaatca ctctggcggt tttctccaca aagaacacga aattggaaaa
      841 aggaaaagat tgatttgttt gaaggcgtga ggaaatcgaa aaaaaattgt ttcaaaacaa
      901 atgaaatatt ttaacgaata aacccaggaa tgccactttt tggtgcccgc ctacaaaaaa
      961 gtcaactgtt cttaaactcc tacatagttg ccctctctaa cgaatgttgc tttttttaaa
     1021 ttttcattga atttgattat cgtctgtatg cgcctactct atcagtggaa aacatgccca
     1081 tttcccacaa atttactcgt ttggagctca aaatgttaaa tggggcatca gggaattggc
     1141 atctcacaga aagtggcgga tgagttatat gtaataataa tggtggctga agtggaaacc
     1201 gcaaaaaaag gatgtgtatt tgagattttg caacggaatt tgaatcagat acacaaaatg
     1261 agagaaattt atagattctt taaaatccac ctgaagtcgg caaatttttg aaaggtctgt
     1321 tcgttcagtt tttcataagt tttatgaatt ttgatttact actgaagttc gagaaaagcc
     1381 ctttaaccat accaataaga aattatgtta actttacaag aatattaaca aaaaattatt
     1441 aaaaaactgt gttcccctgg cattatgata tttccaccgt cagttctcta ttaaaacttc
     1501 aactctacac agtttcaact ctcaaatgag ctgtccatct tttgccttca ggaaattatg
     1561 ctaatctgct gatttcgcgc actttcacca cactatttcc aaatagaagg aggggtggtg
     1621 caacacacga gacactgcaa aaaggttcag tgaagattta tctacttgtg agatgagagc
     1681 aggaggggtg ggagaagtgt gatgattgac gggatgacaa gaacagtgat gtggtgtcga
     1741 aatggaagat gcgctaattt gtggagtttt atgaaatttg aataattaca tttataacat
     1801 ctactagtta ggcgtctgtg taggtattgc ctgcctgcct acaaggcata tgtacgaggc
     1861 aggttaaacc tcctagaaag ctacggtact ttcaaaccac ataacattta tactggtgtc
     1921 ttttcgcgtc ttcttcttga acatgtagac cctcaaaaag ctaagtttta cactgtgtag
     1981 tttgctttga aatctacaaa aattggaggc acgccttctg gactttgccg acacgcctac
     2041 tttctaccta ccaactgcct acttccacag tctagcgtga actccaatac cattttaaaa
     2101 attgttttat ctcaaatttt caatttttaa atccaatttc ccagttctaa aaaaaaaatt
     2161 ttttttttaa tttaaaaact ctcatccttt ttaatgctcg ttcccattcg gcaagacgtg
     2221 taccaaacac aaaacaaatt gactggtgat ccatctcgaa tttctcgttt caaattagtc
     2281 gttgctatat atattgtctt atgtgtggaa aacgaataag aaaaattaga gaaagatgag
     2341 agagcagggg cagaatgaaa atttgcatgg aaattgaaga agatcactgc tttttccgat
     2401 gaatgatgac taaatagaat gagttgagag gaaaataatc ataattctaa ttttaattag
     2461 aaatatgata tggaaagtaa gcaaaaccac aaggcttttg ctctgaactt atttccgaaa
     2521 aaaaaagtga aactgaaagt tagaattttc aaaataaata ttgcgagagt caaccttcag
     2581 aaaaattggc gagaaatcaa atttgaattc tctgaaaatt tagttaaaat acatatgtta
     2641 tccttctttg ttttttggcc ttagcctgcc ttcaaggcga cctgagccta tgcgggtgcc
     2701 taataccgaa acacacataa tttttttttt taatctttaa atactgtaac gagtcgcgag
     2761 atccagcgag acccatagaa tatgttgagc gctgcaaaca atcgtataac tcttgtccaa
     2821 atttttccac tctgtatttt tgcaagcagt tcaacaaatg gtcatcattt ttataaccca
     2881 tatctatatt cgaatctgac ataacaaata atactaagca catcctccca catctctttt
     2941 cattcaagtt gcgcaattcc cctaataact atggatggag gggggaccga gcccaaagat
     3001 tattcaaaac gataacgact ttttcatgcg gccaacgaaa aataaaactt ttctcatatt
     3061 ctcctatttc ccatttcaag aattacatcc ccatgggacg ttgaaatcac atcatattag
     3121 tatatgttaa ttgttctttt cactggaata tcgccacgcc ccccttccgg tggctcattg
     3181 gtttcgagaa gaaggcggag ccgaagtggg ttcttctttc ttttttttgt ggtagaagtc
     3241 tagcaggatt aagctactat tccattagag gatcttcttc ttccaaaaag tgaaaatgtc
     3301 acgactggat aaatcgaagg tcatcaactc tgctctggaa ctgctgaatg gagtgggaat
     3361 tgaaggactt acaacccgaa agcttgcaca aaaattggga gttgaacaac caacacttta
     3421 ctggcatgtt aagaataagc gagcactttt ggatgctctt ccaattgaaa tgttggatag
     3481 acatcatact cattcatgtc cacttgaagg agaatcttgg caagatttct tgagaaacaa
     3541 cgcaaagtca taccgatgcg ctcttttgtc tcatagagat ggagcaaaag ttcatcttgg
     3601 aacacgacca actgaaaagc aatacgaaac attggaaaac caacttgctt tcttgtgtca
     3661 acaaggattt tcattggaaa atgctcttta cgcattgtct gctgttggac attttacact
     3721 tggatgcgtt ttggaagaac aagaacatca agttgcaaaa gaagaacgag aaactccaac
     3781 aactgattca atgccaccac ttttgaagca agctatcgaa cttttcgata gacaaggagc
     3841 agaaccagct ttcctttttg gactggaact tattatctgt ggacttgaga aacaactgaa
     3901 atgtgaatcg ggaggaggat ctggatctgg atctggatct ggttccggtc gacaacttga
     3961 gcttccgccg actgcgactg ccactgcaag tatcatgccc catgtcatgg aacagcccct
     4021 gtcaacgtca ataaatccgg tcaatgatcg gtttaatgga ataccgaatc ctacccccta
     4081 caatagcgat gcggccctag acgcaattac tcaaaccaat gactacggat cagtcaatac
     4141 ccatggcatt ctcagtacat atccaccacc tgccacccac ttgaacgagg cctcggtggc
     4201 ccttgcgcct ggaggtgctc ctcccagacc acccccgcca tacgttgata gtacaacgaa
     4261 ccatcctcct taccatagta acctcgtccc catggccaat ttcggatatt cgactgtaga
     4321 ctatgatgcg atggtggatg atctggcatc gatcgagtac acggatgcgg ttgacgttga
     4381 tccgcagttc atgacgaacc tcgggtttgt cccagggtgt aattttagcg atatcaacac
     4441 gtatgagcaa ccatggggca gtggggctac taacttcagc ctgctgaagc aggctggaga
     4501 cgtggaggag aaccctggac ctatggtctc cgagctcatt aaagaaaaca tgcatatgaa
     4561 gctctacatg gagggaaccg tcaacaacca ccacttcaag tgcacctccg agggagaggg
     4621 aaagccatac gagggaaccc aaaccatgcg tatcaaggcc gtcgagggag gaccactccc
     4681 attcgccttc gacatcctcg ccacctcctt catgtacgga tccaaggtaa gtttaaacat
     4741 atatatacta actaaccctg attatttaaa ttttcagacc ttcatcaacc acacccaagg
     4801 aatcccagac ttcttcaagc aatccttccc agagggattc acctgggagc gtgtcaccac
     4861 ctacgaggac ggaggagtcc tcaccgccac ccaagacacc tccctccaag acggatgcct
     4921 catctacaac gtcaaggtaa gtttaaacag ttcggtacta actaaccata catatttaaa
     4981 ttttcagatc cgtggagtca acttcccatc caacggacca gtcatgcaaa agaagaccct
     5041 cggatgggag gcctccaccg agaccctcta cccagccgac ggaggacttg agggacgtgc
     5101 cgacatggcc ctcaagctcg tcggaggagg acacctcatc tgcaacctca aggtaagttt
     5161 aaacatgatt ttactaacta actaatctga tttaaatttt cagaccacct accgttccaa
     5221 gaagccagcc aagaacctca agatgccagg agtctactac gtcgaccgtc gtcttgagcg
     5281 tatcaaggag gccgacaagg agacctacgt cgagcaacac gaggtcgccg tcgcccgtta
     5341 ctgcgacctc ccatccaagc tcggacaccg tggatcggga gagggacgag gaagtctttt
     5401 gacctgcgga gatgttgagg agaacccagg accaatgtcg cgtcttgata aatcgaaagt
     5461 tattaattca gctcttgaac ttttgaacga agttggaatt gaaggattga caactagaaa
     5521 gcttgcacaa aagttgggag ttgaacaacc aacactttat tggcatgttc gaaataagca
     5581 aactcttatg aacatgttgt ctgaagctat tcttgcaaaa catcatacaa gatcagctcc
     5641 attgccaact gaatcttggc aacaattcct tcaagaaaat gctttgtcat ttcgaaaagc
     5701 acttttggtt catagagatg gagcacgatt gcatattgga acatctccaa ctccaccaca
     5761 attcgaacaa gctgaagcac aacttagatg tttgtgcgat gctggattct cagttgaaga
     5821 agcacttttc atcttgcaat caatctctca tttcacactt ggagctgttt tggaagaaca
     5881 agcaactaac caaatcgaaa acaaccatgt tattgatgct gcaccaccac ttttgcaaga
     5941 agctttcaat attcaagcac gaacatctgc tgaaatggca ttccatttcg gacttaagtc
     6001 attgatcttc ggattttctg ctcaacttga tgaaaagaaa catactccaa ttgaagatgg
     6061 aaacaagcca aagaagaaaa gaaaactggg aagtggaagt aatggaagta gtggatccag
     6121 atctgcgcca ccttcttccg attgtccgcc gccgattgcc tactaccatc atcatccaca
     6181 acatcagcaa caattcctgc catttccaat gccatatttc ttggctccac cgccgcaagc
     6241 tcaacaagga gctccttttc cagtgcaata tattccacag caacatgatt tgatgaatag
     6301 ccagccaatg tatgcaccaa tggcaccgac atactactat caaccaatta attcgaatgg
     6361 catgcccatg aattccaact gagcgccggt cgctaccatt accaacttgt ctggtgtcaa
     6421 aaataatagg ggccgctgtc atcagagtaa gtttaaactg agttctacta actaacgagt
     6481 aatatttaaa ttttcagcat ctcgcgcccg tgcctctgac ttctaagtcc aattactctt
     6541 caacatccct acatgctctt tctccctgtg ctcccacccc ctatttttgt tattatcaaa
     6601 aaacttctct taatttcttt gttttttagc ttcttttaag tcacctctaa caatgaaatt
     6661 gtgtagattc aaaaatagaa ttaattcgta ataaaaagtc gaaaaaaatt gtgctccctc
     6721 cccccattaa taataattct atcccaaaat ctacacaatg ttctgtgtac acttcttatg
     6781 ttttttactt ctgataaatt tttttgaaac atcatagaaa aaaccgcaca caaaatacct
     6841 tatcatatgt tacgtttcag tttatgaccg caatttttat ttcttcgcac gtctgggcct
     6901 ctcatgacgt caaatcatgc tcatcgtgaa aaagttttgg agtatttttg gaatttttca
     6961 atcaagtgaa agtttatgaa attaattttc ctgcttttgc tttttggggt ttcccctatt
     7021 gtttgtcaag atttcgagga cggcgttttt cttgctaaaa tcacaagtat tgatgagcac
     7081 gatgcaagaa agatcggaag aaggtttggg tttgaggctc agtggaaggt gagtagaagt
     7141 tgataatttg aaagtggagt agtgtctatg gggtttttgc cttaaatgac agaatacatt
     7201 cccaatatac caaacataac tgtttcctac tagtcggccg tacgggccct ttcgtctcgc
     7261 gcgtttcggt gatgacggtg aaaacctctg acacatgcag ctcccggaga cggtcacagc
     7321 ttgtctgtaa gcggatgccg ggagcagaca agcccgtcag ggcgcgtcag cgggtgttgg
     7381 cgggtgtcgg ggctggctta actatgcggc atcagagcag attgtactga gagtgcacca
     7441 tatgcggtgt gaaataccgc acagatgcgt aaggagaaaa taccgcatca ggcggcctta
     7501 agggcctcgt gatacgccta tttttatagg ttaatgtcat gataataatg gtttcttaga
     7561 cgtcaggtgg cacttttcgg ggaaatgtgc gcggaacccc tatttgttta tttttctaaa
     7621 tacattcaaa tatgtatccg ctcatgagac aataaccctg ataaatgctt caataatatt
     7681 gaaaaaggaa gagtatgagt attcaacatt tccgtgtcgc ccttattccc ttttttgcgg
     7741 cattttgcct tcctgttttt gctcacccag aaacgctggt gaaagtaaaa gatgctgaag
     7801 atcagttggg tgcacgagtg ggttacatcg aactggatct caacagcggt aagatccttg
     7861 agagttttcg ccccgaagaa cgttttccaa tgatgagcac ttttaaagtt ctgctatgtg
     7921 gcgcggtatt atcccgtatt gacgccgggc aagagcaact cggtcgccgc atacactatt
     7981 ctcagaatga cttggttgag tactcaccag tcacagaaaa gcatcttacg gatggcatga
     8041 cagtaagaga attatgcagt gctgccataa ccatgagtga taacactgcg gccaacttac
     8101 ttctgacaac gatcggagga ccgaaggagc taaccgcttt tttgcacaac atgggggatc
     8161 atgtaactcg ccttgatcgt tgggaaccgg agctgaatga agccatacca aacgacgagc
     8221 gtgacaccac gatgcctgta gcaatggcaa caacgttgcg caaactatta actggcgaac
     8281 tacttactct agcttcccgg caacaattaa tagactggat ggaggcggat aaagttgcag
     8341 gaccacttct gcgctcggcc cttccggctg gctggtttat tgctgataaa tctggagccg
     8401 gtgagcgtgg gtctcgcggt atcattgcag cactggggcc agatggtaag ccctcccgta
     8461 tcgtagttat ctacacgacg gggagtcagg caactatgga tgaacgaaat agacagatcg
     8521 ctgagatagg tgcctcactg attaagcatt ggtaactgtc agaccaagtt tactcatata
     8581 tactttagat tgatttaaaa cttcattttt aatttaaaag gatctaggtg aagatccttt
     8641 ttgataatct catgaccaaa atcccttaac gtgagttttc gttccactga gcgtcagacc
     8701 ccgtagaaaa gatcaaagga tcttc
//
