LOCUS       sequence-320310-        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      71..659
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     560..579
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             4536..4592
                     /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             5451..5504
                     /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(7355..7377)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     7477..7496
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(7574..7592)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        7660..7764
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(7765..7833,7834..8625)
                     /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(7983..8002)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc agaccccgta gaaaagatca
       61 aaggatcttc ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac
      121 caccgctacc agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg
      181 taactggctt cagcagagcg cagataccaa atactgtcct tctagtgtag ccgtagttag
      241 gccaccactt caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac
      301 cagtggctgc tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt
      361 taccggataa ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg
      421 agcgaacgac ctacaccgaa ctgagatacc tacagcgtga gcattgagaa agcgccacgc
      481 ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc
      541 gcacgaggga gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc
      601 acctctgact tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa
      661 acgccagcaa cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt
      721 tctttcctgc gttatcccct gattctgtgg ataaccgtat taccgcggcc gccaacaata
      781 taggatgctc aggagactat aattcaaaaa acccaaccca aattagcata agcttcccca
      841 aattggaaca gtgaaatact caatcaatca ctctggcggt tttctccaca aagaacacga
      901 aattggaaaa aggaaaagat tgatttgttt gaaggcgtga ggaaatcgaa aaaaaattgt
      961 ttcaaaacaa atgaaatatt ttaacgaata aacccaggaa tgccactttt tggtgcccgc
     1021 ctacaaaaaa gtcaactgtt cttaaactcc tacatagttg ccctctctaa cgaatgttgc
     1081 tttttttaaa ttttcattga atttgattat cgtctgtatg cgcctactct atcagtggaa
     1141 aacatgccca tttcccacaa atttactcgt ttggagctca aaatgttaaa tggggcatca
     1201 gggaattggc atctcacaga aagtggcgga tgagttatat gtaataataa tggtggctga
     1261 agtggaaacc gcaaaaaaag gatgtgtatt tgagattttg caacggaatt tgaatcagat
     1321 acacaaaatg agagaaattt atagattctt taaaatccac ctgaagtcgg caaatttttg
     1381 aaaggtctgt tcgttcagtt tttcataagt tttatgaatt ttgatttact actgaagttc
     1441 gagaaaagcc ctttaaccat accaataaga aattatgtta actttacaag aatattaaca
     1501 aaaaattatt aaaaaactgt gttcccctgg cattatgata tttccaccgt cagttctcta
     1561 ttaaaacttc aactctacac agtttcaact ctcaaatgag ctgtccatct tttgccttca
     1621 ggaaattatg ctaatctgct gatttcgcgc actttcacca cactatttcc aaatagaagg
     1681 aggggtggtg caacacacga gacactgcaa aaaggttcag tgaagattta tctacttgtg
     1741 agatgagagc aggaggggtg ggagaagtgt gatgattgac gggatgacaa gaacagtgat
     1801 gtggtgtcga aatggaagat gcgctaattt gtggagtttt atgaaatttg aataattaca
     1861 tttataacat ctactagtta ggcgtctgtg taggtattgc ctgcctgcct acaaggcata
     1921 tgtacgaggc aggttaaacc tcctagaaag ctacggtact ttcaaaccac ataacattta
     1981 tactggtgtc ttttcgcgtc ttcttcttga acatgtagac cctcaaaaag ctaagtttta
     2041 cactgtgtag tttgctttga aatctacaaa aattggaggc acgccttctg gactttgccg
     2101 acacgcctac tttctaccta ccaactgcct acttccacag tctagcgtga actccaatac
     2161 cattttaaaa attgttttat ctcaaatttt caatttttaa atccaatttc ccagttctaa
     2221 aaaaaaaatt ttttttttaa tttaaaaact ctcatccttt ttaatgctcg ttcccattcg
     2281 gcaagacgtg taccaaacac aaaacaaatt gactggtgat ccatctcgaa tttctcgttt
     2341 caaattagtc gttgctatat atattgtctt atgtgtggaa aacgaataag aaaaattaga
     2401 gaaagatgag agagcagggg cagaatgaaa atttgcatgg aaattgaaga agatcactgc
     2461 tttttccgat gaatgatgac taaatagaat gagttgagag gaaaataatc ataattctaa
     2521 ttttaattag aaatatgata tggaaagtaa gcaaaaccac aaggcttttg ctctgaactt
     2581 atttccgaaa aaaaaagtga aactgaaagt tagaattttc aaaataaata ttgcgagagt
     2641 caaccttcag aaaaattggc gagaaatcaa atttgaattc tctgaaaatt tagttaaaat
     2701 acatatgtta tccttctttg ttttttggcc ttagcctgcc ttcaaggcga cctgagccta
     2761 tgcgggtgcc taataccgaa acacacataa tttttttttt taatctttaa atactgtaac
     2821 gagtcgcgag atccagcgag acccatagaa tatgttgagc gctgcaaaca atcgtataac
     2881 tcttgtccaa atttttccac tctgtatttt tgcaagcagt tcaacaaatg gtcatcattt
     2941 ttataaccca tatctatatt cgaatctgac ataacaaata atactaagca catcctccca
     3001 catctctttt cattcaagtt gcgcaattcc cctaataact atggatggag gggggaccga
     3061 gcccaaagat tattcaaaac gataacgact ttttcatgcg gccaacgaaa aataaaactt
     3121 ttctcatatt ctcctatttc ccatttcaag aattacatcc ccatgggacg ttgaaatcac
     3181 atcatattag tatatgttaa ttgttctttt cactggaata tcgccacgcc ccccttccgg
     3241 tggctcattg gtttcgagaa gaaggcggag ccgaagtggg ttcttctttc ttttttttgt
     3301 ggtagaagtc tagcaggatt aagctactat tccattagag gatcttcttc ttccaaaaag
     3361 tgaaaatgtc acgactggat aaatcgaagg tcatcaactc tgctctggaa ctgctgaatg
     3421 gagtgggaat tgaaggactt acaacccgaa agcttgcaca aaaattggga gttgaacaac
     3481 caacacttta ctggcatgtt aagaataagc gagcactttt ggatgctctt ccaattgaaa
     3541 tgttggatag acatcatact cattcatgtc cacttgaagg agaatcttgg caagatttct
     3601 tgagaaacaa cgcaaagtca taccgatgcg ctcttttgtc tcatagagat ggagcaaaag
     3661 ttcatcttgg aacacgacca actgaaaagc aatacgaaac attggaaaac caacttgctt
     3721 tcttgtgtca acaaggattt tcattggaaa atgctcttta cgcattgtct gctgttggac
     3781 attttacact tggatgcgtt ttggaagaac aagaacatca agttgcaaaa gaagaacgag
     3841 aaactccaac aactgattca atgccaccac ttttgaagca agctatcgaa cttttcgata
     3901 gacaaggagc agaaccagct ttcctttttg gactggaact tattatctgt ggacttgaga
     3961 aacaactgaa atgtgaatcg ggaggaggat ctggatctgg atctggatct ggttccggtc
     4021 gacaacttga gcttccgccg actgcgactg ccactgcaag tatcatgccc catgtcatgg
     4081 aacagcccct gtcaacgtca ataaatccgg tcaatgatcg gtttaatgga ataccgaatc
     4141 ctacccccta caatagcgat gcggccctag acgcaattac tcaaaccaat gactacggat
     4201 cagtcaatac ccatggcatt ctcagtacat atccaccacc tgccacccac ttgaacgagg
     4261 cctcggtggc ccttgcgcct ggaggtgctc ctcccagacc acccccgcca tacgttgata
     4321 gtacaacgaa ccatcctcct taccatagta acctcgtccc catggccaat ttcggatatt
     4381 cgactgtaga ctatgatgcg atggtggatg atctggcatc gatcgagtac acggatgcgg
     4441 ttgacgttga tccgcagttc atgacgaacc tcgggtttgt cccagggtgt aattttagcg
     4501 atatcaacac gtatgagcaa ccatggggca gtggggctac taacttcagc ctgctgaagc
     4561 aggctggaga cgtggaggag aaccctggac ctatggtctc cgagctcatt aaagaaaaca
     4621 tgcatatgaa gctctacatg gagggaaccg tcaacaacca ccacttcaag tgcacctccg
     4681 agggagaggg aaagccatac gagggaaccc aaaccatgcg tatcaaggcc gtcgagggag
     4741 gaccactccc attcgccttc gacatcctcg ccacctcctt catgtacgga tccaaggtaa
     4801 gtttaaacat atatatacta actaaccctg attatttaaa ttttcagacc ttcatcaacc
     4861 acacccaagg aatcccagac ttcttcaagc aatccttccc agagggattc acctgggagc
     4921 gtgtcaccac ctacgaggac ggaggagtcc tcaccgccac ccaagacacc tccctccaag
     4981 acggatgcct catctacaac gtcaaggtaa gtttaaacag ttcggtacta actaaccata
     5041 catatttaaa ttttcagatc cgtggagtca acttcccatc caacggacca gtcatgcaaa
     5101 agaagaccct cggatgggag gcctccaccg agaccctcta cccagccgac ggaggacttg
     5161 agggacgtgc cgacatggcc ctcaagctcg tcggaggagg acacctcatc tgcaacctca
     5221 aggtaagttt aaacatgatt ttactaacta actaatctga tttaaatttt cagaccacct
     5281 accgttccaa gaagccagcc aagaacctca agatgccagg agtctactac gtcgaccgtc
     5341 gtcttgagcg tatcaaggag gccgacaagg agacctacgt cgagcaacac gaggtcgccg
     5401 tcgcccgtta ctgcgacctc ccatccaagc tcggacaccg tggatcggga gagggacgag
     5461 gaagtctttt gacctgcgga gatgttgagg agaacccagg accaatgtcg cgtcttgata
     5521 aatcgaaagt tattaattca gctcttgaac ttttgaacga agttggaatt gaaggattga
     5581 caactagaaa gcttgcacaa aagttgggag ttgaacaacc aacactttat tggcatgttc
     5641 gaaataagca aactcttatg aacatgttgt ctgaagctat tcttgcaaaa catcatacaa
     5701 gatcagctcc attgccaact gaatcttggc aacaattcct tcaagaaaat gctttgtcat
     5761 ttcgaaaagc acttttggtt catagagatg gagcacgatt gcatattgga acatctccaa
     5821 ctccaccaca attcgaacaa gctgaagcac aacttagatg tttgtgcgat gctggattct
     5881 cagttgaaga agcacttttc atcttgcaat caatctctca tttcacactt ggagctgttt
     5941 tggaagaaca agcaactaac caaatcgaaa acaaccatgt tattgatgct gcaccaccac
     6001 ttttgcaaga agctttcaat attcaagcac gaacatctgc tgaaatggca ttccatttcg
     6061 gacttaagtc attgatcttc ggattttctg ctcaacttga tgaaaagaaa catactccaa
     6121 ttgaagatgg aaacaagcca aagaagaaaa gaaaactggg aagtggaagt aatggaagta
     6181 gtggatccag atctgcgcca ccttcttccg attgtccgcc gccgattgcc tactaccatc
     6241 atcatccaca acatcagcaa caattcctgc catttccaat gccatatttc ttggctccac
     6301 cgccgcaagc tcaacaagga gctccttttc cagtgcaata tattccacag caacatgatt
     6361 tgatgaatag ccagccaatg tatgcaccaa tggcaccgac atactactat caaccaatta
     6421 attcgaatgg catgcccatg aattccaact gagcgccggt cgctaccatt accaacttgt
     6481 ctggtgtcaa aaataatagg ggccgctgtc atcagagtaa gtttaaactg agttctacta
     6541 actaacgagt aatatttaaa ttttcagcat ctcgcgcccg tgcctctgac ttctaagtcc
     6601 aattactctt caacatccct acatgctctt tctccctgtg ctcccacccc ctatttttgt
     6661 tattatcaaa aaacttctct taatttcttt gttttttagc ttcttttaag tcacctctaa
     6721 caatgaaatt gtgtagattc aaaaatagaa ttaattcgta ataaaaagtc gaaaaaaatt
     6781 gtgctccctc cccccattaa taataattct atcccaaaat ctacacaatg ttctgtgtac
     6841 acttcttatg ttttttactt ctgataaatt tttttgaaac atcatagaaa aaaccgcaca
     6901 caaaatacct tatcatatgt tacgtttcag tttatgaccg caatttttat ttcttcgcac
     6961 gtctgggcct ctcatgacgt caaatcatgc tcatcgtgaa aaagttttgg agtatttttg
     7021 gaatttttca atcaagtgaa agtttatgaa attaattttc ctgcttttgc tttttggggt
     7081 ttcccctatt gtttgtcaag atttcgagga cggcgttttt cttgctaaaa tcacaagtat
     7141 tgatgagcac gatgcaagaa agatcggaag aaggtttggg tttgaggctc agtggaaggt
     7201 gagtagaagt tgataatttg aaagtggagt agtgtctatg gggtttttgc cttaaatgac
     7261 agaatacatt cccaatatac caaacataac tgtttcctac tagtcggccg tacgggccct
     7321 ttcgtctcgc gcgtttcggt gatgacggtg aaaacctctg acacatgcag ctcccggaga
     7381 cggtcacagc ttgtctgtaa gcggatgccg ggagcagaca agcccgtcag ggcgcgtcag
     7441 cgggtgttgg cgggtgtcgg ggctggctta actatgcggc atcagagcag attgtactga
     7501 gagtgcacca tatgcggtgt gaaataccgc acagatgcgt aaggagaaaa taccgcatca
     7561 ggcggcctta agggcctcgt gatacgccta tttttatagg ttaatgtcat gataataatg
     7621 gtttcttaga cgtcaggtgg cacttttcgg ggaaatgtgc gcggaacccc tatttgttta
     7681 tttttctaaa tacattcaaa tatgtatccg ctcatgagac aataaccctg ataaatgctt
     7741 caataatatt gaaaaaggaa gagtatgagt attcaacatt tccgtgtcgc ccttattccc
     7801 ttttttgcgg cattttgcct tcctgttttt gctcacccag aaacgctggt gaaagtaaaa
     7861 gatgctgaag atcagttggg tgcacgagtg ggttacatcg aactggatct caacagcggt
     7921 aagatccttg agagttttcg ccccgaagaa cgttttccaa tgatgagcac ttttaaagtt
     7981 ctgctatgtg gcgcggtatt atcccgtatt gacgccgggc aagagcaact cggtcgccgc
     8041 atacactatt ctcagaatga cttggttgag tactcaccag tcacagaaaa gcatcttacg
     8101 gatggcatga cagtaagaga attatgcagt gctgccataa ccatgagtga taacactgcg
     8161 gccaacttac ttctgacaac gatcggagga ccgaaggagc taaccgcttt tttgcacaac
     8221 atgggggatc atgtaactcg ccttgatcgt tgggaaccgg agctgaatga agccatacca
     8281 aacgacgagc gtgacaccac gatgcctgta gcaatggcaa caacgttgcg caaactatta
     8341 actggcgaac tacttactct agcttcccgg caacaattaa tagactggat ggaggcggat
     8401 aaagttgcag gaccacttct gcgctcggcc cttccggctg gctggtttat tgctgataaa
     8461 tctggagccg gtgagcgtgg gtctcgcggt atcattgcag cactggggcc agatggtaag
     8521 ccctcccgta tcgtagttat ctacacgacg gggagtcagg caactatgga tgaacgaaat
     8581 agacagatcg ctgagatagg tgcctcactg attaagcatt ggtaactgtc agaccaagtt
     8641 tactcatata tactttagat tgatttaaaa cttcattttt aatttaaaag gatctaggtg
     8701 aagatccttt ttgataatct catga
//
