LOCUS       sequence-358575-        7177 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  3xTRE Tet-On cassette with 3xp3 DsRed maker for generating knockins 
            with doxycycline-inducible rescue.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7177)
  AUTHORS   Chen J, Luo J, Gurav AS, Chen Z, Wang Y, Montell C
  TITLE     A DREaMR system to simplify combining mutations with rescue 
            transgenes in Aedes aegypti.
  JOURNAL   Genetics. 2021 Nov 5;219(3). pii: 6368066. doi: 
            10.1093/genetics/iyab146.
  PUBMED    34740249
REFERENCE   2  (bases 1 to 7177)
  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: "Genetics.";
            date: "2021-11-5"; volume: "219(3). pii: 6368066. doi"; pages: " 
            10.1093/genetics/iyab146"
COMMENT     Sequence Label: 3xTRE DREaMR 3p3DsR
FEATURES             Location/Qualifiers
     source          1..7177
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        24..128
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(129..197,198..989)
                     /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(347..366)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      1160..1748
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1649..1668
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     1902..1919
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    2036..2057
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(2072..2077,2078..2095,2096..2102)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2110..2126
                     /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)."
     primer_bind     2115..2137
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     2134..2150
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2134..2150
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     repeat_region   2641..2675
                     /label=piggyBac left (5') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     polyA_signal    3726..3860
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     regulatory      4121..4130
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             4127..4804
                     /codon_start=1
                     /product="rapidly maturing tetrameric variant of DsRed 
                     fluorescent protein (Bevis and Glick, 2002)"
                     /label=DsRed-Express
                     /note="mammalian codon-optimized"
                     /translation="MASSEDVIKEFMRFKVRMEGSVNGHEFEIEGEGEGRPYEGTQTAK
                     LKVTKGGPLPFAWDILSPQFQYGSKVYVKHPADIPDYKKLSFPEGFKWERVMNFEDGGV
                     VTVTQDSSLQDGSFIYKVKFIGVNFPSDGPVMQKKTMGWEASTERLYPRDGVLKGEIHK
                     ALKLKDGGHYLVEFKSIYMAKKPVQLPGYYYVDSKLDITSHNEDYTIVEQYERAEGRHH
                     LFL"
     primer_bind     complement(4257..4275)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     complement(4307..4327)
                     /label=DsRed1-N
                     /note="DsRed1, reverse primer"
     primer_bind     4525..4544
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     4719..4742
                     /label=DsRed1-C
                     /note="DsRed1, forward primer"
     polyA_signal    4925..5046
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     protein_bind    5085..5103
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    5121..5139
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    5156..5174
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    5192..5210
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    5228..5246
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    5263..5281
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    5299..5317
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    5353..5371
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    5389..5407
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    5424..5442
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    5460..5478
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    5496..5514
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    5531..5549
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    5567..5585
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    5622..5640
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    5658..5676
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    5693..5711
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    5729..5747
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    5765..5783
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    5800..5818
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    5836..5854
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     CDS             6039..6128
                     /codon_start=1
                     /product="three tandem HA epitope tags"
                     /label=3xHA
                     /translation="YPYDVPDYAGYPYDVPDYAGSYPYDVPDYA"
     repeat_region   6370..6432
                     /label=piggyBac right (3') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     primer_bind     complement(6562..6579)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(6562..6578)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6571..6593)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      6720..7175
                     /direction=RIGHT
                     /label=M13 ori
                     /note="M13 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6807..6826)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     7017..7038
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
ORIGIN
        1 gtggcacttt tcggggaaat gtgcgcggaa cccctatttg tttatttttc taaatacatt
       61 caaatatgta tccgctcatg agacaataac cctgataaat gcttcaataa tattgaaaaa
      121 ggaagagtat gagtattcaa catttccgtg tcgcccttat tccctttttt gcggcatttt
      181 gccttcctgt ttttgctcac ccagaaacgc tggtgaaagt aaaagatgct gaagatcagt
      241 tgggtgcacg agtgggttac atcgaactgg atctcaacag cggtaagatc cttgagagtt
      301 ttcgccccga agaacgtttt ccaatgatga gcacttttaa agttctgcta tgtggcgcgg
      361 tattatcccg tattgacgcc gggcaagagc aactcggtcg ccgcatacac tattctcaga
      421 atgacttggt tgagtactca ccagtcacag aaaagcatct tacggatggc atgacagtaa
      481 gagaattatg cagtgctgcc ataaccatga gtgataacac tgcggccaac ttacttctga
      541 caacgatcgg aggaccgaag gagctaaccg cttttttgca caacatgggg gatcatgtaa
      601 ctcgccttga tcgttgggaa ccggagctga atgaagccat accaaacgac gagcgtgaca
      661 ccacgatgcc tgtagcaatg gcaacaacgt tgcgcaaact attaactggc gaactactta
      721 ctctagcttc ccggcaacaa ttaatagact ggatggaggc ggataaagtt gcaggaccac
      781 ttctgcgctc ggcccttccg gctggctggt ttattgctga taaatctgga gccggtgagc
      841 gtgggtctcg cggtatcatt gcagcactgg ggccagatgg taagccctcc cgtatcgtag
      901 ttatctacac gacggggagt caggcaacta tggatgaacg aaatagacag atcgctgaga
      961 taggtgcctc actgattaag cattggtaac tgtcagacca agtttactca tatatacttt
     1021 agattgattt aaaacttcat ttttaattta aaaggatcta ggtgaagatc ctttttgata
     1081 atctcatgac caaaatccct taacgtgagt tttcgttcca ctgagcgtca gaccccgtag
     1141 aaaagatcaa aggatcttct tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa
     1201 caaaaaaacc accgctacca gcggtggttt gtttgccgga tcaagagcta ccaactcttt
     1261 ttccgaaggt aactggcttc agcagagcgc agataccaaa tactgttctt ctagtgtagc
     1321 cgtagttagg ccaccacttc aagaactctg tagcaccgcc tacatacctc gctctgctaa
     1381 tcctgttacc agtggctgct gccagtggcg ataagtcgtg tcttaccggg ttggactcaa
     1441 gacgatagtt accggataag gcgcagcggt cgggctgaac ggggggttcg tgcacacagc
     1501 ccagcttgga gcgaacgacc tacaccgaac tgagatacct acagcgtgag ctatgagaaa
     1561 gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa
     1621 caggagagcg cacgagggag cttccagggg gaaacgcctg gtatctttat agtcctgtcg
     1681 ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc
     1741 tatggaaaaa cgccagcaac gcggcctttt tacggttcct ggccttttgc tggccttttg
     1801 ctcacatgtt ctttcctgcg ttatcccctg attctgtgga taaccgtatt accgcctttg
     1861 agtgagctga taccgctcgc cgcagccgaa cgaccgagcg cagcgagtca gtgagcgagg
     1921 aagcggaaga gcgcccaata cgcaaaccgc ctctccccgc gcgttggccg attcattaat
     1981 gcagctggca cgacaggttt cccgactgga aagcgggcag tgagcgcaac gcaattaatg
     2041 tgagttagct cactcattag gcaccccagg ctttacactt tatgcttccg gctcgtatgt
     2101 tgtgtggaat tgtgagcgga taacaatttc acacaggaaa cagctatgac catgattacg
     2161 ccaagcgcgc ccgccgggta actcacgggg tatccatgtc catttctgcg gcatccagcc
     2221 aggatacccg tcctcgctga cgtaatatcc cagcgccgca ccgctgtcat taatctgcac
     2281 accggcacgg cagttccggc tgtcgccggt attgttcggg ttgctgatgc gcttcgggct
     2341 gaccatccgg aactgtgtcc ggaaaagccg cgacgaactg gtatcccagg tggcctgaac
     2401 gaacagttca ccgttaaagg cgtgcatggc cacaccttcc cgaatcatca tggtaaacgt
     2461 gcgttttcgc tcaacgtcaa tgcagcagca gtcatcctcg gcaaactctt tccatgccgc
     2521 ttcaacctcg cgggaaaagg cacgggcttc ttcctccccg atgcccagat agcgccagct
     2581 tgggcgatga ctgagccgga aaaaagaccc gacgatatga tcctgatgca gctagattaa
     2641 ccctagaaag atagtctgcg taaaattgac gcatgcattc ttgaaatatt gctctctctt
     2701 tctaaatagc gcgaatccgt cgctgtgcat ttaggacatc tcagtcgccg cttggagctc
     2761 ccgtgaggcg tgcttgtcaa tgcggtaagt gtcactgatt ttgaactata acgaccgcgt
     2821 gagtcaaaat gacgcatgat tatcttttac gtgactttta agatttaact catacgataa
     2881 ttatattgtt atttcatgtt ctacttacgt gataacttat tatatatata ttttcttgtt
     2941 atagatatca agctacgtag ctagccacca tgagtaggtt ggacaagtcc aaggttatca
     3001 acggcgcttt ggagttgttg aacggtgttg gcattgaggg tctgaccact cgcaaactgg
     3061 cccaaaagct cggagtagaa cagcccacct tgtattggca tgtgaaaaac aagagggctt
     3121 tgttggacgc tctgccgata gagatgttgg accgtcatca cactcatttt tgccccctgg
     3181 aaggagagtc ctggcaggat ttcctccgta acaatgccaa aagttttcgg tgcgcactcc
     3241 tgagtcatcg agacggcgct aaagtacacc tgggaacccg gccaactgag aagcagtatg
     3301 agacactgga gaaccaactg gcattcctgt gtcaacaagg cttcagtttg gaaaatgccc
     3361 tgtacgcctt gagcgcggtt ggacacttta cactgggttg cgttttggag gaacaggaac
     3421 atcaggtcgc gaaggaggaa agggagacac cgaccacaga ttcgatgccg cctctgctcc
     3481 gtcaagcaat tgagctcttt gaccgacaag gagcagaacc tgcctttctc tttggtctcg
     3541 aactcatcat ttgtggattg gaaaagcagc tcaaatgtga gtcgggagga cctgccgacg
     3601 cgctggatga ctttgatttg gacatgctcc cggcagacgc cctggatgat ttcgatctcg
     3661 atatgctgcc cgcagacgct ttggatgact ttgacctgga tatgttgccg ggataactaa
     3721 gtaaggatcc agacatgata agatacattg atgagtttgg acaaaccaca actagaatgc
     3781 agtgaaaaaa atgctttatt tgtgaaattt gtgatgctat tgctttattt gtaaccatta
     3841 taagctgcaa taaacaagtt gcggccgctc tagaactagt gccctttcgt cttcaagaat
     3901 tggatctaat tcaattagag actaattcaa ttagagctaa ttcaattagg atccaagctt
     3961 atcgatttcg aaccctcgac cgccggagta taaatagagg cgcttcgtct acggagcgac
     4021 aattcaattc aaacaagcaa agtgaacacg tcgctaagcg aaagctaagc aaataaacaa
     4081 gcgcagctga acaagctaaa caatcggctc gaagccggtc gccaccatgg cctcctccga
     4141 ggacgtcatc aaggagttca tgcgcttcaa ggtgcgcatg gagggctccg tgaacggcca
     4201 cgagttcgag atcgagggcg agggcgaggg ccgcccctac gagggcaccc agaccgccaa
     4261 gctgaaggtg accaagggcg gccccctgcc cttcgcctgg gacatcctgt ccccccagtt
     4321 ccagtacggc tccaaggtgt acgtgaagca ccccgccgac atccccgact acaagaagct
     4381 gtccttcccc gagggcttca agtgggagcg cgtgatgaac ttcgaggacg gcggcgtggt
     4441 gaccgtgacc caggactcct ccctccagga cggctccttc atctacaagg tgaagttcat
     4501 cggcgtgaac ttcccctccg acggccccgt aatgcagaag aagactatgg gctgggaggc
     4561 gtccaccgag cgcctgtacc cccgcgacgg cgtgctgaag ggcgagatcc acaaggccct
     4621 gaagctgaag gacggcggcc actacctggt ggagttcaag tccatctaca tggccaagaa
     4681 gcccgtgcag ctgcccggct actactacgt ggactccaag ctggacatca cctcccacaa
     4741 cgaggactac accatcgtgg agcagtacga gcgcgccgag ggccgccacc acctgttcct
     4801 gtaggggccg cgactctaga tcataatcag ccataccaca tttgtagagg ttttacttgc
     4861 tttaaaaaac ctcccacacc tccccctgaa cctgaaacat aaaatgaatg caattgttgt
     4921 tgttaacttg tttattgcag cttataatgg ttacaaataa agcaatagca tcacaaattt
     4981 cacaaataaa gcattttttt cactgcattc tagttgtggt ttgtccaaac tcatcaatgt
     5041 atcttaacgc gagttaagct tatcgatacc gtcgacgagt ttactcccta tcagtgatag
     5101 agaacgtatg tcgagtttac tccctatcag tgatagagaa cgatgtcgag tttactccct
     5161 atcagtgata gagaacgtat gtcgagttta ctccctatca gtgatagaga acgtatgtcg
     5221 agtttactcc ctatcagtga tagagaacgt atgtcgagtt tatccctatc agtgatagag
     5281 aacgtatgtc gagtttactc cctatcagtg atagagaacg tatgtcgagg agctcgccct
     5341 cgacgagttt actccctatc agtgatagag aacgtatgtc gagtttactc cctatcagtg
     5401 atagagaacg atgtcgagtt tactccctat cagtgataga gaacgtatgt cgagtttact
     5461 ccctatcagt gatagagaac gtatgtcgag tttactccct atcagtgata gagaacgtat
     5521 gtcgagttta tccctatcag tgatagagaa cgtatgtcga gtttactccc tatcagtgat
     5581 agagaacgta tgtcgaggag cctcgccctc gacgagttta ctccctatca gtgatagaga
     5641 acgtatgtcg agtttactcc ctatcagtga tagagaacga tgtcgagttt actccctatc
     5701 agtgatagag aacgtatgtc gagtttactc cctatcagtg atagagaacg tatgtcgagt
     5761 ttactcccta tcagtgatag agaacgtatg tcgagtttat ccctatcagt gatagagaac
     5821 gtatgtcgag tttactccct atcagtgata gagaacgtat gtcgaggagc tcgcccgggg
     5881 atcgagcgca gcggtataaa agggcgcggg gtggctgaga gcatcagttg tgaatgaatg
     5941 ttcgagccga gcagacgtgc cgctgccttc gttaatatcc tttgaataag ccaactttga
     6001 atcacaagac gcataccaaa cggtaccgcg gcaccatgta cccctacgac gtgcccgact
     6061 acgccggcta cccatacgac gtcccagatt acgctggttc gtacccgtac gacgtgccag
     6121 attacgcctc cggcggttcc ggtggcagcg gcggcgccgt taattaagag gatcccccgg
     6181 gctgcaggaa ttcgataaaa gttttgttac tttatagaag aaattttgag tttttgtttt
     6241 ttttaataaa taaataaaca taaataaatt gtttgttgaa tttattatta gtatgtaagt
     6301 gtaaatataa taaaacttaa tatctattca aattaataaa taaacctcga tatacagacc
     6361 gataaaacac atgcgtcaat tttacacatg attatcttta acgtacgtca caatatgatt
     6421 atctttctag ggttaatcta gctgcgtgtt ctgcagcgtg tcgagcatct tcatctgctc
     6481 catcacgctg taaaacacat ttgcaccgcg agtctgcccg tcctccacgg gttcaaaaac
     6541 gtgaatgaac gaggcgcgct cactggccgt cgttttacaa cgtcgtgact gggaaaaccc
     6601 tggcgttacc caacttaatc gccttgcagc acatccccct ttcgccagct ggcgtaatag
     6661 cgaagaggcc cgcaccgatc gcccttccca acagttgcgc agcctgaatg gcgaatggga
     6721 cgcgccctgt agcggcgcat taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc
     6781 tacacttgcc agcgccctag cgcccgctcc tttcgctttc ttcccttcct ttctcgccac
     6841 gttcgccggc tttccccgtc aagctctaaa tcgggggctc cctttagggt tccgatttag
     6901 tgctttacgg cacctcgacc ccaaaaaact tgattagggt gatggttcac gtagtgggcc
     6961 atcgccctga tagacggttt ttcgcccttt gacgttggag tccacgttct ttaatagtgg
     7021 actcttgttc caaactggaa caacactcaa ccctatctcg gtctattctt ttgatttata
     7081 agggattttg ccgatttcgg cctattggtt aaaaaatgag ctgatttaac aaaaatttaa
     7141 cgcgaatttt aacaaaatat taacgcttac aatttag
//
