LOCUS       sequence-476961-        4278 bp    DNA     circular SYN 24-JUN-2025
DEFINITION  Plasmid expressing a doxycycline-inducible transcriptional 
            activator.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4278)
  AUTHORS   Peterman EL, Ploessl DS, Love KS, Sanabria V, Daniels RF, Johnstone 
            CP, Godavarti DR, Kabaria SR, Oakes CG, Pai AA, Galloway KE
  TITLE     High-resolution profiling reveals coupled transcriptional and 
            translational regulation of transgenes.
  JOURNAL   bioRxiv [Preprint]. 2024 Dec 3:2024.11.26.625483. doi: 
            10.1101/2024.11.26.625483.
  PUBMED    39651241
REFERENCE   2  (bases 1 to 4278)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Jun 24, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "bioRxiv 
            [Preprint]. 2024 Dec 3:2024.11.26.625483. doi: 
            10.1101/2024.11.26.625483."
COMMENT     Sequence Label: pKG3668_pGEEC578_pShip-EFS-rtTA-P2A-iRFP720-bGH
FEATURES             Location/Qualifiers
     source          1..4278
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        5..216
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     CDS             228..971
                     /codon_start=1
                     /product="modified rtTA protein that binds tightly to 
                     promoters containing the tet operator in the presence of 
                     doxycycline"
                     /label=Tet-On(R) 3G
                     /translation="MSRLDKSKVINSALELLNGVGIEGLTTRKLAQKLGVEQPTLYWHV
                     KNKRALLDALPIEMLDRHHTHSCPLEGESWQDFLRNNAKSYRCALLSHRDGAKVHLGTR
                     PTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEEQEHQVAKEERETPTT
                     DSMPPLLKQAIELFDRQGAEPAFLFGLELIICGLEKQLKCESGGPTDALDDFDLDMLPA
                     DALDDFDLDMLPADALDDFDLDMLPG"
     CDS             993..1049
                     /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             1050..2000
                     /codon_start=1
                     /product="near-infrared fluorescent protein with an 
                     emission peak at 720 nm, engineered from a bacterial 
                     phytochrome (Shcherbakova and Verkhusha, 2013)"
                     /label=iRFP720
                     /note="derived from Rhodopseudomonas palustris RpBphP2"
                     /translation="MAEGSVARQPDLLTCDDEPIHIPGAIQPHGLLLALAADMTIVAGS
                     DNLPELTGLAIGALIGRSAADVFDSETHNRLTIALAEPGAAVGAPITVGFTMRKDAGFI
                     GSWHRHDQLIFLELEPPQRDVAEPQAFFRRTNSAIRRLQAAETLESACAAAAQEVRKIT
                     GFDRVMIYRFASDFSGSVIAEDRCAEVESKLGLHYPASFIPAQARRLYTINPVRIIPDI
                     NYRPVPVTPDLNPVTGRPIDLSFAILRSVSPNHLEFMRNIGMHGTMSISILRGERLWGL
                     IVCHHRTPYYVDLDGRQACELVAQVLAWQIGVMEE"
     polyA_signal    2012..2236
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(2347..2366)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             2358..3167
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYGKP
                     DAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPGDAWLLTTAIPGKTA
                     FQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDASD
                     FDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGIAD
                     RYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     complement(2425..2444)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      3257..3845
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3746..3765
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     terminator      4010..4096
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      4188..4215
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     primer_bind     4223..4246
                     /label=pENTR-F
                     /note="pENTR vectors, forward primer"
ORIGIN
        1 tactgggcag agcgcacatc gcccacagtc cccgagaagt tggggggagg ggtcggcaat
       61 tgaaccggtg cctagagaag gtggcgcggg gtaaactggg aaagtgatgt cgtgtactgg
      121 ctccgccttt ttcccgaggg tgggggagaa ccgtatataa gtgcagtagt cgccgtgaac
      181 gttctttttc gcaacgggtt tgccgccaga acacagtaat ggccaccatg agccggctgg
      241 acaagagcaa agtgatcaac agcgccctgg aactgctgaa cggcgtgggc atcgagggcc
      301 tgaccacccg gaagctggcc cagaaactgg gcgtggaaca gcccaccctg tactggcacg
      361 tgaagaacaa gcgggccctg ctggacgccc tgcccatcga gatgctggac cggcaccaca
      421 cccacagctg ccctctggaa ggcgagagct ggcaggactt cctgcggaac aacgccaaga
      481 gctacagatg cgccctgctg agccaccggg acggcgccaa agtgcacctg ggcaccagac
      541 ccaccgagaa gcagtacgag acactggaaa accagctggc cttcctgtgc cagcagggct
      601 tcagcctgga aaacgccctg tacgccctga gcgccgtggg ccacttcacc ctgggctgcg
      661 tgctggaaga acaggaacac caggtcgcca aagaggaaag agagacaccc accaccgaca
      721 gcatgccccc cctgctgaag caggccatcg agctgttcga cagacagggc gccgagcccg
      781 ccttcctgtt cggcctggaa ctgatcatct gcggcctcga gaagcagctg aagtgcgaga
      841 gcggcggacc caccgacgcc ctggacgact tcgacctgga catgctgccc gccgatgccc
      901 tggatgattt tgatctggat atgctccctg ctgacgctct cgatgacttc gatctcgata
      961 tgctgccagg cgctagcggc cgcggaagcg gagctactaa cttcagcctg ctgaagcagg
     1021 ctggagacgt ggaggagaac cctggaccta tggcggaagg atccgtcgcc aggcagcctg
     1081 acctcttgac ctgcgacgat gagccgatcc atatccccgg tgccatccaa ccgcatggac
     1141 tgctgctcgc cctcgccgcc gacatgacga tcgttgccgg cagcgacaac cttcccgaac
     1201 tcaccggact ggcgatcggc gccctgatcg gccgctctgc ggccgatgtc ttcgactcgg
     1261 agacgcacaa ccgtctgacg atcgccttgg ccgagcccgg ggcggccgtc ggagcaccga
     1321 tcactgtcgg cttcacgatg cgaaaggacg caggcttcat cggctcctgg catcgccatg
     1381 atcagctcat cttcctcgag ctcgagcctc cccagcggga cgtcgccgag ccgcaggcgt
     1441 tcttccgccg caccaacagc gccatccgcc gcctgcaggc cgccgaaacc ttggaaagcg
     1501 cctgcgccgc cgcggcgcaa gaggtgcgga agattaccgg cttcgatcgg gtgatgatct
     1561 atcgcttcgc ctccgacttc agcgggtccg tgatcgcaga ggatcggtgc gccgaggtcg
     1621 agtcaaaact aggcctgcac tatcctgcct cattcatccc ggcgcaggcc cgtcggctct
     1681 ataccatcaa cccggtacgg atcattcccg atatcaatta tcggccggtg ccggtcaccc
     1741 cagacctcaa tccggtcacc gggcggccga ttgatcttag cttcgccatc ctgcgcagcg
     1801 tctcgcccaa ccatctggag ttcatgcgca acataggcat gcacggcacg atgtcgatct
     1861 cgattttgcg cggcgagcga ctgtggggat tgatcgtttg ccatcaccga acgccgtact
     1921 acgtcgatct cgatggccgc caagcctgcg agctagtcgc ccaggttctg gcctggcaga
     1981 tcggcgtgat ggaagagtaa caacgttgca cctgtgcctt ctagttgcca gccatctgtt
     2041 gtttgcccct cccccgtgcc ttccttgacc ctggaaggtg ccactcccac tgtcctttcc
     2101 taataaaatg aggaaattgc atcgcattgt ctgagtaggt gtcattctat tctggggggt
     2161 ggggtggggc aggacagcaa gggggaggat tgggaagaca atagcaggca tgctggggat
     2221 gcggtgggct ctatggcgct cgctgactgc aggtgcccgt gtctcaaaat ctctgatgtt
     2281 acattgcaca agataaaaat atatcatcat gaacaataaa actgtctgct tacataaaca
     2341 gtaatacaag gggtgttatg agccatattc aacgggaaac gtcgaggccg cgattaaatt
     2401 ccaacatgga tgctgattta tatgggtata aatgggctcg cgataatgtc gggcaatcag
     2461 gtgcgacaat ctatcgcttg tatgggaagc ccgatgcgcc agagttgttt ctgaaacatg
     2521 gcaaaggtag cgttgccaat gatgttacag atgagatggt cagactaaac tggctgacgg
     2581 aatttatgcc tcttccgacc atcaagcatt ttatccgtac tcctggtgat gcatggttac
     2641 tcaccactgc gatccccgga aaaacagcat tccaggtatt agaagaatat cctgattcag
     2701 gtgaaaatat tgttgatgcg ctggcagtgt tcctgcgccg gttgcattcg attcctgttt
     2761 gtaattgtcc ttttaacagc gatcgcgtat ttcgtctagc tcaggcgcaa tcacgaatga
     2821 ataacggttt ggttgatgcg agtgattttg atgacgagcg taatggctgg cctgttgaac
     2881 aagtctggaa agaaatgcat aaacttttgc cattctcacc ggattcagtc gtcactcatg
     2941 gtgatttctc acttgataac cttatttttg acgaggggaa attaataggt tgtattgatg
     3001 ttggacgagt cggaatcgca gaccgatacc aggatcttgc catcctatgg aactgcctcg
     3061 gtgagttttc tccttcatta cagaaacggc tttttcaaaa atatggtatt gataatcctg
     3121 atatgaataa attgcagttt catttgatgc tcgatgagtt tttctaatca gaattggtta
     3181 attggttgta acattattca gattgggccc cgttccactg agcgtcagac cccgtagaaa
     3241 agatcaaagg atcttcttga gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa
     3301 aaaaaccacc gctaccagcg gtggtttgtt tgccggatca agagctacca actctttttc
     3361 cgaaggtaac tggcttcagc agagcgcaga taccaaatac tgttcttcta gtgtagccgt
     3421 agttaggcca ccacttcaag aactctgtag caccgcctac atacctcgct ctgctaatcc
     3481 tgttaccagt ggctgctgcc agtggcgata agtcgtgtct taccgggttg gactcaagac
     3541 gatagttacc ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc acacagccca
     3601 gcttggagcg aacgacctac accgaactga gatacctaca gcgtgagcta tgagaaagcg
     3661 ccacgcttcc cgaagggaga aaggcggaca ggtatccggt aagcggcagg gtcggaacag
     3721 gagagcgcac gagggagctt ccagggggaa acgcctggta tctttatagt cctgtcgggt
     3781 ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg cggagcctat
     3841 ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc cttttgctgg ccttttgctc
     3901 acatgttctt tcctgcgtta tcccctgatt ctgtggataa ccgtattacc gctagcatgg
     3961 atctcgggga cgtctaacta ctaagcgaga gtagggaact gccaggcatc aaataaaacg
     4021 aaaggctcag tcggaagact gggcctttcg ttttatctgt tgtttgtcgg tgaacgctct
     4081 cctgagtagg acaaatccgc cgggagcgga tttgaacgtt gtgaagcaac ggcccggagg
     4141 gtggcgggca ggacgcccgc cataaactgc caggcatcaa actaagcaga aggccatcct
     4201 gacggatggc ctttttgcgt ttctacaaac tcttcctgtt agttagttac ttaagctcgg
     4261 gcccacctgc agtctact
//
