LOCUS       sequence-333551-        6972 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  piggyBac vector for expression of reverse tetracycline 
            transactivator from a CAG constitutive promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6972)
  AUTHORS   Woltjen K, Michael IP, Mohseni P, Desai R, Mileikovsky M, Hamalainen
            R, Cowling R, Wang W, Liu P, Gertsenstein M, Kaji K, Sung HK, Nagy A
  TITLE     piggyBac transposition reprograms fibroblasts to induced pluripotent
            stem cells.
  JOURNAL   Nature. 2009 Apr 9;458(7239):766-70. doi: 10.1038/nature07863
  PUBMED    19252478
REFERENCE   2  (bases 1 to 6972)
  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"; doi: "10"; journalName: 
            "Nature"; date: "2009-04-9- 9"; volume: "458"; issue: "7239"; pages:
            "766-70"
COMMENT     Sequence Label: PB-CA-rtTA Adv
FEATURES             Location/Qualifiers
     source          1..6972
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        27..131
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(132..200,201..992)
                     /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(350..369)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      1163..1751
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1652..1671
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     1905..1922
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    2039..2060
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(2075..2080,2081..2098,2099..2105)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2113..2129
                     /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     2118..2140
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     2137..2153
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2137..2153
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     promoter        2174..2192
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     repeat_region   2341..2375
                     /label=piggyBac left (5') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     enhancer        2754..3133
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     intron          3373..4389
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     4313..4335
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     4397..4416
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     protein_bind    4483..4507
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             join(4513..5130,5131..5136,5137..5172,5173..5175,5176..5211,
                     5212..5214,5215..5250,5251..5259)
                     /codon_start=1
                     /product="improved tetracycline-controlled transactivator"
                     /label=rtTA-Advanced
                     /note="In the Tet-On(R) system, rtTA-Advanced binds to the 
                     Tet-responsive element and stimulates transcription only in
                     the presence of tetracycline or doxycycline."
                     /translation="MSRLDKSKVINGALELLNGVGIEGLTTRKLAQKLGVEQPTLYWHV
                     KNKRALLDALPIEMLDRHHTHFCPLEGESWQDFLRNNAKSFRCALLSHRDGAKVHLGTR
                     PTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEEQEHQVAKEERETPTT
                     DSMPPLLRQAIELFDRQGAEPAFLFGLELIICGLEKQLKCESGGPADALDDFDLDMLPA
                     DALDDFDLDMLPADALDDFDLDMLPG"
     protein_bind    complement(5262..5286)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     primer_bind     complement(5382..5401)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    5447..5502
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(5501..5520)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     repeat_region   6206..6268
                     /label=piggyBac right (3') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     primer_bind     complement(6334..6353)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(6335..6353)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(6360..6377)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(6360..6376)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6369..6391)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      6518..6946
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6605..6624)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     6815..6836
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
ORIGIN
        1 taggtggcac ttttcgggga aatgtgcgcg gaacccctat ttgtttattt ttctaaatac
       61 attcaaatat gtatccgctc atgagacaat aaccctgata aatgcttcaa taatattgaa
      121 aaaggaagag tatgagtatt caacatttcc gtgtcgccct tattcccttt tttgcggcat
      181 tttgccttcc tgtttttgct cacccagaaa cgctggtgaa agtaaaagat gctgaagatc
      241 agttgggtgc acgagtgggt tacatcgaac tggatctcaa cagcggtaag atccttgaga
      301 gttttcgccc cgaagaacgt tttccaatga tgagcacttt taaagttctg ctatgtggcg
      361 cggtattatc ccgtattgac gccgggcaag agcaactcgg tcgccgcata cactattctc
      421 agaatgactt ggttgagtac tcaccagtca cagaaaagca tcttacggat ggcatgacag
      481 taagagaatt atgcagtgct gccataacca tgagtgataa cactgcggcc aacttacttc
      541 tgacaacgat cggaggaccg aaggagctaa ccgctttttt gcacaacatg ggggatcatg
      601 taactcgcct tgatcgttgg gaaccggagc tgaatgaagc cataccaaac gacgagcgtg
      661 acaccacgat gcctgtagca atggcaacaa cgttgcgcaa actattaact ggcgaactac
      721 ttactctagc ttcccggcaa caattaatag actggatgga ggcggataaa gttgcaggac
      781 cacttctgcg ctcggccctt ccggctggct ggtttattgc tgataaatct ggagccggtg
      841 agcgtgggtc tcgcggtatc attgcagcac tggggccaga tggtaagccc tcccgtatcg
      901 tagttatcta cacgacgggg agtcaggcaa ctatggatga acgaaataga cagatcgctg
      961 agataggtgc ctcactgatt aagcattggt aactgtcaga ccaagtttac tcatatatac
     1021 tttagattga tttaaaactt catttttaat ttaaaaggat ctaggtgaag atcctttttg
     1081 ataatctcat gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg tcagaccccg
     1141 tagaaaagat caaaggatct tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc
     1201 aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc
     1261 tttttccgaa ggtaactggc ttcagcagag cgcagatacc aaatactgtt cttctagtgt
     1321 agccgtagtt aggccaccac ttcaagaact ctgtagcacc gcctacatac ctcgctctgc
     1381 taatcctgtt accagtggct gctgccagtg gcgataagtc gtgtcttacc gggttggact
     1441 caagacgata gttaccggat aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac
     1501 agcccagctt ggagcgaacg acctacaccg aactgagata cctacagcgt gagctatgag
     1561 aaagcgccac gcttcccgaa gggagaaagg cggacaggta tccggtaagc ggcagggtcg
     1621 gaacaggaga gcgcacgagg gagcttccag ggggaaacgc ctggtatctt tatagtcctg
     1681 tcgggtttcg ccacctctga cttgagcgtc gatttttgtg atgctcgtca ggggggcgga
     1741 gcctatggaa aaacgccagc aacgcggcct ttttacggtt cctggccttt tgctggcctt
     1801 ttgctcacat gttctttcct gcgttatccc ctgattctgt ggataaccgt attaccgcct
     1861 ttgagtgagc tgataccgct cgccgcagcc gaacgaccga gcgcagcgag tcagtgagcg
     1921 aggaagcgga agagcgccca atacgcaaac cgcctctccc cgcgcgttgg ccgattcatt
     1981 aatgcagctg gcacgacagg tttcccgact ggaaagcggg cagtgagcgc aacgcaatta
     2041 atgtgagtta gctcactcat taggcacccc aggctttaca ctttatgctt ccggctcgta
     2101 tgttgtgtgg aattgtgagc ggataacaat ttcacacagg aaacagctat gaccatgatt
     2161 acgccaagct cgaaattaac cctcactaaa gggaacaaaa gctggtacct cgcgcgactt
     2221 ggtttgccat tctttagcgc gcgtcgcgtc acacagcttg gccacaatgt ggtttttgtc
     2281 aaacgaagat tctatgacgt gtttaaagtt taggtcgagt aaagcgcaaa tcttttttaa
     2341 ccctagaaag atagtctgcg taaaattgac gcatgcattc ttgaaatatt gctctctctt
     2401 tctaaatagc gcgaatccgt cgctgtgcat ttaggacatc tcagtcgccg cttggagctc
     2461 ccgtgaggcg tgcttgtcaa tgcggtaagt gtcactgatt ttgaactata acgaccgcgt
     2521 gagtcaaaat gacgcatgat tatcttttac gtgactttta agatttaact catacgataa
     2581 ttatattgtt atttcatgtt ctacttacgt gataacttat tatatatata ttttcttgtt
     2641 atagatatcg tgactaatat ataataaaat gggtagttct ttagacgatg agcatatcct
     2701 ctctgctctt ctgcaaagcg atgacgagct tgttggctag cggtggcggc ctcgacattg
     2761 attattgact agttattaat agtaatcaat tacggggtca ttagttcata gcccatatat
     2821 ggagttccgc gttacataac ttacggtaaa tggcccgcct ggctgaccgc ccaacgaccc
     2881 ccgcccattg acgtcaataa tgacgtatgt tcccatagta acgccaatag ggactttcca
     2941 ttgacgtcaa tgggtggagt atttacggta aactgcccac ttggcagtac atcaagtgta
     3001 tcatatgcca agtacgcccc ctattgacgt caatgacggt aaatggcccg cctggcatta
     3061 tgcccagtac atgaccttat gggactttcc tacttggcag tacatctacg tattagtcat
     3121 cgctattacc atggtcgagg tgagccccac gttctgcttc actctcccca tctccccccc
     3181 ctccccaccc ccaattttgt atttatttat tttttaatta ttttgtgcag cgatgggggc
     3241 gggggggggg ggggcggggc gaggcggaga ggtgcggcgg cagccaatca gagcggcgcg
     3301 ctccgaaagt ttccttttat ggcgaggcgg cggcggcggc ggccctataa aaagcgaagc
     3361 gcgcggcggg cgggagtcgc tgcgcgctgc cttcgccccg tgccccgctc cgccgccgcc
     3421 tcgcgccgcc cgccccggct ctgactgacc gcgttactcc cacaggtgag cgggcgggac
     3481 ggcccttctc ctccgggctg taattagcgc ttggtttaat gacggcttgt ttcttttctg
     3541 tggctgcgtg aaagccttga ggggctccgg gagggccctt tgtgcggggg gagcggctcg
     3601 gggggtgcgt gcgtgtgtgt gtgcgtgggg agcgccgcgt gcggctccgc gctgcccggc
     3661 ggctgtgagc gctgcgggcg cggcgcgggg ctttgtgcgc tccgcagtgt gcgcgagggg
     3721 agcgcggccg ggggcggtgc cccgcggtgc ggggggggct gcgaggggaa caaaggctgc
     3781 gtgcggggtg tgtgcgtggg ggggtgagca gggggtgtgg gcgcgtcggt cgggctgcaa
     3841 ccccccctgc acccccctcc ccgagttgct gagcacggcc cggcttcggg tgcggggctc
     3901 cgtacggggc gtggcgcggg gctcgccgtg ccgggcgggg ggtggcggca ggtgggggtg
     3961 ccgggcgggg cggggccgcc tcgggccggg gagggctcgg gggaggggcg cggcggcccc
     4021 cggagcgccg gcggctgtcg aggcgcggcg agccgcagcc attgcctttt atggtaatcg
     4081 tgcgagaggg cgcagggact tcctttgtcc caaatctgtg cggagccgaa atctgggagg
     4141 cgccgccgca ccccctctag cgggcgcggg gcgaagcggt gcggcgccgg caggaaggaa
     4201 atgggcgggg agggccttcg tgcgtcgccg cgccgccgtc cccttctccc tctccagcct
     4261 cggggctgtc cgcgggggga cggctgcctt cgggggggac ggggcagggc ggggttcggc
     4321 ttctggcgtg tgaccggcgg ctctagagcc tctgctaacc atgttcatgc cttcttcttt
     4381 ttcctacagc tcctgggcaa cgtgctggtt attgtgctgt ctcatcattt tggcaaagaa
     4441 ttcgcggccg cggtaccggc gcgccggatc ctgcaggtta tcacaagttt gtacaaaaaa
     4501 gcaggcttca ccatgtctag actggacaag agcaaagtca taaacggcgc tctggaatta
     4561 ctcaatggag tcggtatcga aggcctgacg acaaggaaac tcgctcaaaa gctgggagtt
     4621 gagcagccta ccctgtactg gcacgtgaag aacaagcggg ccctgctcga tgccctgcca
     4681 atcgagatgc tggacaggca tcatacccac ttctgccccc tggaaggcga gtcatggcaa
     4741 gactttctgc ggaacaacgc caagtcattc cgctgtgctc tcctctcaca tcgcgacggg
     4801 gctaaagtgc atctcggcac ccgcccaaca gagaaacagt acgaaaccct ggaaaatcag
     4861 ctcgcgttcc tgtgtcagca aggcttctcc ctggagaacg cactgtacgc tctgtccgcc
     4921 gtgggccact ttacactggg ctgcgtattg gaggaacagg agcatcaagt agcaaaagag
     4981 gaaagagaga cacctaccac cgattctatg cccccacttc tgagacaagc aattgagctg
     5041 ttcgaccggc agggagccga acctgccttc cttttcggcc tggaactaat catatgtggc
     5101 ctggagaaac agctaaagtg cgaaagcggc gggccggccg acgcccttga cgattttgac
     5161 ttagacatgc tcccagccga tgcccttgac gactttgacc ttgatatgct gcctgctgac
     5221 gctcttgacg attttgacct tgacatgctc cccgggtaac tacccagctt tcttgtacaa
     5281 agtggtgata actctagaga attcactcct caggtgcagg ctgcctatca gaaggtggtg
     5341 gctggtgtgg ccaatgccct ggctcacaaa taccactgag atctttttcc ctctgccaaa
     5401 aattatgggg acatcatgaa gccccttgag catctgactt ctggctaata aaggaaattt
     5461 attttcattg caatagtgtg ttggaatttt ttgtgtctct cactcggaag gacatatggg
     5521 agggcaaatc atttaaaaca tcagaatgag tatttggttt agagtttggc aacatatgcc
     5581 catatgctgg ctgccatgaa caaaggttgg ctataaagag gtcatcagta tatgaaacag
     5641 ccccctgctg tccattcctt attccataga aaagccttga cttgaggtta gatttttttt
     5701 atattttgtt ttgtgttatt tttttcttta acatccctaa aattttcctt acatgtttta
     5761 ctagccagat ttttcctcct ctcctgacta ctcccagtca tagctgtccc tcttctctta
     5821 tggagatccc tcgacctgca gcccaagctt ggatccctcg agttaattaa cgagagcata
     5881 atattgatat gtgccaaagt tgtttctgac tgactaataa gtataatttg tttctattat
     5941 gtataggtta agctaattac ttattttata atacaacatg actgttttta aagtacaaaa
     6001 taagtttatt tttgtaaaag agagaatgtt taaaagtttt gttactttat agaagaaatt
     6061 ttgagttttt gttttttttt aataaataaa taaacataaa taaattgttt gttgaattta
     6121 ttattagtat gtaagtgtaa atataataaa acttaatatc tattcaaatt aataaataaa
     6181 cctcgatata cagaccgata aaacacatgc gtcaatttta cgcatgatta tctttaacgt
     6241 acgtcacaat atgattatct ttctagggtt aaataatagt ttctaatttt tttattattc
     6301 agcctgctgt cgtgaatacc gagctccaat tcgccctata gtgagtcgta ttacaattca
     6361 ctggccgtcg ttttacaacg tcgtgactgg gaaaaccctg gcgttaccca acttaatcgc
     6421 cttgcagcac atcccccttt cgccagctgg cgtaatagcg aagaggcccg caccgatcgc
     6481 ccttcccaac agttgcgcag cctgaatggc gaatgggacg cgccctgtag cggcgcatta
     6541 agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag cgccctagcg
     6601 cccgctcctt tcgctttctt cccttccttt ctcgccacgt tcgccggctt tccccgtcaa
     6661 gctctaaatc gggggctccc tttagggttc cgatttagtg ctttacggca cctcgacccc
     6721 aaaaaacttg attagggtga tggttcacgt agtgggccat cgccctgata gacggttttt
     6781 cgccctttga cgttggagtc cacgttcttt aatagtggac tcttgttcca aactggaaca
     6841 acactcaacc ctatctcggt ctattctttt gatttataag ggattttgcc gatttcggcc
     6901 tattggttaa aaaatgagct gatttaacaa aaatttaacg cgaattttaa caaaatatta
     6961 acgcttacaa tt
//
