LOCUS       sequence-362474-        7739 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  piggyBac co-transfection transposon containing M2rtTA promoter 
            linked with Puromycin resistance gene, through T2A with GSG linker, 
            subcloned between ends of two ITRs. Pairs with #169174.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7739)
  AUTHORS   Jung HS, Uenishi G, Park MA, Liu P, Suknuntha K, Raymond M, Choi YJ,
            Thomson JA, Ong IM, Slukvin II
  TITLE     SOX17 integrates HOXA and arterial programs in hemogenic endothelium
            to drive definitive lympho-myeloid hematopoiesis.
  JOURNAL   Cell Rep. 2021 Feb 16;34(7):108758. doi: 
            10.1016/j.celrep.2021.108758.
  PUBMED    33596423
REFERENCE   2  (bases 1 to 7739)
  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: "Cell Rep.";
            date: "2021-02-16"; volume: "34(7):108758. doi"; pages: " 
            10.1016/j.celrep.2021.108758"
COMMENT     Sequence Label: pEF1-alpha-M2rtTA-T2A-Puro
FEATURES             Location/Qualifiers
     source          1..7739
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        complement(1..105)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(131..586)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(268..289)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     480..499
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     713..735
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     727..744
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     728..744
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     repeat_region   874..936
                     /label=piggyBac right (3') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     primer_bind     complement(1132..1148)
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1132..1148)
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     primer_bind     complement(1477..1497)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        1540..2702
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          1771..2693
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     2650..2670
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     CDS             join(2758..3375,3376..3381,3382..3417,3418..3420,3421..3456,
                     3457..3459,3460..3495,3496..3501)
                     /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"
     CDS             3517..3570
                     /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"
     CDS             3577..4176
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTKLQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(3577..3596)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     4073..4093
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    4177..4298
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4188..4207)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     4242..4261
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     promoter        complement(4825..4843)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     4850..4869
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        4850..4868
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     4893..4909
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     4894..4910
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     repeat_region   5193..5227
                     /label=piggyBac left (5') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     primer_bind     complement(5718..5734)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5718..5734)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5731..5753)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5742..5758
                     /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)."
     promoter        complement(join(5766..5772,5773..5790,5791..5796))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5811..5832
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5949..5966)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6120..6708)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6200..6219)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6879..7670,7671..7739))
                     /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     7502..7521
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata
       61 catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa
      121 agtgccacct aaattgtaag cgttaatatt ttgttaaaat tcgcgttaaa tttttgttaa
      181 atcagctcat tttttaacca ataggccgaa atcggcaaaa tcccttataa atcaaaagaa
      241 tagaccgaga tagggttgag tgttgttcca gtttggaaca agagtccact attaaagaac
      301 gtggactcca acgtcaaagg gcgaaaaacc gtctatcagg gcgatggccc actacgtgaa
      361 ccatcaccct aatcaagttt tttggggtcg aggtgccgta aagcactaaa tcggaaccct
      421 aaagggagcc cccgatttag agcttgacgg ggaaagccgg cgaacgtggc gagaaaggaa
      481 gggaagaaag cgaaaggagc gggcgctagg gcgctggcaa gtgtagcggt cacgctgcgc
      541 gtaaccacca cacccgccgc gcttaatgcg ccgctacagg gcgcgtccca ttcgccattc
      601 aggctgcgca actgttggga agggcgatcg gtgcgggcct cttcgctatt acgccagctg
      661 gcgaaagggg gatgtgctgc aaggcgatta agttgggtaa cgccagggtt ttcccagtca
      721 cgacgttgta aaacgacggc cagtgagcgc gcctcgttca ttcacgtttt tgaacccgtg
      781 gaggacgggc agactcgcgg tgcaaatgtg ttttacagcg tgatggagca gatgaagatg
      841 ctcgacacgc tgcagaacac gcagctagat taaccctaga aagataatca tattgtgacg
      901 tacgttaaag ataatcatgt gtaaaattga cgcatgtgtt ttatcggtct gtatatcgag
      961 gtttatttat taatttgaat agatattaag ttttattata tttacactta catactaata
     1021 ataaattcaa caaacaattt atttatgttt atttatttat taaaaaaaac aaaaactcaa
     1081 aatttcttct ataaagtaac aaaactttta tcgaattcct gcagcccggg ggatccacta
     1141 gttctagagg gacagccccc ccccaaagcc cccagggatg taattacgtc cctcccccgc
     1201 tagggggcag cagcgagccg cccggggctc cgctccggtc cggcgctccc cccgcatccc
     1261 cgagccggca gcgtgcgggg acagcccggg cacggggaag gtggcacggg atcgctttcc
     1321 tctgaacgct tctcgctgct ctttgagcct gcagacacct ggggggatac ggggaaaagg
     1381 cctccaaggc cagcttccca caataagttg ggtgaatttt ggctcattcc tcctttctat
     1441 aggattgagg tcagagcttt gtgatgggaa ttctgtggaa tgtgtgtcag ttagggtgtg
     1501 gaaagtcccg cgatcgtcga gagatttcta gcttcgtgag gctccggtgc ccgtcagtgg
     1561 gcagagcgca catcgcccac agtccccgag aagttggggg gaggggtcgg caattgaacc
     1621 ggtgcctaga gaaggtggcg cggggtaaac tgggaaagtg atgtcgtgta ctggctccgc
     1681 ctttttcccg agggtggggg agaaccgtat ataagtgcag tagtcgccgt gaacgttctt
     1741 tttcgcaacg ggtttgccgc cagaacacag gtaagtgccg tgtgtggttc ccgcgggcct
     1801 ggcctcttta cgggttatgg cccttgcgtg ccttgaatta cttccacctg gctccagtac
     1861 gtgattcttg atcccgagct ggagccaggg gcgggccttg cgctttagga gccccttcgc
     1921 ctcgtgcttg agttgaggcc tggcctgggc gctggggccg ccgcgtgcga atctggtggc
     1981 accttcgcgc ctgtctcgct gctttcgata agtctctagc catttaaaat ttttgatgac
     2041 ctgctgcgac gctttttttc tggcaagata gtcttgtaaa tgcgggccag gatctgcaca
     2101 ctggtatttc ggtttttggg gccgcgggcg gcgacggggc ccgtgcgtcc cagcgcacat
     2161 gttcggcgag gcggggcctg cgagcgcggc caccgagaat cggacggggg tagtctcaag
     2221 ctggccggcc tgctctggtg cctggcctcg cgccgccgtg tatcgccccg ccctgggcgg
     2281 caaggctggc ccggtcggca ccagttgcgt gagcggaaag atggccgctt cccggccctg
     2341 ctccaggggg ctcaaaatgg aggacgcggc gctcgggaga gcgggcgggt gagtcaccca
     2401 cacaaaggaa aggggccttt ccgtcctcag ccgtcgcttc atgtgactcc acggagtacc
     2461 gggcgccgtc caggcacctc gattagttct ggagcttttg gagtacgtcg tctttaggtt
     2521 ggggggaggg gttttatgcg atggagtttc cccacactga gtgggtggag actgaagtta
     2581 ggccagcttg gcacttgatg taattctcct tggaatttgc cctttttgag tttggatctt
     2641 ggttcattct caagcctcag acagtggttc aaagtttttt tcttccattt caggtgtcgt
     2701 gaacacgtgg tcgcggccat ctagagcaac tagcttgggc ccgggaatta attcaccatg
     2761 tctagactgg acaagagcaa agtcataaac ggcgctctgg aattactcaa tggagtcggt
     2821 atcgaaggcc tgacgacaag gaaactcgct caaaagctgg gagttgagca gcctaccctg
     2881 tactggcacg tgaagaacaa gcgggccctg ctcgatgccc tgccaatcga gatgctggac
     2941 aggcatcata cccacttctg ccccctggaa ggcgagtcat ggcaagactt tctgcggaac
     3001 aacgccaagt cattccgctg tgctctcctc tcacatcgcg acggggctaa agtgcatctc
     3061 ggcacccgcc caacagagaa acagtacgaa accctggaaa atcagctcgc gttcctgtgt
     3121 cagcaaggct tctccctgga gaacgcactg tacgctctgt ccgccgtggg ccactttaca
     3181 ctgggctgcg tattggagga acaggagcat caagtagcaa aagaggaaag agagacacct
     3241 accaccgatt ctatgccccc acttctgaga caagcaattg agctgttcga ccggcaggga
     3301 gccgaacctg ccttcctttt cggcctggaa ctaatcatat gtggcctgga gaaacagcta
     3361 aagtgcgaaa gcggcgggcc ggccgacgcc cttgacgatt ttgacttaga catgctccca
     3421 gccgatgccc ttgacgactt tgaccttgat atgctgcctg ctgacgctct tgacgatttt
     3481 gaccttgaca tgctccccgg ggaattcgga agcggagagg gcagaggaag tcttctaaca
     3541 tgcggtgacg tggaggagaa tcccggccct gatatcatga ccgagtacaa gcccacggtg
     3601 cgcctcgcca cccgcgacga cgtcccccgg gccgtacgca ccctcgccgc cgcgttcgcc
     3661 gactaccccg ccacgcgcca caccgtcgac ccggaccgcc acatcgagcg ggtcaccaag
     3721 ctgcaagaac tcttcctcac gcgcgtcggg ctcgacatcg gcaaggtgtg ggtcgcggac
     3781 gacggcgccg cggtggcggt ctggaccacg ccggagagcg tcgaagcggg ggcggtgttc
     3841 gccgagatcg gcccgcgcat ggccgagttg agcggttccc ggctggccgc gcagcaacag
     3901 atggaaggcc tcctggcgcc gcaccggccc aaggagcccg cgtggttcct ggccaccgtc
     3961 ggcgtctcgc ccgaccacca gggcaagggt ctgggcagcg ccgtcgtgct ccccggagtg
     4021 gaggcggccg agcgcgccgg ggtgcccgcc ttcctggaga cctccgcgcc ccgcaacctc
     4081 cccttctacg agcggctcgg cttcaccgtc accgccgacg tcgaggtgcc cgaaggaccg
     4141 cgcacctggt gcatgacccg caagcccggt gcctgataag atacattgat gagtttggac
     4201 aaaccacaac tagaatgcag tgaaaaaaat gctttatttg tgaaatttgt gatgctattg
     4261 ctttatttgt aaccattata agctgcaata aacaagttaa caacaacaat tgcattcatt
     4321 ttatgcttca ggttcagggg gaggtgtggg aggtttttta aagcaagtaa aacctctaca
     4381 aatgtggtat ggctgattat gatccggctg cctcgcgcgg tacgacgcgt atacctagga
     4441 tccggccggc ctgcaggtgt cctcacagga acgaagtccc taaagaaaca gtggcagcca
     4501 ggtttagccc cggaattgac tggattcctt tttagggccc attggtatgg ctttttcccc
     4561 gtatcccccc aggtgtctgc aggctcaaag agcagcgaga agcgttcaga ggaaagcgat
     4621 cccgtgccac cttccccgtg cccgggctgt ccccgcacgc tgccggctcg gggatgcggg
     4681 gggagcgccg gaccggagcg gagccccggg cggctcgctg ctgcccccta gcgggggagg
     4741 gacgtaatta catccctggg ggctttgggg gggggctgtc cctctagagc ggccgccacc
     4801 gcggtggagc tccagctttt gttcccttta gtgagggtta attagatctt aatacgactc
     4861 actatagggc gaattgggta ccgggccccc cctcgaggtc gacggtatcg ataagcttga
     4921 tatctataac aagaaaatat atatataata agttatcacg taagtagaac atgaaataac
     4981 aatataatta tcgtatgagt taaatcttaa aagtcacgta aaagataatc atgcgtcatt
     5041 ttgactcacg cggtcgttat agttcaaaat cagtgacact taccgcattg acaagcacgc
     5101 ctcacgggag ctccaagcgg cgactgagat gtcctaaatg cacagcgacg gattcgcgct
     5161 atttagaaag agagagcaat atttcaagaa tgcatgcgtc aattttacgc agactatctt
     5221 tctagggtta atctagctgc atcaggatca tatcgtcggg tcttttttcc ggctcagtca
     5281 tcgcccaagc tggcgctatc tgggcatcgg ggaggaagaa gcccgtgcct tttcccgcga
     5341 ggttgaagcg gcatggaaag agtttgccga ggatgactgc tgctgcattg acgttgagcg
     5401 aaaacgcacg tttaccatga tgattcggga aggtgtggcc atgcacgcct ttaacggtga
     5461 actgttcgtt caggccacct gggataccag ttcgtcgcgg cttttccgga cacagttccg
     5521 gatggtcagc ccgaagcgca tcagcaaccc gaacaatacc ggcgacagcc ggaactgccg
     5581 tgccggtgtg cagattaatg acagcggtgc ggcgctggga tattacgtca gcgaggacgg
     5641 gtatcctggc tggatgccgc agaaatggac atggataccc cgtgagttac ccggcgggcg
     5701 cgcttggcgt aatcatggtc atagctgttt cctgtgtgaa attgttatcc gctcacaatt
     5761 ccacacaaca tacgagccgg aagcataaag tgtaaagcct ggggtgccta atgagtgagc
     5821 taactcacat taattgcgtt gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc
     5881 cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat tgggcgctct
     5941 tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca
     6001 gctcactcaa aggcggtaat acggttatcc acagaatcag gggataacgc aggaaagaac
     6061 atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt
     6121 ttccataggc tccgcccccc tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg
     6181 cgaaacccga caggactata aagataccag gcgtttcccc ctggaagctc cctcgtgcgc
     6241 tctcctgttc cgaccctgcc gcttaccgga tacctgtccg cctttctccc ttcgggaagc
     6301 gtggcgcttt ctcatagctc acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc
     6361 aagctgggct gtgtgcacga accccccgtt cagcccgacc gctgcgcctt atccggtaac
     6421 tatcgtcttg agtccaaccc ggtaagacac gacttatcgc cactggcagc agccactggt
     6481 aacaggatta gcagagcgag gtatgtaggc ggtgctacag agttcttgaa gtggtggcct
     6541 aactacggct acactagaag aacagtattt ggtatctgcg ctctgctgaa gccagttacc
     6601 ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt
     6661 ttttttgttt gcaagcagca gattacgcgc agaaaaaaag gatctcaaga agatcctttg
     6721 atcttttcta cggggtctga cgctcagtgg aacgaaaact cacgttaagg gattttggtc
     6781 atgagattat caaaaaggat cttcacctag atccttttaa attaaaaatg aagttttaaa
     6841 tcaatctaaa gtatatatga gtaaacttgg tctgacagtt accaatgctt aatcagtgag
     6901 gcacctatct cagcgatctg tctatttcgt tcatccatag ttgcctgact ccccgtcgtg
     6961 tagataacta cgatacggga gggcttacca tctggcccca gtgctgcaat gataccgcga
     7021 gacccacgct caccggctcc agatttatca gcaataaacc agccagccgg aagggccgag
     7081 cgcagaagtg gtcctgcaac tttatccgcc tccatccagt ctattaattg ttgccgggaa
     7141 gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat tgctacaggc
     7201 atcgtggtgt cacgctcgtc gtttggtatg gcttcattca gctccggttc ccaacgatca
     7261 aggcgagtta catgatcccc catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg
     7321 atcgttgtca gaagtaagtt ggccgcagtg ttatcactca tggttatggc agcactgcat
     7381 aattctctta ctgtcatgcc atccgtaaga tgcttttctg tgactggtga gtactcaacc
     7441 aagtcattct gagaatagtg tatgcggcga ccgagttgct cttgcccggc gtcaatacgg
     7501 gataataccg cgccacatag cagaacttta aaagtgctca tcattggaaa acgttcttcg
     7561 gggcgaaaac tctcaaggat cttaccgctg ttgagatcca gttcgatgta acccactcgt
     7621 gcacccaact gatcttcagc atcttttact ttcaccagcg tttctgggtg agcaaaaaca
     7681 ggaaggcaaa atgccgcaaa aaagggaata agggcgacac ggaaatgttg aatactcat
//
