LOCUS       sequence-442451-        6760 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  PiggyBac transposon plasmid for doxycycline inducible expression of 
            HOXA10.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6760)
  AUTHORS   Pierson Smela M, Adams J, Ma C, Breimann L, Widocki U, Shioda T, 
            Church GM
  TITLE     Induction of Meiosis from Human Pluripotent Stem Cells
  JOURNAL   bioRxiv 2024.05.31.596483
REFERENCE   2  (bases 1 to 6760)
  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: "bioRxiv 
            2024.05.31.596483"
COMMENT     Sequence Label: pPB-cT3G-cERP2-HOXA10
FEATURES             Location/Qualifiers
     source          1..6760
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    17..35
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    53..71
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    89..107
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    125..143
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    161..179
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    197..215
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    233..251
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     278..302
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     protein_bind    368..392
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     protein_bind    complement(675..699)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             701..727
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     polyA_signal    894..1028
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     repeat_region   1300..1334
                     /label=piggyBac left (5') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     primer_bind     complement(1345..1362)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(1345..1361)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1354..1376)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     complement(1570..1589)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     1691..1713
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(1752..1770)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        1838..1942
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1943..2011,2012..2803)
                     /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(2161..2180)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2974..3562
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3463..3482
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3716..3733
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    3939..3960
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(3975..3980,3981..3998,3999..4005)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4013..4029
                     /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     4018..4040
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     4037..4053
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     4037..4053
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     repeat_region   4079..4141
                     /label=piggyBac right (3') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     polyA_signal    4317..4398
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     CDS             complement(5115..5171)
                     /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             complement(5181..5924)
                     /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             complement(5925..5948)
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     regulatory      5948..5957
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     promoter        complement(5977..6188)
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     enhancer        6189..6568
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
ORIGIN
        1 atcacctcga gtttactccc tatcagtgat agagaacgta tgaagagttt actccctatc
       61 agtgatagag aacgtatgca gactttactc cctatcagtg atagagaacg tataaggagt
      121 ttactcccta tcagtgatag agaacgtatg accagtttac tccctatcag tgatagagaa
      181 cgtatctaca gtttactccc tatcagtgat agagaacgta tatccagttt actccctatc
      241 agtgatagag aacgtataag cttgcctata taagcagagc tcgtttagtg aaccgtcaga
      301 tcgcctggag acgccatcca cgctgttttg acctccatag aagacaccgg gaccgatcca
      361 gcctaccaca agtttgtaca aaaaagttgg catgtgtcaa ggcaattcca aaggtgaaaa
      421 cgcagccaac tggctcacgg caaagagtgg tcggaagaag cgctgcccct acacgaagca
      481 ccagacactg gagctggaga aggagtttct gttcaatatg taccttactc gagagcggcg
      541 cctagagatt agccgcagcg tccacctcac ggacagacaa gtgaaaatct ggtttcagaa
      601 ccgcaggatg aaactgaaga aaatgaatcg agaaaaccgg atccgggagc tcacagccaa
      661 ctttaatttt tcctgcccaa ctttcttgta caaagtggta tacccatacg atgttccaga
      721 ttacgcttag taatgagaaa tactcagaag atgtcactca ccatagtccg cagtgaaaga
      781 taaatgatct tttctggggt ggctcccaag ttttagagct agaaatagca agtaggccac
      841 tgagcaagaa ctatgaatga gatctttaga catattgcgg ccgctttctc cgcgatccag
      901 acatgataag atacattgat gagtttggac aaaccaaaac tagaatgcag tgaaaaaaat
      961 gccttatttg tgaaatttgt gatgctattg ccttatttgt aaccattata agctgcaata
     1021 aacaagttat ctataacaag aaaatatata tataataagt tatcacgtaa gtagaacatg
     1081 aaataacaat ataattatcg tatgagttaa atcttaaaag tcacgtaaaa gataatcatg
     1141 cgtcattttg actcacgcgg tcgttatagt tcaaaatcag tgacacttac cgcattgaca
     1201 agcacgcctc acgggagctc caagcggcga ctgagatgtc ctaaatgcac agcgacggat
     1261 tcgcgctatt tagaaagaga gagcaatatt tcaagaatgc atgcgtcaat tttacgcaga
     1321 ctatctttct agggttaaga attcactggc cgtcgtttta caacgtcgtg actgggaaaa
     1381 ccctggcgtt acccaactta atcgccttgc agcacatccc cctttcgcca gctggcgtaa
     1441 tagcgaagag gcccgcaccg atcgcccttc ccaacagttg cgcagcctga atggcgaatg
     1501 gcgcctgatg cggtattttc tccttacgca tctgtgcggt atttcacacc gcatatggtg
     1561 cactctcagt acaatctgct ctgatgccgc atagttaagc cagccccgac acccgccaac
     1621 acccgctgac gcgccctgac gggcttgtct gctcccggca tccgcttaca gacaagctgt
     1681 gaccatgtct ccgggagctg catgtgtcag aggttttcac cgtcatcacc gaaacacgcg
     1741 agttcgaaag ggcctcgtga tacgcctatt tttataggtt aatgtcatga taataatggt
     1801 ttcttagacg tcaggtggca cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt
     1861 tttctaaata cattcaaata tgtatccgct catgagacaa taaccctgat aaatgcttca
     1921 ataatattga aaaaggaaga gtatgagtat tcaacatttc cgtgtcgccc ttattccctt
     1981 ttttgcggca ttttgccttc ctgtttttgc tcacccagaa acgctggtga aagtaaaaga
     2041 tgctgaagat cagttgggtg cacgagtggg ttacatcgaa ctggatctca acagcggtaa
     2101 gatccttgag agttttcgcc ccgaagaacg ttttccaatg atgagcactt ttaaagttct
     2161 gctatgtggc gcggtattat cccgtattga cgccgggcaa gagcaactcg gtcgccgcat
     2221 acactattct cagaatgact tggttgagta ctcaccagtc acagaaaagc atcttacgga
     2281 tggcatgaca gtaagagaat tatgcagtgc tgccataacc atgagtgata acactgcggc
     2341 caacttactt ctgacaacga tcggaggacc gaaggagcta accgcttttt tgcacaacat
     2401 gggggatcat gtaactcgcc ttgatcgttg ggaaccggag ctgaatgaag ccataccaaa
     2461 cgacgagcgt gacaccacga tgcctgtagc aatggcaaca acgttgcgca aactattaac
     2521 tggcgaacta cttactctag cttcccggca acaattaata gactggatgg aggcggataa
     2581 agttgcagga ccacttctgc gctcggccct tccggctggc tggtttattg ctgataaatc
     2641 tggagccggt gagcgtggtt cacgcggtat cattgcagca ctggggccag atggtaagcc
     2701 ctcccgtatc gtagttatct acacgacggg gagtcaggca actatggatg aacgaaatag
     2761 acagatcgct gagataggtg cctcactgat taagcattgg taactgtcag accaagttta
     2821 ctcatatata ctttagattg atttaaaact tcatttttaa tttaaaagga tctaggtgaa
     2881 gatccttttt gataatctca tgaccaaaat cccttaacgt gagttttcgt tccactgagc
     2941 gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat
     3001 ctgctgcttg caaacaaaaa aaccaccgct accagcggtg gtttgtttgc cggatcaaga
     3061 gctaccaact ctttttccga aggtaactgg cttcagcaga gcgcagatac caaatactgt
     3121 tcttctagtg tagccgtagt taggccacca cttcaagaac tctgtagcac cgcctacata
     3181 cctcgctctg ctaatcctgt taccagtggc tgctgccagt ggcgataagt cgtgtcttac
     3241 cgggttggac tcaagacgat agttaccgga taaggcgcag cggtcgggct gaacgggggg
     3301 ttcgtgcaca cagcccagct tggagcgaac gacctacacc gaactgagat acctacagcg
     3361 tgagctatga gaaagcgcca cgcttcccga agggagaaag gcggacaggt atccggtaag
     3421 cggcagggtc ggaacaggag agcgcacgag ggagcttcca gggggaaacg cctggtatct
     3481 ttatagtcct gtcgggtttc gccacctctg acttgagcgt cgatttttgt gatgctcgtc
     3541 aggggggcgg agcctatgga aaaacgccag caacgcggcc tttttacggt tcctggcctt
     3601 ttgctggcct tttgctcaca tgttctttcc tgcgttatcc cctgattctg tggataaccg
     3661 tattaccgcc tttgagtgag ctgataccgc tcgccgcagc cgaacgaccg agcgcagcga
     3721 gtcagtgagc gaggaagcgg atgagcgccc aatacgcaaa ccgcctctca ccgcgcgttg
     3781 gccgattcat taatgcagct ggcacgacag gtttcccgac tggaaagcgg gcagtgagcg
     3841 caacgcaaac cgcctctccc cgcgcgttgg ccgattcatt aatgcagctg gcacgacagg
     3901 tttcccgact ggaaagcggg cagtgagcgc aacgcaatta atgtgagtta gctcactcat
     3961 taggcacccc aggctttaca ctttatgctt ccggctcgta tgttgtgtgg aattgtgagc
     4021 ggataacaat ttcacacagg aaacagctat gaccatgatt acgccaaggt cgacttaacc
     4081 ctagaaagat aatcatattg tgacgtacgt taaagataat catgcgtaaa attgacgcat
     4141 gtgttttatc ggtctgtata tcgaggttta tttattaatt tgaatagata ttaagtttta
     4201 ttatatttac acttacatac taataataaa ttcaacaaac aatttattta tgtttattta
     4261 tttattaaaa aaaaacaaaa actcaaaatt tcttctataa agtaacaaaa cttttagcag
     4321 tgaaaaaaat gctttatttg tgaaatttgt gatgctattg ctttatttgt aaccattata
     4381 agctgcaata aacaagttaa caacaacaat tgcattcatt ttatgtttca ggttcagggg
     4441 gaggtgtggg aggtttttta aagcaagtaa aacctctaca aatgtggtat ggctgattat
     4501 gatcctctgg agattcatgc gccaggtttt ctggtcatgc accacgttcg tgggccctca
     4561 ggaacctcca cgtcagcggt aactgtgaag cccagtcgct cgtaaaaagg aagattccgg
     4621 ggagcactgg tttcgaggaa cgccggcacg ccagcccgtt ccgcggcctc aacaccagga
     4681 agaacgacag cgcttcccaa acctttacct tggtgatctg ggctgacacc gacagtggca
     4741 agaaaccacg caggctcctt tggtctatga ggagccagca agccctccat ttgctgttgt
     4801 gccgccaacc tactccctga cagctcggcc atccgcggac ctatttccgc gaacactgcg
     4861 ccggcttcga cgctttccgg agtagtccag accgcaactg cggcaccatc gtccgcaacc
     4921 caaactttac caatgtccag acccacgcgc gtaagaaaca gctcttgcaa ctcagttacc
     4981 ctctctatgt gtctgtccgg atctacggta tggcgggtcg ctggataatc tgcaaatgct
     5041 gcggcaagtg ttcggacagc ccgtggtaca tcgtcacggg tagcgagcct aacagtgggc
     5101 ttatattccg tcatggggcc ggggttctcc tccacgtcgc cggcctgctt cagcaggctg
     5161 aagttggtgg cgccgctgcc cccggggagc atgtcaaggt caaaatcgtc aagagcgtca
     5221 gcaggcagca tatcaaggtc aaagtcgtca agggcatcgg ctgggagcat gtctaagtca
     5281 aaatcgtcaa gggcgtcggt cggcccgccg ctttcgcact ttagctgttt ctccaggcca
     5341 catatgatta gttccaggcc gaaaaggaag gcaggttcgg ctccctgccg gtcgaacagc
     5401 tcaattgctt gtttcagaag tgggggcata gaatcggtgg taggtgtctc tctttcctct
     5461 tttgctactt gatgctcctg ttcctccaat acgcagccca gtgtaaagtg gcccacggcg
     5521 gacagagcgt acagtgcgtt ctccagggag aagccttgct gacacaggaa cgcgagctga
     5581 ttttccaggg tttcgtactg tttctctgtt gggcgggtgc cgagatgcac tttagccccg
     5641 tcacgatgtg agaggagagc acagcggtat gacttggcgt tgttccgcag aaagtcttgc
     5701 catgactcgc cttccagggg gcaggagtgg gtatgatgcc tgtccagcat ctcgattggc
     5761 agggcatcga gcagggcccg cttgttcttc acgtgccagt acagggtagg ctgctcaact
     5821 cccagctttt gagcgagttt ccttgtcgtc aggccttcga taccgactcc attgagtaat
     5881 tccagagcag agtttatgac tttgctcttg tccagtctag acatcttatc gtcatcgtct
     5941 ttgtaatcca tggtggcgga tcccgcgtca cgacacctgt gttctggcgg caaacccgtt
     6001 gcgaaaaaga acgttcacgg cgactactgc acttatatac ggttctcccc caccctcggg
     6061 aaaaaggcgg agccagtaca cgacatcact ttcccagttt accccgcgcc accttctcta
     6121 ggcaccggtt caattgccga cccctccccc caacttctcg ggcactgtgg gccatgtgcg
     6181 ctctgcccca tggtaatagc catgactaat acgtagatgt actgccaagt aggaaagtgc
     6241 cataaggtca tgtactgggc ataatgccag gcgggccatt taccgtcatt gacgtcaata
     6301 gggggcgtac ttggcatatg atacacttga tgtactgcca agtgggcagt ttaccgtaaa
     6361 tactccaccc attgacgtca atggaaagtg cctattggcg ttactatggg aacatacgtc
     6421 attattgacg tcaatgggcg ggggtcgttg ggcggtcagc caggcgggcc atttaccgta
     6481 agttatgtaa cgcggaactc catatatggg ctatgaacta atgaccccgt aattgattac
     6541 tattaataac tagtcaataa tcaatgtcat aagctctaat cgcataatcg agctaagagc
     6601 gattaatatg cttagctagc ttagctaagc tagatttctc tattctccca caatatatag
     6661 cagctattag cttagcagtc cgaatacgat cgtccacgat ccataagcgt aacacatttg
     6721 agtagcgact gacgggcacc ggagccactc gagtggaatt
//
