LOCUS       sequence-236043-        4370 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  piggyBac transposon vector for sgRNA expression..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4370)
  AUTHORS   Kagawa H, Shimamoto R, Kim SI, Oceguera-Yanez F, Yamamoto T, 
            Schroeder T, Woltjen K
  TITLE     OVOL1 Influences the Determination and Expansion of iPSC 
            Reprogramming Intermediates.
  JOURNAL   Stem Cell Reports. 2018 Dec 28. pii: S2213-6711(18)30527-7. doi: 
            10.1016/j.stemcr.2018.12.008.
  PUBMED    30639212
REFERENCE   2  (bases 1 to 4370)
  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: "Stem Cell 
            Reports. 2018 Dec 28. pii: S2213-6711(18)30527-7. doi: 
            10.1016/j.stemcr.2018.12.008."
COMMENT     Sequence Label: PB-U6
FEATURES             Location/Qualifiers
     source          1..4370
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      complement(28..456)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(138..159)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     350..369
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     583..605
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     597..614
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     598..614
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     621..640
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        621..639
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     repeat_region   706..768
                     /label=piggyBac right (3') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     promoter        1178..1426
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     1178..1198
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     1349..1368
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_RNA        1445..1520
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     repeat_region   1997..2031
                     /label=piggyBac left (5') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     promoter        complement(2180..2198)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(2219..2235)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2219..2235)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2232..2254)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2243..2259
                     /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(2267..2273,2274..2291,2292..2297))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2312..2333
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2450..2467)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2621..3209)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2701..2720)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3380..4171,4172..4240))
                     /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     4003..4022
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4241..4345)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 aaattgtaag cgttaatatt ttgttaaaat tcgcgttaaa tttttgttaa atcagctcat
       61 tttttaacca ataggccgaa atcggcaaaa tcccttataa atcaaaagaa tagaccgaga
      121 tagggttgag tgttgttcca gtttggaaca agagtccact attaaagaac gtggactcca
      181 acgtcaaagg gcgaaaaacc gtctatcagg gcgatggccc actacgtgaa ccatcaccct
      241 aatcaagttt tttggggtcg aggtgccgta aagcactaaa tcggaaccct aaagggagcc
      301 cccgatttag agcttgacgg ggaaagccgg cgaacgtggc gagaaaggaa gggaagaaag
      361 cgaaaggagc gggcgctagg gcgctggcaa gtgtagcggt cacgctgcgc gtaaccacca
      421 cacccgccgc gcttaatgcg ccgctacagg gcgcgtccca ttcgccattc aggctgcgca
      481 actgttggga agggcgatcg gtgcgggcct cttcgctatt acgccagctg gcgaaagggg
      541 gatgtgctgc aaggcgatta agttgggtaa cgccagggtt ttcccagtca cgacgttgta
      601 aaacgacggc cagtgaattg taatacgact cactataggg cgaattggag ctcggtattc
      661 acgacagcag gctgaataat aaaaaaatta gaaactatta tttaacccta gaaagataat
      721 catattgtga cgtacgttaa agataatcat gcgtaaaatt gacgcatgtg ttttatcggt
      781 ctgtatatcg aggtttattt attaatttga atagatatta agttttatta tatttacact
      841 tacatactaa taataaattc aacaaacaat ttatttatgt ttatttattt attaaaaaaa
      901 aacaaaaact caaaatttct tctataaagt aacaaaactt ttaaacattc tctcttttac
      961 aaaaataaac ttattttgta ctttaaaaac agtcatgttg tattataaaa taagtaatta
     1021 gcttaaccta tacataatag aaacaaatta tacttattag tcagtcagaa acaactttgg
     1081 cacatatcaa tattatgctc tcgttaatta actcgaggta tacttaagcc taggatccag
     1141 atctgtacat tcgaatcgat gaattcgtta accatgtgag ggcctatttc ccatgattcc
     1201 ttcatatttg catatacgat acaaggctgt tagagagata attggaatta atttgactgt
     1261 aaacacaaag atattagtac aaaatacgtg acgtagaaag taataatttc ttgggtagtt
     1321 tgcagtttta aaattatgtt ttaaaatgga ctatcatatg cttaccgtaa cttgaaagta
     1381 tttcgatttc ttggctttat atatcttgtg gaaaggacga aacaccgggt cttcgagaag
     1441 acctgtttta gagctagaaa tagcaagtta aaataaggct agtccgttat caacttgaaa
     1501 aagtggcacc gagtcggtgc ttttttgttt tagagctaga aatagcaagt taaaataagg
     1561 ctagtccgtt tttagcgcgt gcgccaattc tgcagacaaa tggctctaga gcgatcgccc
     1621 gggtttaaac gctagccaac aagctcgtca tcgctttgca gaagagcaga gaggatatgc
     1681 tcatcgtcta aagaactacc cattttatta tatattagtc acgatatcta taacaagaaa
     1741 atatatatat aataagttat cacgtaagta gaacatgaaa taacaatata attatcgtat
     1801 gagttaaatc ttaaaagtca cgtaaaagat aatcatgcgt cattttgact cacgcggtcg
     1861 ttatagttca aaatcagtga cacttaccgc attgacaagc acgcctcacg ggagctccaa
     1921 gcggcgactg agatgtccta aatgcacagc gacggattcg cgctatttag aaagagagag
     1981 caatatttca agaatgcatg cgtcaatttt acgcagacta tctttctagg gttaaaaaag
     2041 atttgcgctt tactcgacct aaactttaaa cacgtcatag aatcttcgtt tgacaaaaac
     2101 cacattgtgg ccaagctgtg tgacgcgacg cgcgctaaag aatggcaaac caagtcgcgc
     2161 gaggtaccag cttttgttcc ctttagtgag ggttaatttc gagcttggcg taatcatggt
     2221 catagctgtt tcctgtgtga aattgttatc cgctcacaat tccacacaac atacgagccg
     2281 gaagcataaa gtgtaaagcc tggggtgcct aatgagtgag ctaactcaca ttaattgcgt
     2341 tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat taatgaatcg
     2401 gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc tcgctcactg
     2461 actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc agctcactca aaggcggtaa
     2521 tacggttatc cacagaatca ggggataacg caggaaagaa catgtgagca aaaggccagc
     2581 aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg ctccgccccc
     2641 ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg acaggactat
     2701 aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc
     2761 cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt tctcatagct
     2821 cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg
     2881 aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc
     2941 cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt agcagagcga
     3001 ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc tacactagaa
     3061 gaacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa agagttggta
     3121 gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt tgcaagcagc
     3181 agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct acggggtctg
     3241 acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgagatta tcaaaaagga
     3301 tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa agtatatatg
     3361 agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc tcagcgatct
     3421 gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact acgatacggg
     3481 agggcttacc atctggcccc agtgctgcaa tgataccgcg agacccacgc tcaccggctc
     3541 cagatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt ggtcctgcaa
     3601 ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta agtagttcgc
     3661 cagttaatag tttgcgcaac gttgttgcca ttgctacagg catcgtggtg tcacgctcgt
     3721 cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt acatgatccc
     3781 ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc agaagtaagt
     3841 tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt actgtcatgc
     3901 catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc tgagaatagt
     3961 gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg ggataatacc gcgccacata
     4021 gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa ctctcaagga
     4081 tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac tgatcttcag
     4141 catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa
     4201 aaaagggaat aagggcgaca cggaaatgtt gaatactcat actcttcctt tttcaatatt
     4261 attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa tgtatttaga
     4321 aaaataaaca aataggggtt ccgcgcacat ttccccgaaa agtgccacct
//
