LOCUS       sequence-478112-        3708 bp    DNA     circular SYN 10-JUL-2025
DEFINITION  PaqCI Golden Gate assembly plasmid encoding the backbone for 
            PiggyBac-mediated integration.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3708)
  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 3708)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Jul 10, 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: pKG1334_pHarbor-PiggyBac_backbone
FEATURES             Location/Qualifiers
     source          1..3708
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        11..115
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(116..184,185..976)
                     /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(334..353)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      1147..1735
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1636..1655
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     repeat_region   1817..1851
                     /label=piggyBac left (5') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     protein_bind    1888..2012
                     /gene="mutant version of attR"
                     /label=attR1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        join(2045..2050,2051..2068,2069..2075)
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     CDS             2129..2809
                     /codon_start=1
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQAGRNLE
                     DPAY"
     primer_bind     complement(2195..2214)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     CDS             3129..3434
                     /codon_start=1
                     /gene="ccdB"
                     /product="CcdB, a bacterial toxin that poisons DNA gyrase"
                     /label=ccdB
                     /note="Plasmids containing the ccdB gene cannot be 
                     propagated in standard E. coli strains."
                     /translation="MQFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDK
                     VPRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     primer_bind     3313..3332
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     protein_bind    complement(3471..3595)
                     /gene="mutant version of attR"
                     /label=attR2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     repeat_region   3643..3705
                     /label=piggyBac right (3') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
ORIGIN
        1 ctataatgtg cgcggaaccc ctatttgttt atttttctaa atacattcaa atatgtatcc
       61 gctcatgaga caataaccct gataaatgct tcaataatat tgaaaaagga agagtatgag
      121 tattcaacat ttccgtgtcg cccttattcc cttttttgcg gcattttgcc ttcctgtttt
      181 tgctcaccca gaaacgctgg tgaaagtaaa agatgctgaa gatcagttgg gtgcacgagt
      241 gggttacatc gaactggatc tcaacagcgg taagatcctt gagagttttc gccccgaaga
      301 acgttttcca atgatgagca cttttaaagt tctgctatgt ggcgcggtat tatcccgtat
      361 tgacgccggg caagagcaac tcggtcgccg catacactat tctcagaatg acttggttga
      421 gtactcacca gtcacagaaa agcatcttac ggatggcatg acagtaagag aattatgcag
      481 tgctgccata accatgagtg ataacactgc ggccaactta cttctgacaa cgatcggagg
      541 accgaaggag ctaaccgctt ttttgcacaa catgggggat catgtaactc gccttgatcg
      601 ttgggaaccg gagctgaatg aagccatacc aaacgacgag cgtgacacca cgatgcctgt
      661 agcaatggca acaacgttgc gcaaactatt aactggcgaa ctacttactc tagcttcccg
      721 gcaacaatta atagactgga tggaggcgga taaagttgca ggaccacttc tgcgctcggc
      781 ccttccggct ggctggttta ttgctgataa atctggagcc ggtgagcgtg ggtctcgcgg
      841 tatcattgca gcactggggc cagatggtaa gccctcccgt atcgtagtta tctacacgac
      901 ggggagtcag gcaactatgg atgaacgaaa tagacagatc gctgagatag gtgcctcact
      961 gattaagcat tggtaactgt cagaccaagt ttactcatat atactttaga ttgatttaaa
     1021 acttcatttt taatttaaaa ggatctaggt gaagatcctt tttgataatc tcatgaccaa
     1081 aatcccttaa cgtgagtttt cgttccactg agcgtcagac cccgtagaaa agatcaaagg
     1141 atcttcttga gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc
     1201 gctaccagcg gtggtttgtt tgccggatca agagctacca actctttttc cgaaggtaac
     1261 tggcttcagc agagcgcaga taccaaatac tgttcttcta gtgtagccgt agttaggcca
     1321 ccacttcaag aactctgtag caccgcctac atacctcgct ctgctaatcc tgttaccagt
     1381 ggctgctgcc agtggcgata agtcgtgtct taccgggttg gactcaagac gatagttacc
     1441 ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc acacagccca gcttggagcg
     1501 aacgacctac accgaactga gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc
     1561 cgaagggaga aaggcggaca ggtatccggt aagcggcagg gtcggaacag gagagcgcac
     1621 gagggagctt ccagggggaa acgcctggta tctttatagt cctgtcgggt ttcgccacct
     1681 ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc
     1741 cagcaacgcg gcctttttac ggttcctggc cttttgctgg ccttttgctc acatgttctt
     1801 tcctgcgtta tcttcaccct agaaagatag tctgcgtaaa attgacgcat gtccttactc
     1861 atagcaggtg ccactagtgg agttccgaca agtttgtaca aaaaagctga acgagaaacg
     1921 taaaatgata taaatatcaa tatattaaat tagattttgc ataaaaaaca gactacataa
     1981 tactgtaaaa cacaacatat ccagtcacta tggcggattc gcggccgcat taggcacccc
     2041 aggctttaca ctttatgctt ccggctcgta taatgtgtgg attttgagtt aggatccgtc
     2101 gagattttca ggagctaagg aagctaaaat ggagaaaaaa atcactggat ataccacagt
     2161 tgatatatcc caatggcatc gtaaagaaca ttttgaggca tttcagtcag ttgctcaatg
     2221 tacctataac cagaccgttc agctggatat tacggccttt ttaaagaccg taaagaaaaa
     2281 taagcacaag ttttatccgg cctttattca cattcttgcc cgcctgatga atgctcatcc
     2341 ggaattccgt atggcaatga aagacggtga gctggtgata tgggatagtg ttcacccttg
     2401 ttacaccgtt ttccatgagc aaactgaaac gttttcatcg ctctggagtg aataccacga
     2461 cgatttccgg cagtttctac acatatattc gcaagatgtg gcgtgttacg gtgaaaacct
     2521 ggcctatttc cctaaagggt ttattgagaa tatgtttttc gtttcagcca atccctgggt
     2581 gagtttcacc agttttgatt taaacgtggc caatatggac aacttcttcg cccccgtttt
     2641 caccatgggc aaatattata cgcaaggcga caaggtgctg atgccgctgg cgattcaggt
     2701 tcatcatgcc gtttgtgatg gcttccatgt cggcagaatg cttaatgaat tacaacagta
     2761 ctgcgatgag tggcaggcgg ggcgtaatct agaggatccg gcttactaaa agccagataa
     2821 cagtatgcgt atttgcgcgc tgatttttgc ggtataagaa tatatactga tatgtatacc
     2881 cgaagtatgt caaaaagagg tatgctatga agcagcgtat tacagtgaca gttgacagcg
     2941 acagctatca gttgctcaag gcatatatga tgtcaatatc tccggtctgg taagcacaac
     3001 catgcagaat gaagcccgtc gtctgcgtgc cgaacgctgg aaagcggaaa atcaggaagg
     3061 gatggctgag gtcgcccggt ttattgaaat gaacggctct tttgctgacg agaacagggg
     3121 ctggtgaaat gcagtttaag gtttacacct ataaaagaga gagccgttat cgtctgtttg
     3181 tggatgtaca gagtgatatt attgacacgc ccgggcgacg gatggtgatc cccctggcca
     3241 gtgcacgtct gctgtcagat aaagtccccc gtgaacttta cccggtggtg catatcgggg
     3301 atgaaagctg gcgcatgatg accaccgata tggccagtgt gccggtttcc gttatcgggg
     3361 aagaagtggc tgatctcagc caccgcgaaa atgacatcaa aaacgccatt aacctgatgt
     3421 tctggggaat ataaatgtca ggctccctta tacacagcct gcaggtcgac catagtgact
     3481 ggatatgttg tgttttacag tattatgtag tctgtttttt atgcaaaatc taatttaata
     3541 tattgatatt tatatcattt tacgtttctc gttcagcttt cttgtacaaa gtggtcacct
     3601 gctatgcgct gatataggta ggctcgagca gagaagctta cacatgcgtc aattttacgc
     3661 atgattatct ttaacgtacg tcacaatatg attatctttc tagggtta
//
