LOCUS       sequence-460481-        5214 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  A PiggyBac destination vector compatible with gateway cloning 
            system. .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5214)
  AUTHORS   Gillespie WB 3rd, Zhang Y, Ruiz OE, Cerda J 3rd, Ortiz-Guzman J, 
            Sherman M, Turner WD, Largoza G, Mosser LE, Fujimoto E, Chien CB, 
            Kwan KM, Arenkiel BR, Devine WP, Wythe JD
  TITLE     MultiSite Assembly of Gateway Induced Clones (MAGIC): a flexible 
            cloning toolbox for use in vertebrate model systems.
  JOURNAL   Development. 2025 Nov 15;152(22):dev204308. doi: 10.1242/dev.204308.
            Epub 2025 Nov 13.
  PUBMED    41230856
REFERENCE   2  (bases 1 to 5214)
  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: 
            "Development."; date: "2025-11-15"; volume: "152(22):dev204308. 
            doi"; pages: " 10.1242/dev.204308. Epub 2025 Nov 13"
COMMENT     Sequence Label: pB-DEST (JDW 1205)
FEATURES             Location/Qualifiers
     source          1..5214
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      complement(30..458)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(140..161)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     352..371
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     585..607
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     600..616
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     623..642
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        623..641
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     repeat_region   660..694
                     /label=piggyBac left (5') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     protein_bind    1002..1125
                     /gene="mutant version of attR"
                     /label=attR4
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     CDS             complement(1166..1471)
                     /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     complement(1268..1287)
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     CDS             complement(1791..2471)
                     /codon_start=1
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQAGRNLE
                     DPAY"
     primer_bind     2386..2405
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     promoter        complement(join(2525..2531,2532..2549,2550..2555))
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     protein_bind    complement(2580..2703)
                     /gene="mutant version of attR"
                     /label=attR3
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(2711..2727)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     repeat_region   2940..3002
                     /label=piggyBac right (3') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     promoter        complement(3026..3044)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(3065..3081)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3065..3081)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3078..3100)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3089..3105
                     /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(3113..3119,3120..3137,3138..3143))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3158..3179
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3296..3313)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3467..4055)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3547..3566)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4226..5017,5018..5086))
                     /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     4849..4868
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5087..5191)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ctaaattgta agcgttaata ttttgttaaa attcgcgtta aatttttgtt aaatcagctc
       61 attttttaac caataggccg aaatcggcaa aatcccttat aaatcaaaag aatagaccga
      121 gatagggttg agtgttgttc cagtttggaa caagagtcca ctattaaaga acgtggactc
      181 caacgtcaaa gggcgaaaaa ccgtctatca gggcgatggc ccactacgtg aaccatcacc
      241 ctaatcaagt tttttggggt cgaggtgccg taaagcacta aatcggaacc ctaaagggag
      301 cccccgattt agagcttgac ggggaaagcc ggcgaacgtg gcgagaaagg aagggaagaa
      361 agcgaaagga gcgggcgcta gggcgctggc aagtgtagcg gtcacgctgc gcgtaaccac
      421 cacacccgcc gcgcttaatg cgccgctaca gggcgcgtcc cattcgccat tcaggctgcg
      481 caactgttgg gaagggcgat cggtgcgggc ctcttcgcta ttacgccagc tggcgaaagg
      541 gggatgtgct gcaaggcgat taagttgggt aacgccaggg ttttcccagt cacgacgttg
      601 taaaacgacg gccagtgaat tgtaatacga ctcactatag ggcgaattgg gtaccttaac
      661 cctagaaaga tagtctgcgt aaaattgacg catgcattct tgaaatattg ctctctcttt
      721 ctaaatagcg cgaatccgtc gctgtgcatt taggacatct cagtcgccgc ttggagctcc
      781 cgtgaggcgt gcttgtcaat gcggtaagtg tcactgattt tgaactataa cgaccgcgtg
      841 agtcaaaatg acgcatgatt atcttttacg tgacttttaa gatttaactc atacgataat
      901 tatattgtta tttcatgttc tacttacgtg ataacttatt atatatatat tttcttgtta
      961 tagatatcgt cgacttaatt aacatgatta cgccaagcta tcaactttgt atagaaaagt
     1021 tgaacgagaa acgtaaaatg atataaatat caatatatta aattagattt tgcataaaaa
     1081 acagactaca taatactgta aaacacaaca tatccagtca ctatggtcga cctgcagact
     1141 ggctgtgtat aagggagcct gacatttata ttccccagaa catcaggtta atggcgtttt
     1201 tgatgtcatt ttcgcggtgg ctgagatcag ccacttcttc cccgataacg gagaccggca
     1261 cactggccat atcggtggtc atcatgcgcc agctttcatc cccgatatgc accaccgggt
     1321 aaagttcacg ggggacttta tctgacagca gacgtgcact ggccaggggg atcaccatcc
     1381 gtcgcccggg cgtgtcaata atatcactct gtacatccac aaacagacga taacggctct
     1441 ctcttttata ggtgtaaacc ttaaactgca tttcaccagc ccctgttctc gtcggcaaag
     1501 gagccgttca tttcaataaa ccgggcgacc tcagccatcc cttcctgatt ttccgctttc
     1561 cagcgttcgg cacgcagacg acgggcttca ttctgcatgg ttgtgcttac cgaaccggag
     1621 atattgacat catatatgcc ttgagcaact gatagctgtc gctgtcaact gtcactgtaa
     1681 tacgctgctt catagcatac ctctttttga catacttcgg gtatacatat cagtatatat
     1741 tcttataccg caaaaatcag cgcgcaaata cgcatactgt tatctggctt ttagtaagcc
     1801 ggatcctcta gattacgccc cgcctgccac tcatcgcagt actgttgtaa ttcattaagc
     1861 attctgccga catggaagcc atcacaaacg gcatgatgaa cctgaatcgc cagcggcatc
     1921 agcaccttgt cgccttgcgt ataatatttg cccatggtga aaacgggggc gaagaagttg
     1981 tccatattgg ccacgtttaa atcaaaactg gtgaaactca cccagggatt ggctgagacg
     2041 aaaaacatat tctcaataaa ccctttaggg aaataggcca ggttttcacc gtaacacgcc
     2101 acatcttgcg aatatatgtg tagaaactgc cggaaatcgt cgtggtattc actccagagc
     2161 gatgaaaacg tttcagtttg ctcatggaaa acggtgtaac aagggtgaac actatcccat
     2221 atcaccagct caccgtcttt cattgccata cggaattccg gatgagcatt catcaggcgg
     2281 gcaagaatgt gaataaaggc cggataaaac ttgtgcttat ttttctttac ggtctttaaa
     2341 aaggccgtaa tatccagctg aacggtctgg ttataggtac attgagcaac tgactgaaat
     2401 gcctcaaaat gttctttacg atgccattgg gatatatcaa cggtggtata tccagtgatt
     2461 tttttctcca ttttagcttc cttagctcct gaaaatctcg acggatccta actcaaaatc
     2521 cacacattat acgagccgga agcataaagt gtaaagcctg gggtgcctaa tgcggccgcc
     2581 atagtgactg gatatgttgt gttttacagt attatgtagt ctgtttttta tgcaaaatct
     2641 aatttaatat attgatattt atatcatttt acgtttctcg ttcaacttta ttatacatag
     2701 ttgataattc actggccgtc gttttacggt accatcgatg atgataagct tgtttaaacg
     2761 ttaacggatc ctttgttact ttatagaaga aattttgagt ttttgttttt ttttaataaa
     2821 taaataaaca taaataaatt gtttgttgaa tttattatta gtatgtaagt gtaaatataa
     2881 taaaacttaa tatctattca aattaataaa taaacctcga tatacagacc gataaaacac
     2941 atgcgtcaat tttacgcatg attatcttta acgtacgtca caatatgatt atctttctag
     3001 ggttaaagag ctccagcttt tgttcccttt agtgagggtt aatttcgagc ttggcgtaat
     3061 catggtcata gctgtttcct gtgtgaaatt gttatccgct cacaattcca cacaacatac
     3121 gagccggaag cataaagtgt aaagcctggg gtgcctaatg agtgagctaa ctcacattaa
     3181 ttgcgttgcg ctcactgccc gctttccagt cgggaaacct gtcgtgccag ctgcattaat
     3241 gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg gcgctcttcc gcttcctcgc
     3301 tcactgactc gctgcgctcg gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg
     3361 cggtaatacg gttatccaca gaatcagggg ataacgcagg aaagaacatg tgagcaaaag
     3421 gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc
     3481 gcccccctga cgagcatcac aaaaatcgac gctcaagtca gaggtggcga aacccgacag
     3541 gactataaag ataccaggcg tttccccctg gaagctccct cgtgcgctct cctgttccga
     3601 ccctgccgct taccggatac ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc
     3661 atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg
     3721 tgcacgaacc ccccgttcag cccgaccgct gcgccttatc cggtaactat cgtcttgagt
     3781 ccaacccggt aagacacgac ttatcgccac tggcagcagc cactggtaac aggattagca
     3841 gagcgaggta tgtaggcggt gctacagagt tcttgaagtg gtggcctaac tacggctaca
     3901 ctagaagaac agtatttggt atctgcgctc tgctgaagcc agttaccttc ggaaaaagag
     3961 ttggtagctc ttgatccggc aaacaaacca ccgctggtag cggtggtttt tttgtttgca
     4021 agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg
     4081 ggtctgacgc tcagtggaac gaaaactcac gttaagggat tttggtcatg agattatcaa
     4141 aaaggatctt cacctagatc cttttaaatt aaaaatgaag ttttaaatca atctaaagta
     4201 tatatgagta aacttggtct gacagttacc aatgcttaat cagtgaggca cctatctcag
     4261 cgatctgtct atttcgttca tccatagttg cctgactccc cgtcgtgtag ataactacga
     4321 tacgggaggg cttaccatct ggccccagtg ctgcaatgat accgcgagac ccacgctcac
     4381 cggctccaga tttatcagca ataaaccagc cagccggaag ggccgagcgc agaagtggtc
     4441 ctgcaacttt atccgcctcc atccagtcta ttaattgttg ccgggaagct agagtaagta
     4501 gttcgccagt taatagtttg cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac
     4561 gctcgtcgtt tggtatggct tcattcagct ccggttccca acgatcaagg cgagttacat
     4621 gatcccccat gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc gttgtcagaa
     4681 gtaagttggc cgcagtgtta tcactcatgg ttatggcagc actgcataat tctcttactg
     4741 tcatgccatc cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag tcattctgag
     4801 aatagtgtat gcggcgaccg agttgctctt gcccggcgtc aatacgggat aataccgcgc
     4861 cacatagcag aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct
     4921 caaggatctt accgctgttg agatccagtt cgatgtaacc cactcgtgca cccaactgat
     4981 cttcagcatc ttttactttc accagcgttt ctgggtgagc aaaaacagga aggcaaaatg
     5041 ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat actcatactc ttcctttttc
     5101 aatattattg aagcatttat cagggttatt gtctcatgag cggatacata tttgaatgta
     5161 tttagaaaaa taaacaaata ggggttccgc gcacatttcc ccgaaaagtg ccac
//
