LOCUS       sequence-82382-e        5970 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  for Phic31 integrase-mediated transgene integration.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5970)
  AUTHORS   Tasic B, Hippenmeyer S, Wang C, Gamboa M, Zong H, Chen-Tsai Y, Luo L
  TITLE     Site-specific integrase-mediated transgenesis in mice via pronuclear
            injection.
  JOURNAL   Proc Natl Acad Sci U S A. 2011 May 10;108(19):7902-7. doi: 
            10.1073/pnas.1019507108. Epub 2011 Apr 4.
  PUBMED    21464299
REFERENCE   2  (bases 1 to 5970)
  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"; doi: 
            "10.1073/pnas.1019507108"; journalName: "Proc Natl Acad Sci U S A"; 
            date: "2011-05-10- 10"; volume: "108"; issue: "19"; pages: "7902-7"
COMMENT     Sequence Label: pBT316_pattB-pCA-GFP-pA
FEATURES             Location/Qualifiers
     source          1..5970
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      complement(33..461)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(143..164)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     355..374
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     588..610
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     602..619
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     603..619
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     626..645
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        626..644
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    695..764
                     /label=attB
                     /bound_moiety="phage phi-C31 integrase"
                     /note="attB site for the phi-C31 integrase (Groth et al., 
                     2000)"
     enhancer        1070..1373
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        1375..1652
                     /label=chicken beta-actin promoter
     intron          1654..2671
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     2595..2617
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     2679..2698
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     regulatory      2749..2758
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(2755..2757,2758..2760,2761..3474)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     ANFKIRHNIEDGGVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(2800..2821)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(3061..3080)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3408..3429
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    3596..3717
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3633..3652)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3687..3706
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     promoter        complement(3785..3803)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(3824..3840)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3824..3840)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3837..3859)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3848..3864
                     /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(3872..3878,3879..3896,3897..3902))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3917..3938
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4055..4072)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4226..4814)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4306..4325)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4985..5776,5777..5845))
                     /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     5608..5627
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5846..5950)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 cacctaaatt gtaagcgtta atattttgtt aaaattcgcg ttaaattttt gttaaatcag
       61 ctcatttttt aaccaatagg ccgaaatcgg caaaatccct tataaatcaa aagaatagac
      121 cgagataggg ttgagtgttg ttccagtttg gaacaagagt ccactattaa agaacgtgga
      181 ctccaacgtc aaagggcgaa aaaccgtcta tcagggcgat ggcccactac gtgaaccatc
      241 accctaatca agttttttgg ggtcgaggtg ccgtaaagca ctaaatcgga accctaaagg
      301 gagcccccga tttagagctt gacggggaaa gccggcgaac gtggcgagaa aggaagggaa
      361 gaaagcgaaa ggagcgggcg ctagggcgct ggcaagtgta gcggtcacgc tgcgcgtaac
      421 caccacaccc gccgcgctta atgcgccgct acagggcgcg tcccattcgc cattcaggct
      481 gcgcaactgt tgggaagggc gatcggtgcg ggcctcttcg ctattacgcc agctggcgaa
      541 agggggatgt gctgcaaggc gattaagttg ggtaacgcca gggttttccc agtcacgacg
      601 ttgtaaaacg acggccagtg aattgtaata cgactcacta tagggcgaat tgggtaccgg
      661 gccccccctc gaggtcgacg atgtaggtca cggtctcgaa gccgcggtgc gggtgccagg
      721 gcgtgccctt gggctccccg ggcgcgtact ccacctcacc catctggtcc atcatgatga
      781 acgggtcgag gtggcggtag ttgatcccgg cgaacgcgcg gcgcaccggg aagccctcgc
      841 cctcgaaacc gctgggcgcg gtggtcacgg tgagcacggg acgtgcgacg gcgtcggcgg
      901 gtgcggatac gcggggcagc gtcagcgggt tctcgacggt cacggcgggc atgtcgacgg
      961 tatcgataag cttcccttaa ttaaccgttt aaacaattct gcaggaatct agttattaat
     1021 agtaatcaat tacggggtca ttagttcata gcccatatat ggagttccgc gttacataac
     1081 ttacggtaaa tggcccgcct ggctgaccgc ccaacgaccc ccgcccattg acgtcaataa
     1141 tgacgtatgt tcccatagta acgccaatag ggactttcca ttgacgtcaa tgggtggagt
     1201 atttacggta aactgcccac ttggcagtac atcaagtgta tcatatgcca agtacgcccc
     1261 ctattgacgt caatgacggt aaatggcccg cctggcatta tgcccagtac atgaccttat
     1321 gggactttcc tacttggcag tacatctacg tattagtcat cgctattacc atggtcgagg
     1381 tgagccccac gttctgcttc actctcccca tctccccccc ctccccaccc ccaattttgt
     1441 atttatttat tttttaatta ttttgtgcag cgatgggggc gggggggggg ggggggcgcg
     1501 cgccaggcgg ggcggggcgg ggcgaggggc ggggcggggc gaggcggaga ggtgcggcgg
     1561 cagccaatca gagcggcgcg ctccgaaagt ttccttttat ggcgaggcgg cggcggcggc
     1621 ggccctataa aaagcgaagc gcgcggcggg cggggagtcg ctgcgacgct gccttcgccc
     1681 cgtgccccgc tccgccgccg cctcgcgccg cccgccccgg ctctgactga ccgcgttact
     1741 cccacaggtg agcgggcggg acggcccttc tcctccgggc tgtaattagc gcttggttta
     1801 atgacggctt gtttcttttc tgtggctgcg tgaaagcctt gaggggctcc gggagggccc
     1861 tttgtgcggg gggagcggct cggggggtgc gtgcgtgtgt gtgtgcgtgg ggagcgccgc
     1921 gtgcggctcc gcgctgcccg gcggctgtga gcgctgcggg cgcggcgcgg ggctttgtgc
     1981 gctccgcagt gtgcgcgagg ggagcgcggc cgggggcggt gccccgcggt gcgggggggg
     2041 ctgcgagggg aacaaaggct gcgtgcgggg tgtgtgcgtg ggggggtgag cagggggtgt
     2101 gggcgcgtcg gtcgggctgc aaccccccct gcacccccct ccccgagttg ctgagcacgg
     2161 cccggcttcg ggtgcggggc tccgtacggg gcgtggcgcg gggctcgccg tgccgggcgg
     2221 ggggtggcgg caggtggggg tgccgggcgg ggcggggccg cctcgggccg gggagggctc
     2281 gggggagggg cgcggcggcc cccggagcgc cggcggctgt cgaggcgcgg cgagccgcag
     2341 ccattgcctt ttatggtaat cgtgcgagag ggcgcaggga cttcctttgt cccaaatctg
     2401 tgcggagccg aaatctggga ggcgccgccg caccccctct agcgggcgcg gggcgaagcg
     2461 gtgcggcgcc ggcaggaagg aaatgggcgg ggagggcctt cgtgcgtcgc cgcgccgccg
     2521 tccccttctc cctctccagc ctcggggctg tccgcggggg gacggctgcc ttcggggggg
     2581 acggggcagg gcggggttcg gcttctggcg tgtgaccggc ggctctagag cctctgctaa
     2641 ccatgttcat gccttcttct ttttcctaca gctcctgggc aacgtgctgg ttattgtgct
     2701 gtctcatcat tttggcaaag aattgatttg ataccgcggg cccgggccgc caccatggtg
     2761 agcaagggcg aggagctgtt caccggggtg gtgcccatcc tggtcgagct ggacggcgac
     2821 gtaaacggcc acaagttcag cgtgtccggc gagggcgagg gcgatgccac ctacggcaag
     2881 ctgaccctga agttcatctg caccaccggc aagctgcccg tgccctggcc caccctcgtg
     2941 accaccctga cctacggcgt gcagtgcttc agccgctacc ccgaccacat gaagcagcac
     3001 gacttcttca agtccgccat gcccgaaggc tacgtccagg agcgcaccat cttcttcaag
     3061 gacgacggca actacaagac ccgcgccgag gtgaagttcg agggcgacac cctggtgaac
     3121 cgcatcgagc tgaagggcat cgacttcaag gaggacggca acatcctggg gcacaagctg
     3181 gagtacaact acaacagcca caacgtctat atcatggccg acaagcagaa gaacggcatc
     3241 aaggcgaact tcaagatccg ccacaacatc gaggacggcg gcgtgcagct cgccgaccac
     3301 taccagcaga acacccccat cggcgacggc cccgtgctgc tgcccgacaa ccactacctg
     3361 agcacccagt ccgccctgag caaagacccc aacgagaagc gcgatcacat ggtcctgctg
     3421 gagttcgtga ccgccgccgg gatcactctc ggcatggacg agctttacaa gtaagaattc
     3481 tagataactg atcataatca gccataccac atttgtagag gttttacttg ctttaaaaaa
     3541 cctcccacac ctccccctga acctgaaaca taaaatgaat gcaattgttg ttgttaactt
     3601 gtttattgca gcttataatg gttacaaata aagcaatagc atcacaaatt tcacaaataa
     3661 agcatttttt tcactgcatt ctagttgtgg tttgtccaaa ctcatcaatg tatcttattt
     3721 aaattgggcg cgcccaacta gttctagagc ggccgccacc gcggtggagc tccagctttt
     3781 gttcccttta gtgagggtta atttcgagct tggcgtaatc atggtcatag ctgtttcctg
     3841 tgtgaaattg ttatccgctc acaattccac acaacatacg agccggaagc ataaagtgta
     3901 aagcctgggg tgcctaatga gtgagctaac tcacattaat tgcgttgcgc tcactgcccg
     3961 ctttccagtc gggaaacctg tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga
     4021 gaggcggttt gcgtattggg cgctcttccg cttcctcgct cactgactcg ctgcgctcgg
     4081 tcgttcggct gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg ttatccacag
     4141 aatcagggga taacgcagga aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc
     4201 gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac gagcatcaca
     4261 aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga taccaggcgt
     4321 ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt accggatacc
     4381 tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc
     4441 tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc
     4501 ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta agacacgact
     4561 tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat gtaggcggtg
     4621 ctacagagtt cttgaagtgg tggcctaact acggctacac tagaaggaca gtatttggta
     4681 tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct tgatccggca
     4741 aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa
     4801 aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct cagtggaacg
     4861 aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc acctagatcc
     4921 ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa acttggtctg
     4981 acagttacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta tttcgttcat
     5041 ccatagttgc ctgactcccc gtcgtgtaga taactacgat acgggagggc ttaccatctg
     5101 gccccagtgc tgcaatgata ccgcgagacc cacgctcacc ggctccagat ttatcagcaa
     5161 taaaccagcc agccggaagg gccgagcgca gaagtggtcc tgcaacttta tccgcctcca
     5221 tccagtctat taattgttgc cgggaagcta gagtaagtag ttcgccagtt aatagtttgc
     5281 gcaacgttgt tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt
     5341 cattcagctc cggttcccaa cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa
     5401 aagcggttag ctccttcggt cctccgatcg ttgtcagaag taagttggcc gcagtgttat
     5461 cactcatggt tatggcagca ctgcataatt ctcttactgt catgccatcc gtaagatgct
     5521 tttctgtgac tggtgagtac tcaaccaagt cattctgaga atagtgtatg cggcgaccga
     5581 gttgctcttg cccggcgtca atacgggata ataccgcgcc acatagcaga actttaaaag
     5641 tgctcatcat tggaaaacgt tcttcggggc gaaaactctc aaggatctta ccgctgttga
     5701 gatccagttc gatgtaaccc actcgtgcac ccaactgatc ttcagcatct tttactttca
     5761 ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg
     5821 cgacacggaa atgttgaata ctcatactct tcctttttca atattattga agcatttatc
     5881 agggttattg tctcatgagc ggatacatat ttgaatgtat ttagaaaaat aaacaaatag
     5941 gggttccgcg cacatttccc cgaaaagtgc
//
