LOCUS       sequence-324720-        6486 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  To generate a stable acceptor cell line with a BxbI attP site at the
            AASV1 locus, between an exogenous human pRPL30 promoter and a 
            tripartite V5/SNAPtag/Blast reporter..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6486)
  TITLE     Huang Lab Starport System Plasmids
REFERENCE   2  (bases 1 to 6486)
  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"
COMMENT     Sequence Label: StarPort01
FEATURES             Location/Qualifiers
     source          1..6486
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_feature    412..913
                     /label=HA-L
                     /note="left homology arm from the adeno-associated virus 
                     integration site (AAVS1) within intron 1 of the human 
                     PPP1R12C gene"
     protein_bind    976..1009
                     /label=FRT (minimal)
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="supports FLP-mediated excision but not integration 
                     (Turan and Bode, 2011)"
     protein_bind    1627..1660
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     protein_bind    1860..1907
                     /label=Bxb1 attP
                     /bound_moiety="Mycobacteriophage Bxb1 integrase"
                     /note="Minimal attP site for the Mycopbacteriophage Bxb1 
                     integrase (Voutev and Mann, 2017)"
     CDS             2079..2120
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     CDS             2121..2663
                     /codon_start=1
                     /product="human O6-alkylguanine-DNA-alkyltransferase 
                     variant that forms covalent bonds with benzylguanine 
                     derivatives"
                     /label=SNAP-tag(R)
                     /note="fast-reacting variant, optimized for mammalian 
                     expression"
                     /translation="DKDCEMKRTTLDSPLGKLELSGCEQGLHRIIFLGKGTSAADAVEV
                     PAPAAVLGGPEPLMQATAWLNAYFHQPEAIEEFPVPALHHPVFQQESFTRQVLWKLLKV
                     VKFGEVISYSHLAALAGNPAATAAVKTALSGNPVPILIPCHRVVQGDLDVGGYEGGLAV
                     KEWLLAHEGHRLGKPGLG"
     CDS             2835..2888
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     CDS             2895..3290
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
                     VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
                     KAIVKDSDGQPTAVGIRELLPSGYVWEG"
     polyA_signal    3301..3435
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3325..3344)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     3379..3398
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     protein_bind    3733..3766
                     /label=FRT (minimal)
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="supports FLP-mediated excision but not integration 
                     (Turan and Bode, 2011)"
     misc_feature    3938..4467
                     /label=HA-R
                     /note="right homology arm from the adeno-associated virus 
                     integration site (AAVS1) within intron 1 of the human 
                     PPP1R12C gene"
     CDS             complement(4764..5579)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             complement(4764..5579)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     5487..5506
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     5571..5590
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      5804..6392
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6293..6312
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 accgtgcggc cgccaatcca caaccagtaa gggctggccc aggtgtctgg ccccacagct
       61 gagctcctaa ccgcaccgag cctctccaaa acatggtctt tttttgctcc ccactgccac
      121 caccatcacc gtcccctgcc tcatccagtc ccccttccca ttctccagag acttcctggg
      181 ccatctccat gaccttccag accagcctgg gcttcactct caaatccaat gctttctctg
      241 acctgcattc tctcccctgg gcctgtgtcg ctttctgtct gcagcttgtg gcctgggtca
      301 cctctacggc tggcccagat ccttccctgc cgcctccttc aggttccgtc ttcctccact
      361 ccctcttccc cttgctctct gctgtgttgc tgcccaagga tgctctttcc ggagcacttc
      421 cttctcggcg ctgcaccacg tgatgtcctc tgagcggatc ctccccgtgt ctgggtcctc
      481 tccgggcatc tctcctccct cacccaaccc catgccgtct tcactcgctg ggttcccttt
      541 tccttctcct tctggggcct gtgccatctc tcgtttctta ggatggcctt ctccgacgga
      601 tgtctccctt gcgtcccgcc tccccttctt gtaggcctgc atcatcaccg tttttctgga
      661 caaccccaaa gtaccccgtc tgcctggctt tagccacctc tccatcctct tgctttcttt
      721 gcctggacac cccgttctcc tgtggattcg ggtcacctct cactcctttc atttgggcag
      781 ctcccctacc ccccttacct ctctagtctg tgctagctct tccagccccc tgtcatggca
      841 tcttccaggg gtccgagagc tcagctagtc ttcttcctcc aacccgggcc cctatgtcca
      901 cttcaggaca gcatgtttgc tgcctccagg gatcctgtgt ccccgagctg ggaccacctt
      961 atattcccag gccctgaagt tcctattctc tagaaagtat aggaacttcc tgaactggcc
     1021 gataattgca gacgatcgaa ttcgagttgg cgcgcctgtc attctaaatc tctctttcag
     1081 cctaaagctt tttccccgta tccccccagg tgtctgcagg ctcaaagagc agcgagaagc
     1141 gttcagagga aagcgatccc gtgccacctt ccccgtgccc gggctgtccc cgcacgctgc
     1201 cggctcgggg atgcgggggg agcgccggac cggagcggag ccccgggcgg ctcgctgctg
     1261 ccccctagcg ggggagggac gtaattacat ccctgggggc tttgggaggg ggctgtcccc
     1321 gtgagctccc aggcgcgcct gtcattctaa atctctcttt cagcctaaag ctttttcccc
     1381 gtatcccccc aggtgtctgc aggctcaaag agcagcgaga agcgttcaga ggaaagcgat
     1441 cccgtgccac cttccccgtg cccgggctgt ccccgcacgc tgccggctcg gggatgcggg
     1501 gggagcgccg gaccggagcg gagccccggg cggctcgctg ctgcccccta gcgggggagg
     1561 gacgtaatta catccctggg ggctttgggg gggggctgtc cccgtgagct caataaaaga
     1621 gcccacataa cttcgtataa tgtatgctat acgaagttat aacgccccgc agccattcta
     1681 gctagcggta ccaatagcaa ccggcagctg ccctccgctt ttgctccgcc ccttctgctt
     1741 gcgatctgtt tccgcttccg gtcccgcagt tccggctctg ccgtgaagag ctttgcattg
     1801 tgggaagtct ttcctttctc gttccccggc catcttagcg gctgctgttg gtgagtggtg
     1861 gtttgtctgg tcaaccaccg cggtctcagt ggtgtacggt acaaaccagg agctttgcgg
     1921 gttgccattt gtaactccgg atcctaaaat tcctgtcctg ttctctgtct cttctaggtt
     1981 gggggccgtc ccgctcctaa ggcaggaata tgatggagac tgacaccctc ttgctctggg
     2041 tcctgctgct gtgggttccg ggtagtaccg gtgacatggg taagcctatc cctaaccctc
     2101 tcctcggtct ggattctacg gacaaagact gcgaaatgaa gcgcaccacc ctggatagcc
     2161 ctctgggcaa gctggaactg tctgggtgcg aacagggcct gcaccgtatc atcttcctgg
     2221 gcaaaggaac atctgccgcc gacgccgtgg aagtgcctgc cccagccgcc gtgctgggcg
     2281 gaccagagcc actgatgcag gccaccgcct ggctcaacgc ctactttcac cagcctgagg
     2341 ccatcgagga gttccctgtg ccagccctgc accacccagt gttccagcag gagagcttta
     2401 cccgccaggt gctgtggaaa ctgctgaaag tggtgaagtt cggagaggtc atcagctaca
     2461 gccacctggc cgccctggcc ggcaatcccg ccgccaccgc cgccgtgaaa accgccctga
     2521 gcggaaatcc cgtgcccatt ctgatcccct gccaccgggt ggtgcagggc gacctggacg
     2581 tggggggcta cgagggcggg ctcgccgtga aagagtggct gctggcccac gagggccaca
     2641 gactgggcaa gcctgggctg ggtggttcta acgccgtggg ccaggatacg caggaggtga
     2701 tcgtcgtgcc ccacagcttg cccttcaagg tcgtggttat tagtgctatc ctggcgttgg
     2761 tggtgctgac tattatctcc cttatcatac ttatcatgct gtggcaaaaa aaaccccgag
     2821 gatccggaag cggagagggc agaggaagtc tgctaacatg cggtgacgtc gaggagaatc
     2881 ctggacctgg atccatggcc aagcctttgt ctcaagaaga atccaccctc attgaaagag
     2941 caacggctac aatcaacagc atccccatct ctgaagacta cagcgtcgcc agcgcagctc
     3001 tctctagcga cggccgcatc ttcactggtg tcaatgtata tcattttact gggggacctt
     3061 gtgcagaact cgtggtgctg ggcactgctg ctgctgcggc agctggcaac ctgacttgta
     3121 tcgtcgcgat cggaaatgag aacaggggca tcttgagccc ctgcggacgg tgccgacagg
     3181 tgcttctcga tctgcatcct gggatcaaag ccatagtgaa ggacagtgat ggacagccga
     3241 cggcagttgg gattcgtgaa ttgctgccct ctggttatgt gtgggagggc tggatcctaa
     3301 gatccagaca tgataagata cattgatgag tttggacaaa ccacaactag aatgcagtga
     3361 aaaaaatgct ttatttgtga aatttgtgat gctattgctt tatttgtaac cattataagc
     3421 tgcaataaac aagttaacaa caacaattgc attcatttta tgtttcaggt tcagggggag
     3481 gtgtgggagg ttttttaaag caagtaaaac ctctacaaat gtggtatggc tgattatgat
     3541 cgctgctgaa caggttgact attaattgtg tctgcttgat gtggacacca ggtggcgctg
     3601 gacatcagat ttggagaggc agttgtctag ggaaccgggc tctgtgccag cgcaggaggc
     3661 aggctggctc tcctattcca gggatgctca tccaggaagg aaaggttgca tgctggacac
     3721 actaacctta cagaagttcc tattctctag aaagtatagg aacttcgagt gattggtgac
     3781 agaaaagccc catccttagg cctcctcctt cctagtctcc tgatattggg tctaaccccc
     3841 acctcctgtt aggcagattc cttatctggt gacacacccc catttcctgg agccatctct
     3901 ctccttgcca gaacctctaa ggtttgctta cgatggagcc agagaggatc ctgggaggga
     3961 gagcttggca gggggtggga gggaaggggg ggatgcgtga cctgcccggt tctcagtggc
     4021 caccctgcgc taccctctcc cagaacctga gctgctctga cgcggccgtc tggtgcgttt
     4081 cactgatcct ggtgctgcag cttccttaca cttcccaaga ggagaagcag tttggaaaaa
     4141 caaaatcaga ataagttggt cctgagttct aactttggct cttcaccttt ctagtcccca
     4201 atttatattg ttcctccgtg cgtcagtttt acctgtgaga taaggccagt agccagcccc
     4261 gtcctggcag ggctgtggtg aggagggggg tgtccgtgtg gaaaactccc tttgtgagaa
     4321 tggtgcgtcc taggtgttca ccaggtcgtg gccgcctcta ctccctttct ctttctccat
     4381 ccttctttcc ttaaagagtc cccagtgcta tctgggacat attcctccgc ccagagcagg
     4441 gtcccgcttc cctaaggccc tgctctgggc ttctgggttt gagtccttgg caagcccagg
     4501 agaggcgctc aggcttccct gtcccccttc ctcgtccacc atctcatgcc cctggctctc
     4561 ctgccccttc cctacagggg ttcctggctc tgctctacct actgcattaa aaaaaaattt
     4621 ttttttttct aagacaaggt ctgactctgt ctcccaggct ggagtgcagt ggtgcaatca
     4681 cgctcatagc ctcaacctcc cgggctccag tgggcctccc acctccgagc ggccgcgtgt
     4741 tacaaccaat taaccaattc tgattagaaa aactcatcga gcatcaaatg aaactgcaat
     4801 ttattcatat caggattatc aataccatat ttttgaaaaa gccgtttctg taatgaagga
     4861 gaaaactcac cgaggcagtt ccataggatg gcaagatcct ggtatcggtc tgcgattccg
     4921 actcgtccaa catcaataca acctattaat ttcccctcgt caaaaataag gttatcaagt
     4981 gagaaatcac catgagtgac gactgaatcc ggtgagaatg gcaaaagctt atgcatttct
     5041 ttccagactt gttcaacagg ccagccatta cgctcgtcat caaaatcact cgcatcaacc
     5101 aaaccgttat tcattcgtga ttgcgcctga gcgaggcgaa atacgcgatc gctgttaaaa
     5161 ggacaattac aaacaggaat cgaatgcaac cggcgcagga acactgccag cgcatcaaca
     5221 atattttcac ctgaatcagg atattcttct aatacctgga atgctgtttt cccggggatc
     5281 gcagtggtga gtaaccatgc atcatcagga gtacggataa aatgcttgat ggtcggaaga
     5341 ggcataaatt ccgtcagcca gtttagtctg accatctcat ctgtaacatc attggcaacg
     5401 ctacctttgc catgtttcag aaacaactct ggcgcatcgg gcttcccata caatcgatag
     5461 attgtcgcac ctgattgccc gacattatcg cgagcccatt tatacccata taaatcagca
     5521 tccatgttgg aatttaatcg cggcctggag caagacgttt cccgttgaat atggctcata
     5581 acaccccttg tattactgtt tatgtaagca gacagtttta ttgttcatga tgatatattt
     5641 ttatcttgtg caatgtaaca tcagagattt tgagacacaa cgtggctttg ttgaataaat
     5701 cgaacttttg ctgagttgaa ggatcagtca tgaccaaaat cccttaacgt gagttttcgt
     5761 tccactgagc gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat cctttttttc
     5821 tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct accagcggtg gtttgtttgc
     5881 cggatcaaga gctaccaact ctttttccga aggtaactgg cttcagcaga gcgcagatac
     5941 caaatactgt tcttctagtg tagccgtagt taggccacca cttcaagaac tctgtagcac
     6001 cgcctacata cctcgctctg ctaatcctgt taccagtggc tgctgccagt ggcgataagt
     6061 cgtgtcttac cgggttggac tcaagacgat agttaccgga taaggcgcag cggtcgggct
     6121 gaacgggggg gtcgtgcaca cagcccagct tggagcgaac gacctacacc gaactgagat
     6181 acctacagcg tgagctatga gaaagcgcca cgcttcccga agggagaaag gcggacaggt
     6241 atccggtaag cggcagggtc ggaacaggag agcgcacgag ggagcttcca gggggaaacg
     6301 cctggtatct ttatagtcct gtcgggtttc gccacctctg acttgagcgt cgatttttgt
     6361 gatgctcgtc aggggggcgg agcctatgga aaaacgccag caacgcggcc tttttacggt
     6421 tcctggcctt ttgctggcct tttgctcaca tgttctttcc tgcgttatcc cctgattctg
     6481 tggata
//
