LOCUS       sequence-319378-        6381 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 6381)
  TITLE     Huang Lab Starport System Plasmids
REFERENCE   2  (bases 1 to 6381)
  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..6381
                     /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"
     primer_bind     complement(3331..3350)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     3385..3404
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     protein_bind    3628..3661
                     /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    3833..4362
                     /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(4659..5474)
                     /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(4659..5474)
                     /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     5382..5401
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     5466..5485
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      5699..6287
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6188..6207
                     /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 gcctgtgccg 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 ctcgaggatc cagacatgat aagatacatt gatgagtttg gacaaaccac aactagaatg
     3361 cagtgaaaaa aatgctttat ttgtgaaatt tgtgatgcta ttgctttatt tgtaaccatt
     3421 ataagctgca ataaacgctg ctgaacaggt tgactattaa ttgtgtctgc ttgatgtgga
     3481 caccaggtgg cgctggacat cagatttgga gaggcagttg tctagggaac cgggctctgt
     3541 gccagcgcag gaggcaggct ggctctccta ttccagggat gctcatccag gaaggaaagg
     3601 ttgcatgctg gacacactaa ccttacagaa gttcctattc tctagaaagt ataggaactt
     3661 cgagtgattg gtgacagaaa agccccatcc ttaggcctcc tccttcctag tctcctgata
     3721 ttgggtctaa cccccacctc ctgttaggca gattccttat ctggtgacac acccccattt
     3781 cctggagcca tctctctcct tgccagaacc tctaaggttt gcttacgatg gagccagaga
     3841 ggatcctggg agggagagct tggcaggggg tgggagggaa gggggggatg cgtgacctgc
     3901 ccggttctca gtggccaccc tgcgctaccc tctcccagaa cctgagctgc tctgacgcgg
     3961 ccgtctggtg cgtttcactg atcctggtgc tgcagcttcc ttacacttcc caagaggaga
     4021 agcagtttgg aaaaacaaaa tcagaataag ttggtcctga gttctaactt tggctcttca
     4081 cctttctagt ccccaattta tattgttcct ccgtgcgtca gttttacctg tgagataagg
     4141 ccagtagcca gccccgtcct ggcagggctg tggtgaggag gggggtgtcc gtgtggaaaa
     4201 ctccctttgt gagaatggtg cgtcctaggt gttcaccagg tcgtggccgc ctctactccc
     4261 tttctctttc tccatccttc tttccttaaa gagtccccag tgctatctgg gacatattcc
     4321 tccgcccaga gcagggtccc gcttccctaa ggccctgctc tgggcttctg ggtttgagtc
     4381 cttggcaagc ccaggagagg cgctcaggct tccctgtccc ccttcctcgt ccaccatctc
     4441 atgcccctgg ctctcctgcc ccttccctac aggggttcct ggctctgctc tacctactgc
     4501 attaaaaaaa aatttttttt tttctaagac aaggtctgac tctgtctccc aggctggagt
     4561 gcagtggtgc aatcacgctc atagcctcaa cctcccgggc tccagtgggc ctcccacctc
     4621 cgagcggccg cgtgttacaa ccaattaacc aattctgatt agaaaaactc atcgagcatc
     4681 aaatgaaact gcaatttatt catatcagga ttatcaatac catatttttg aaaaagccgt
     4741 ttctgtaatg aaggagaaaa ctcaccgagg cagttccata ggatggcaag atcctggtat
     4801 cggtctgcga ttccgactcg tccaacatca atacaaccta ttaatttccc ctcgtcaaaa
     4861 ataaggttat caagtgagaa atcaccatga gtgacgactg aatccggtga gaatggcaaa
     4921 agcttatgca tttctttcca gacttgttca acaggccagc cattacgctc gtcatcaaaa
     4981 tcactcgcat caaccaaacc gttattcatt cgtgattgcg cctgagcgag gcgaaatacg
     5041 cgatcgctgt taaaaggaca attacaaaca ggaatcgaat gcaaccggcg caggaacact
     5101 gccagcgcat caacaatatt ttcacctgaa tcaggatatt cttctaatac ctggaatgct
     5161 gttttcccgg ggatcgcagt ggtgagtaac catgcatcat caggagtacg gataaaatgc
     5221 ttgatggtcg gaagaggcat aaattccgtc agccagttta gtctgaccat ctcatctgta
     5281 acatcattgg caacgctacc tttgccatgt ttcagaaaca actctggcgc atcgggcttc
     5341 ccatacaatc gatagattgt cgcacctgat tgcccgacat tatcgcgagc ccatttatac
     5401 ccatataaat cagcatccat gttggaattt aatcgcggcc tggagcaaga cgtttcccgt
     5461 tgaatatggc tcataacacc ccttgtatta ctgtttatgt aagcagacag ttttattgtt
     5521 catgatgata tatttttatc ttgtgcaatg taacatcaga gattttgaga cacaacgtgg
     5581 ctttgttgaa taaatcgaac ttttgctgag ttgaaggatc agtcatgacc aaaatccctt
     5641 aacgtgagtt ttcgttccac tgagcgtcag accccgtaga aaagatcaaa ggatcttctt
     5701 gagatccttt ttttctgcgc gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag
     5761 cggtggtttg tttgccggat caagagctac caactctttt tccgaaggta actggcttca
     5821 gcagagcgca gataccaaat actgttcttc tagtgtagcc gtagttaggc caccacttca
     5881 agaactctgt agcaccgcct acatacctcg ctctgctaat cctgttacca gtggctgctg
     5941 ccagtggcga taagtcgtgt cttaccgggt tggactcaag acgatagtta ccggataagg
     6001 cgcagcggtc gggctgaacg gggggttcgt gcacacagcc cagcttggag cgaacgacct
     6061 acaccgaact gagataccta cagcgtgagc tatgagaaag cgccacgctt cccgaaggga
     6121 gaaaggcgga caggtatccg gtaagcggca gggtcggaac aggagagcgc acgagggagc
     6181 ttccaggggg aaacgcctgg tatctttata gtcctgtcgg gtttcgccac ctctgacttg
     6241 agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct atggaaaaac gccagcaacg
     6301 cggccttttt acggttcctg gccttttgct ggccttttgc tcacatgttc tttcctgcgt
     6361 tatcccctga ttctgtggat a
//
