LOCUS       sequence-200286-        7027 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7027)
  AUTHORS   Cai D, Cohen KB, Luo T, Lichtman JW, Sanes JR
  TITLE     Improved tools for the Brainbow toolbox.
  JOURNAL   Nat Methods. 2013 May 5;10(6):540-7. doi: 10.1038/nmeth.2450. Epub 
            2013 May 5.
  PUBMED    23817127
REFERENCE   2  (bases 1 to 7027)
  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: 
            "10.1038/nmeth.2450"; journalName: "Nat Methods"; date: "2013-05-5- 
            5"; volume: "10"; issue: "6"; pages: "540-7"
COMMENT     Sequence Label: AAV-EF1a-BbTagBY
FEATURES             Location/Qualifiers
     source          1..7027
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   51..191
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   51..180
                     /label=AAV2 ITR
     rep_origin      266..721
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(353..372)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     563..584
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(738..757)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     857..879
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(917..935)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        1003..1107
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1108..1176,1177..1968)
                     /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(1326..1345)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2139..2727
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2628..2647
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     repeat_region   2789..2918
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     promoter        3019..4197
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          3250..4188
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     4145..4165
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     protein_bind    complement(4204..4237)
                     /label=lox71
                     /bound_moiety="Cre recombinase"
                     /note="Left element (LE) mutant of loxP (Araki et al., 
                     2010). Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     regulatory      5008..5017
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     protein_bind    5018..5051
                     /label=loxKR3
                     /bound_moiety="Cre recombinase"
                     /note="Right element (RE) mutant of loxP (Araki et al., 
                     2010). Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     protein_bind    complement(5055..5088)
                     /label=lox66
                     /bound_moiety="Cre recombinase"
                     /note="Right element (RE) mutant of loxP (Araki et al., 
                     2010). Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     CDS             complement(join(5167..5877,5878..5880,5881..5883))
                     /codon_start=1
                     /product="enhanced YFP"
                     /label=EYFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(5209..5230)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     5558..5577
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     5817..5838
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     regulatory      5880..5889
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     protein_bind    5890..5923
                     /label=loxKR3
                     /bound_moiety="Cre recombinase"
                     /note="Right element (RE) mutant of loxP (Araki et al., 
                     2010). Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     misc_feature    5942..6530
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(5995..6015)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(6413..6424)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    join(6562..7027,1..11)
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(6761..6780)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
ORIGIN
        1 tccctgtcct tctgattttg taggtaacca cgtgcggacc gagcggccgc aggaacccct
       61 agtgatggag ttggccactc cctctctgcg cgctcgctcg ctcactgagg ccgggcgacc
      121 aaaggtcgcc cgacgcccgg gctttgcccg ggcggcctca gtgagcgagc gagcgcgcag
      181 ctgcctgcag gggcgcctga tgcggtattt tctccttacg catctgtgcg gtatttcaca
      241 ccgcatacgt caaagcaacc atagtacgcg ccctgtagcg gcgcattaag cgcggcgggt
      301 gtggtggtta cgcgcagcgt gaccgctaca cttgccagcg ccttagcgcc cgctcctttc
      361 gctttcttcc cttcctttct cgccacgttc gccggctttc cccgtcaagc tctaaatcgg
      421 gggctccctt tagggttccg atttagtgct ttacggcacc tcgaccccaa aaaacttgat
      481 ttgggtgatg gttcacgtag tgggccatcg ccctgataga cggtttttcg ccctttgacg
      541 ttggagtcca cgttctttaa tagtggactc ttgttccaaa ctggaacaac actcaactct
      601 atctcgggct attcttttga tttataaggg attttgccga tttcggtcta ttggttaaaa
      661 aatgagctga tttaacaaaa atttaacgcg aattttaaca aaatattaac gtttacaatt
      721 ttatggtgca ctctcagtac aatctgctct gatgccgcat agttaagcca gccccgacac
      781 ccgccaacac ccgctgacgc gccctgacgg gcttgtctgc tcccggcatc cgcttacaga
      841 caagctgtga ccgtctccgg gagctgcatg tgtcagaggt tttcaccgtc atcaccgaaa
      901 cgcgcgagac gaaagggcct cgtgatacgc ctatttttat aggttaatgt catgataata
      961 atggtttctt agacgtcagg tggcactttt cggggaaatg tgcgcggaac ccctatttgt
     1021 ttatttttct aaatacattc aaatatgtat ccgctcatga gacaataacc ctgataaatg
     1081 cttcaataat attgaaaaag gaagagtatg agtattcaac atttccgtgt cgcccttatt
     1141 cccttttttg cggcattttg ccttcctgtt tttgctcacc cagaaacgct ggtgaaagta
     1201 aaagatgctg aagatcagtt gggtgcacga gtgggttaca tcgaactgga tctcaacagc
     1261 ggtaagatcc ttgagagttt tcgccccgaa gaacgttttc caatgatgag cacttttaaa
     1321 gttctgctat gtggcgcggt attatcccgt attgacgccg ggcaagagca actcggtcgc
     1381 cgcatacact attctcagaa tgacttggtt gagtactcac cagtcacaga aaagcatctt
     1441 acggatggca tgacagtaag agaattatgc agtgctgcca taaccatgag tgataacact
     1501 gcggccaact tacttctgac aacgatcgga ggaccgaagg agctaaccgc ttttttgcac
     1561 aacatggggg atcatgtaac tcgccttgat cgttgggaac cggagctgaa tgaagccata
     1621 ccaaacgacg agcgtgacac cacgatgcct gtagcaatgg caacaacgtt gcgcaaacta
     1681 ttaactggcg aactacttac tctagcttcc cggcaacaat taatagactg gatggaggcg
     1741 gataaagttg caggaccact tctgcgctcg gcccttccgg ctggctggtt tattgctgat
     1801 aaatctggag ccggtgagcg tgggtctcgc ggtatcattg cagcactggg gccagatggt
     1861 aagccctccc gtatcgtagt tatctacacg acggggagtc aggcaactat ggatgaacga
     1921 aatagacaga tcgctgagat aggtgcctca ctgattaagc attggtaact gtcagaccaa
     1981 gtttactcat atatacttta gattgattta aaacttcatt tttaatttaa aaggatctag
     2041 gtgaagatcc tttttgataa tctcatgacc aaaatccctt aacgtgagtt ttcgttccac
     2101 tgagcgtcag accccgtaga aaagatcaaa ggatcttctt gagatccttt ttttctgcgc
     2161 gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg tttgccggat
     2221 caagagctac caactctttt tccgaaggta actggcttca gcagagcgca gataccaaat
     2281 actgttcttc tagtgtagcc gtagttaggc caccacttca agaactctgt agcaccgcct
     2341 acatacctcg ctctgctaat cctgttacca gtggctgctg ccagtggcga taagtcgtgt
     2401 cttaccgggt tggactcaag acgatagtta ccggataagg cgcagcggtc gggctgaacg
     2461 gggggttcgt gcacacagcc cagcttggag cgaacgacct acaccgaact gagataccta
     2521 cagcgtgagc tatgagaaag cgccacgctt cccgaaggga gaaaggcgga caggtatccg
     2581 gtaagcggca gggtcggaac aggagagcgc acgagggagc ttccaggggg aaacgcctgg
     2641 tatctttata gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc
     2701 tcgtcagggg ggcggagcct atggaaaaac gccagcaacg cggccttttt acggttcctg
     2761 gccttttgct ggccttttgc tcacatgtcc tgcaggcagc tgcgcgctcg ctcgctcact
     2821 gaggccgccc gggcgtcggg cgacctttgg tcgcccggcc tcagtgagcg agcgagcgcg
     2881 cagagaggga gtggccaact ccatcactag gggttcctgc ggccgcacgc gtaagctttg
     2941 caaagatgga taaagtttta aacagagagg aatctttgca gctaatggac cttctaggtc
     3001 ttgaaaggag tgggaattgg ctccggtgcc cgtcagtggg cagagcgcac atcgcccaca
     3061 gtccccgaga agttgggggg aggggtcggc aattgaaccg gtgcctagag aaggtggcgc
     3121 ggggtaaact gggaaagtga tgtcgtgtac tggctccgcc tttttcccga gggtggggga
     3181 gaaccgtata taagtgcagt agtcgccgtg aacgttcttt ttcgcaacgg gtttgccgcc
     3241 agaacacagg taagtgccgt gtgtggttcc cgcgggcctg gcctctttac gggttatggc
     3301 ccttgcgtgc cttgaattac ttccacctgg ctgcagtacg tgattcttga tcccgagctt
     3361 cgggttggaa gtgggtggga gagttcgagg ccttgcgctt aaggagcccc ttcgcctcgt
     3421 gcttgagttg aggcctggcc tgggcgctgg ggccgccgcg tgcgaatctg gtggcacctt
     3481 cgcgcctgtc tcgctgcttt cgataagtct ctagccattt aaaatttttg atgacctgct
     3541 gcgacgcttt ttttctggca agatagtctt gtaaatgcgg gccaagatct gcacactggt
     3601 atttcggttt ttggggccgc gggcggcgac ggggcccgtg cgtcccagcg cacatgttcg
     3661 gcgaggcggg gcctgcgagc gcggccaccg agaatcggac gggggtagtc tcaagctggc
     3721 cggcctgctc tggtgcctgg cctcgcgccg ccgtgtatcg ccccgccctg ggcggcaagg
     3781 ctggcccggt cggcaccagt tgcgtgagcg gaaagatggc cgcttcccgg ccctgctgca
     3841 gggagctcaa aatggaggac gcggcgctcg ggagagcggg cgggtgagtc acccacacaa
     3901 aggaaaaggg cctttccgtc ctcagccgtc gcttcatgtg actccacgga gtaccgggcg
     3961 ccgtccaggc acctcgatta gttctcgagc ttttggagta cgtcgtcttt aggttggggg
     4021 gaggggtttt atgcgatgga gtttccccac actgagtggg tggagactga agttaggcca
     4081 gcttggcact tgatgtaatt ctccttggaa tttgcccttt ttgagtttgg atcttggttc
     4141 attctcaagc ctcagacagt ggttcaaagt ttttttcttc catttcaggt gtcgtgaggt
     4201 acctaccgtt cgtatagcat acattatacg aagttattta ggagagcaca cacttgcagc
     4261 tcatgcagcc ggggccactc tcatcaggag ggttcagctt actcacgcgt gcacttctgt
     4321 gccccagttt gctagggagg tcgcagtatc tggccactgc cacctcgtgc tgctcgacgt
     4381 aggtctcgtt gttggcctcc ttgattcttt ccagtctgta gtccacatag tagacgccag
     4441 gcatcttgag gttcttagcg ggtttcttgg atctatatgt ggtcttgatg tttgcgatca
     4501 gatggctccc gcccacgagc ttcagggcca tgtcgtttct gccttccagg ccgccgtcag
     4561 cggggtacag cgtctcggtg aaggcctccc agccgagtgt tttcttctgc atcacagggc
     4621 cgttggatgt gaagttcacc cctctgatct tgacgttgta gatgaggcag ccgtcctgga
     4681 ggctggtgtc ctgggtagcg gtcagcacgc ccccgtcttc gtatgtggtg actctctccc
     4741 atgtgaagcc ctcagggaag gactgcttga agaagtcggg gatgccctgg gtgtggttga
     4801 tgaaggtctt gctgccgtag aggaagctag tagccaggat gtcgaaggcg aaggggagag
     4861 ggccgccctc gaccaccttg attctcatgg tctgggtgcc ctcgtagggc ttgccttcgc
     4921 cctcggatgt gcacttgaag tgatggttgt ccacggtgcc ctccatgtac agcttcatgt
     4981 gcatgttctc cttaatcagc tcgctcacca tggtggcata acaaggtata atgtatgcta
     5041 tacgaagtta tgatataact tcgtatagca tacattatac gaacggtatc aggagagcac
     5101 acacttgcag ctcatgcagc cggggccact ctcatcagga gggttcagct tactcacgcg
     5161 tgcactcttg tacagctcgt ccatgccgag agtgatcccg gcggcggtca cgaactccag
     5221 caggaccatg tgatcgcgct tctcgttggg gtctttgctc agggcggact ggtagctcag
     5281 gtagtggttg tcgggcagca gcacggggcc gtcgccgatg ggggtgttct gctggtagtg
     5341 gtcggcgagc tgcacgctgc cgtcctcgat gttgtggcgg atcttgaagt tcaccttgat
     5401 gccgttcttc tgcttgtcgg ccatgatata gacgttgtgg ctgttgtagt tgtactccag
     5461 cttgtgcccc aggatgttgc cgtcctcctt gaagtcgatg cccttcagct cgatgcggtt
     5521 caccagggtg tcgccctcga acttcacctc ggcgcgggtc ttgtagttgc cgtcgtcctt
     5581 gaagaagatg gtgcgctcct ggacgtagcc ttcgggcatg gcggacttga agaagtcgtg
     5641 ctgcttcatg tggtcggggt agcgggcgaa gcactgcagg ccgtagccga aggtggtcac
     5701 gagggtgggc cagggcacgg gcagcttgcc ggtggtgcag atgaacttca gggtcagctt
     5761 gccgtaggtg gcatcgccct cgccctcgcc ggacacgctg aacttgtggc cgtttacgtc
     5821 gccgtccagc tcgaccagga tgggcaccac cccggtgaac agctcctcgc ccttgctcac
     5881 catggtggca taacaaggta taatgtatgc tatacgaagt tatgatatca agcttatcga
     5941 taatcaacct ctggattaca aaatttgtga aagattgact ggtattctta actatgttgc
     6001 tccttttacg ctatgtggat acgctgcttt aatgcctttg tatcatgcta ttgcttcccg
     6061 tatggctttc attttctcct ccttgtataa atcctggttg ctgtctcttt atgaggagtt
     6121 gtggcccgtt gtcaggcaac gtggcgtggt gtgcactgtg tttgctgacg caacccccac
     6181 tggttggggc attgccacca cctgtcagct cctttccggg actttcgctt tccccctccc
     6241 tattgccacg gcggaactca tcgccgcctg ccttgcccgc tgctggacag gggctcggct
     6301 gttgggcact gacaattccg tggtgttgtc ggggaaatca tcgtcctttc cttggctgct
     6361 cgcctatgtt gccacctgga ttctgcgcgg gacgtccttc tgctacgtcc cttcggccct
     6421 caatccagcg gaccttcctt cccgcggcct gctgccggct ctgcggcctc ttccgcgtct
     6481 tcgccttcgc cctcagacga gtcggatctc cctttgggcc gcctccccgc atcgataccg
     6541 agcgctgctc gagagatcta cgggtggcat ccctgtgacc cctccccagt gcctctcctg
     6601 gccctggaag ttgccactcc agtgcccacc agccttgtcc taataaaatt aagttgcatc
     6661 attttgtctg actaggtgtc cttctataat attatggggt ggaggggggt ggtatggagc
     6721 aaggggcaag ttgggaagac aacctgtagg gcctgcgggg tctattggga accaagctgg
     6781 agtgcagtgg cacaatcttg gctcactgca atctccgcct cctgggttca agcgattctc
     6841 ctgcctcagc ctcccgagtt gttgggattc caggcatgca tgaccaggct cagctaattt
     6901 ttgttttttt ggtagagacg gggtttcacc atattggcca ggctggtctc caactcctaa
     6961 tctcaggtga tctacccacc ttggcctccc aaattgctgg gattacaggc gtgaaccact
     7021 gctccct
//
