LOCUS       sequence-181812-        6455 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Can be used to generate AAV virus that will express mRuby3 in the 
            presence of Cre.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6455)
  AUTHORS   Rylan Larsen
  TITLE     AAV red fluorescent protein plasmids
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 6455)
  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: 
            "Unpublished"
COMMENT     Sequence Label: AAV pCAG-FLEX-mRuby3-WPRE
FEATURES             Location/Qualifiers
     source          1..6455
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..141
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   12..141
                     /label=AAV2 ITR
     enhancer        234..537
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        539..816
                     /label=chicken beta-actin promoter
     intron          818..1835
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     1759..1781
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1843..1862
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     protein_bind    complement(1939..1972)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     protein_bind    complement(1983..2016)
                     /label=lox2272
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (AAGTATCC) (Shaw et al., 2021). lox2272 sites are 
                     compatible with each other, but incompatible with loxP or 
                     loxN sites (Lee and Saito, 1988)."
     CDS             complement(2030..2740)
                     /codon_start=1
                     /product="monomeric red fluorescent protein derived from 
                     mRuby2, with improved brightness and photostability (Bajar 
                     et al., 2016)"
                     /label=mRuby3
                     /translation="MVSKGEELIKENMRMKVVMEGSVNGHQFKCTGEGEGRPYEGVQTM
                     RIKVIEGGPLPFAFDILATSFMYGSRTFIKYPADIPDFFKQSFPEGFTWERVTRYEDGG
                     VVTVTQDTSLEDGELVYNVKVRGVNFPSNGPVMQKKTKGWEPNTEMMYPADGGLRGYTD
                     IALKVDGGGHLHCNFVTTYRSKKTVGNIKMPGVHAVDHRLERIEESDNETYVVQREVAV
                     AKYSNLGGGMDELYK"
     protein_bind    2758..2791
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     protein_bind    2802..2835
                     /label=lox2272
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (AAGTATCC) (Shaw et al., 2021). lox2272 sites are 
                     compatible with each other, but incompatible with loxP or 
                     loxN sites (Lee and Saito, 1988)."
     misc_feature    2859..3447
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(2912..2932)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3330..3341)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(3482..3499)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3488..3695
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   3718..3858
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   3718..3847
                     /label=AAV2 ITR
     rep_origin      3933..4388
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4020..4039)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4230..4251
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4405..4424)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4524..4546
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4584..4602)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4670..4774
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4775..4843,4844..5635)
                     /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(4993..5012)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5806..6394
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6295..6314
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cctgcaggca gctgcgcgct cgctcgctca ctgaggccgc ccgggcaaag cccgggcgtc
       61 gggcgacctt tggtcgcccg gcctcagtga gcgagcgagc gcgcagagag ggagtggcca
      121 actccatcac taggggttcc tgcggccgca cgcgaaacaa ttctgcagga atctagttat
      181 taatagtaat caattacggg gtcattagtt catagcccat atatggagtt ccgcgttaca
      241 taacttacgg taaatggccc gcctggctga ccgcccaacg acccccgccc attgacgtca
      301 ataatgacgt atgttcccat agtaacgcca atagggactt tccattgacg tcaatgggtg
      361 gagtatttac ggtaaactgc ccacttggca gtacatcaag tgtatcatat gccaagtacg
      421 ccccctattg acgtcaatga cggtaaatgg cccgcctggc attatgccca gtacatgacc
      481 ttatgggact ttcctacttg gcagtacatc tacgtattag tcatcgctat taccatggtc
      541 gaggtgagcc ccacgttctg cttcactctc cccatctccc ccccctcccc acccccaatt
      601 ttgtatttat ttatttttta attattttgt gcagcgatgg gggcgggggg gggggggggg
      661 cgcgcgccag gcggggcggg gcggggcgag gggcggggcg gggcgaggcg gagaggtgcg
      721 gcggcagcca atcagagcgg cgcgctccga aagtttcctt ttatggcgag gcggcggcgg
      781 cggcggccct ataaaaagcg aagcgcgcgg cgggcgggga gtcgctgcga cgctgccttc
      841 gccccgtgcc ccgctccgcc gccgcctcgc gccgcccgcc ccggctctga ctgaccgcgt
      901 tactcccaca ggtgagcggg cgggacggcc cttctcctcc gggctgtaat tagcgcttgg
      961 tttaatgacg gcttgtttct tttctgtggc tgcgtgaaag ccttgagggg ctccgggagg
     1021 gccctttgtg cggggggagc ggctcggggg gtgcgtgcgt gtgtgtgtgc gtggggagcg
     1081 ccgcgtgcgg ctccgcgctg cccggcggct gtgagcgctg cgggcgcggc gcggggcttt
     1141 gtgcgctccg cagtgtgcgc gaggggagcg cggccggggg cggtgccccg cggtgcgggg
     1201 ggggctgcga ggggaacaaa ggctgcgtgc ggggtgtgtg cgtggggggg tgagcagggg
     1261 gtgtgggcgc gtcggtcggg ctgcaacccc ccctgcaccc ccctccccga gttgctgagc
     1321 acggcccggc ttcgggtgcg gggctccgta cggggcgtgg cgcggggctc gccgtgccgg
     1381 gcggggggtg gcggcaggtg ggggtgccgg gcggggcggg gccgcctcgg gccggggagg
     1441 gctcggggga ggggcgcggc ggcccccgga gcgccggcgg ctgtcgaggc gcggcgagcc
     1501 gcagccattg ccttttatgg taatcgtgcg agagggcgca gggacttcct ttgtcccaaa
     1561 tctgtgcgga gccgaaatct gggaggcgcc gccgcacccc ctctagcggg cgcggggcga
     1621 agcggtgcgg cgccggcagg aaggaaatgg gcggggaggg ccttcgtgcg tcgccgcgcc
     1681 gccgtcccct tctccctctc cagcctcggg gctgtccgcg gggggacggc tgccttcggg
     1741 ggggacgggg cagggcgggg ttcggcttct ggcgtgtgac cggcggctct agagcctctg
     1801 ctaaccatgt tcatgccttc ttctttttcc tacagctcct gggcaacgtg ctggttattg
     1861 tgctgtctca tcattttggc aaagaattga tttgataccg cgggccccgc gtggacggta
     1921 cccttcaggt cgacgtcgat aacttcgtat agcatacatt atacgaagtt atccgaatcg
     1981 caataacttc gtataggata ctttatacga agttatttgg cgcgcctgac ttgtacagct
     2041 cgtccatgcc accaccaagg ttgctgtatt tggcaactgc cacttctctt tgcactacgt
     2101 aggtttcatt gtcactctcc tcgatccttt ccaggcggtg atcaacggca tggacaccgg
     2161 gcatcttgat gttcccgacg gtcttttttg acctgtaagt tgtcacgaag ttgcagtgca
     2221 gatggccacc accatcaact ttcagtgcga tgtcagtgta tcctctcaga ccaccatctg
     2281 ctggatacat catctctgta ttaggctccc aacccttggt cttcttctgc atcacgggac
     2341 cattggaggg aaagtttacc cctctgacct tgacgttgta gacgagctcg ccatcctcaa
     2401 ggctggtgtc ctgcgtgacg gtgacgactc caccatcttc gtatctcgta actctttccc
     2461 aagtaaaacc ctcaggaaag gactgtttaa agaaatcagg gatgtcggcc gggtacttga
     2521 taaaggtacg gctgccatac atgaacgacg tggcaagaat gtcaaaggca aatggcaggg
     2581 gtcctccctc gatgactttg atcctcatgg tttgcactcc ctcgtacggt ctgccttctc
     2641 cttcacctgt gcatttgaat tggtggccgt tgaccgaacc ttccatgacc accttcatac
     2701 gcatattttc cttgatcagc tcttcgccct tagacaccat gccaccggcc ggccttgata
     2761 acttcgtata atgtatgcta tacgaagtta tccgaatcgc aataacttcg tataaagtat
     2821 cctatacgaa gttatcgaag atcatcaagc ttatcgataa tcaacctctg gattacaaaa
     2881 tttgtgaaag attgactggt attcttaact atgttgctcc ttttacgcta tgtggatacg
     2941 ctgctttaat gcctttgtat catgctattg cttcccgtat ggctttcatt ttctcctcct
     3001 tgtataaatc ctggttgctg tctctttatg aggagttgtg gcccgttgtc aggcaacgtg
     3061 gcgtggtgtg cactgtgttt gctgacgcaa cccccactgg ttggggcatt gccaccacct
     3121 gtcagctcct ttccgggact ttcgctttcc ccctccctat tgccacggcg gaactcatcg
     3181 ccgcctgcct tgcccgctgc tggacagggg ctcggctgtt gggcactgac aattccgtgg
     3241 tgttgtcggg gaaatcatcg tcctttcctt ggctgctcgc ctatgttgcc acctggattc
     3301 tgcgcgggac gtccttctgc tacgtccctt cggccctcaa tccagcggac cttccttccc
     3361 gcggcctgct gccggctctg cggcctcttc cgcgtcttcg ccttcgccct cagacgagtc
     3421 ggatctccct ttgggccgcc tccccgcatc gataccgagc gctgctcgag agatcgatct
     3481 gcctcgactg tgccttctag ttgccagcca tctgttgttt gcccctcccc cgtgccttcc
     3541 ttgaccctgg aaggtgccac tcccactgtc ctttcctaat aaaatgagga aattgcatcg
     3601 cattgtctga gtaggtgtca ttctattctg gggggtgggg tggggcagga cagcaagggg
     3661 gaggattggg aagacaatag caggcatgct ggggacacgt gcggaccgag cggccgcagg
     3721 aacccctagt gatggagttg gccactccct ctctgcgcgc tcgctcgctc actgaggccg
     3781 ggcgaccaaa ggtcgcccga cgcccgggct ttgcccgggc ggcctcagtg agcgagcgag
     3841 cgcgcagctg cctgcagggg cgcctgatgc ggtattttct ccttacgcat ctgtgcggta
     3901 tttcacaccg catacgtcaa agcaaccata gtacgcgccc tgtagcggcg cattaagcgc
     3961 ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt gccagcgccc tagcgcccgc
     4021 tcctttcgct ttcttccctt cctttctcgc cacgttcgcc ggctttcccc gtcaagctct
     4081 aaatcggggg ctccctttag ggttccgatt tagtgcttta cggcacctcg accccaaaaa
     4141 acttgatttg ggtgatggtt cacgtagtgg gccatcgccc tgatagacgg tttttcgccc
     4201 tttgacgttg gagtccacgt tctttaatag tggactcttg ttccaaactg gaacaacact
     4261 caaccctatc tcgggctatt cttttgattt ataagggatt ttgccgattt cggcctattg
     4321 gttaaaaaat gagctgattt aacaaaaatt taacgcgaat tttaacaaaa tattaacgtt
     4381 tacaatttta tggtgcactc tcagtacaat ctgctctgat gccgcatagt taagccagcc
     4441 ccgacacccg ccaacacccg ctgacgcgcc ctgacgggct tgtctgctcc cggcatccgc
     4501 ttacagacaa gctgtgaccg tctccgggag ctgcatgtgt cagaggtttt caccgtcatc
     4561 accgaaacgc gcgagacgaa agggcctcgt gatacgccta tttttatagg ttaatgtcat
     4621 gataataatg gtttcttaga cgtcaggtgg cacttttcgg ggaaatgtgc gcggaacccc
     4681 tatttgttta tttttctaaa tacattcaaa tatgtatccg ctcatgagac aataaccctg
     4741 ataaatgctt caataatatt gaaaaaggaa gagtatgagt attcaacatt tccgtgtcgc
     4801 ccttattccc ttttttgcgg cattttgcct tcctgttttt gctcacccag aaacgctggt
     4861 gaaagtaaaa gatgctgaag atcagttggg tgcacgagtg ggttacatcg aactggatct
     4921 caacagcggt aagatccttg agagttttcg ccccgaagaa cgttttccaa tgatgagcac
     4981 ttttaaagtt ctgctatgtg gcgcggtatt atcccgtatt gacgccgggc aagagcaact
     5041 cggtcgccgc atacactatt ctcagaatga cttggttgag tactcaccag tcacagaaaa
     5101 gcatcttacg gatggcatga cagtaagaga attatgcagt gctgccataa ccatgagtga
     5161 taacactgcg gccaacttac ttctgacaac gatcggagga ccgaaggagc taaccgcttt
     5221 tttgcacaac atgggggatc atgtaactcg ccttgatcgt tgggaaccgg agctgaatga
     5281 agccatacca aacgacgagc gtgacaccac gatgcctgta gcaatggcaa caacgttgcg
     5341 caaactatta actggcgaac tacttactct agcttcccgg caacaattaa tagactggat
     5401 ggaggcggat aaagttgcag gaccacttct gcgctcggcc cttccggctg gctggtttat
     5461 tgctgataaa tctggagccg gtgagcgtgg gtctcgcggt atcattgcag cactggggcc
     5521 agatggtaag ccctcccgta tcgtagttat ctacacgacg gggagtcagg caactatgga
     5581 tgaacgaaat agacagatcg ctgagatagg tgcctcactg attaagcatt ggtaactgtc
     5641 agaccaagtt tactcatata tactttagat tgatttaaaa cttcattttt aatttaaaag
     5701 gatctaggtg aagatccttt ttgataatct catgaccaaa atcccttaac gtgagttttc
     5761 gttccactga gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag atcctttttt
     5821 tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt
     5881 gccggatcaa gagctaccaa ctctttttcc gaaggtaact ggcttcagca gagcgcagat
     5941 accaaatact gtccttctag tgtagccgta gttaggccac cacttcaaga actctgtagc
     6001 accgcctaca tacctcgctc tgctaatcct gttaccagtg gctgctgcca gtggcgataa
     6061 gtcgtgtctt accgggttgg actcaagacg atagttaccg gataaggcgc agcggtcggg
     6121 ctgaacgggg ggttcgtgca cacagcccag cttggagcga acgacctaca ccgaactgag
     6181 atacctacag cgtgagctat gagaaagcgc cacgcttccc gaagggagaa aggcggacag
     6241 gtatccggta agcggcaggg tcggaacagg agagcgcacg agggagcttc cagggggaaa
     6301 cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt
     6361 gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg
     6421 gttcctggcc ttttgctggc cttttgctca catgt
//
