LOCUS       sequence-487711-        6428 bp    DNA     circular SYN 02-OCT-2025
DEFINITION  Can be used to generate AAV virus that will express mScarlet in the 
            presence of Cre.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6428)
  AUTHORS   Rylan Larsen
  TITLE     AAV red fluorescent protein plasmids
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 6428)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Oct 2, 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            "Unpublished"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-mScarlet-WPRE
FEATURES             Location/Qualifiers
     source          1..6428
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..130
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     enhancer        223..526
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        528..806
                     /label=chicken beta-actin promoter
     intron          807..1823
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     1747..1769
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1831..1850
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     protein_bind    complement(1927..1960)
                     /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(1971..2004)
                     /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(2018..2713)
                     /codon_start=1
                     /product="bright monomeric red fluorescent protein evolved 
                     from a synthetic template (Bindels et al., 2016)"
                     /label=mScarlet
                     /translation="MVSKGEAVIKEFMRFKVHMEGSMNGHEFEIEGEGEGRPYEGTQTA
                     KLKVTKGGPLPFSWDILSPQFMYGSRAFTKHPADIPDYYKQSFPEGFKWERVMNFEDGG
                     AVTVTQDTSLEDGTLIYKVKLRGTNFPPDGPVMQKKTMGWEASTERLYPEDGVLKGDIK
                     MALRLKDGGRYLADFKTTYKAKKPVQMPGAYNVDRKLDITSHNEDYTVVEQYERSEGRH
                     STGGMDELYK"
     primer_bind     complement(2293..2312)
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     2562..2580
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     protein_bind    2731..2764
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     protein_bind    2775..2808
                     /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    2832..3420
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(2885..2905)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3303..3314)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(3455..3472)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3461..3668
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   3691..3831
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   3691..3820
                     /label=AAV2 ITR
     rep_origin      3906..4361
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3993..4012)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4203..4224
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4378..4397)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4497..4519
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4557..4575)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4643..4747
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4748..4816,4817..5608)
                     /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(4966..4985)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5779..6367
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6268..6287
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cctgcaggca gctgcgcgct cgctcgctca ctgaggccgc ccgggcgtcg ggcgaccttt
       61 ggtcgcccgg cctcagtgag cgagcgagcg cgcagagagg gagtggccaa ctccatcact
      121 aggggttcct gcggccgcac gcgaaacaat tctgcaggaa tctagttatt aatagtaatc
      181 aattacgggg tcattagttc atagcccata tatggagttc cgcgttacat aacttacggt
      241 aaatggcccg cctggctgac cgcccaacga cccccgccca ttgacgtcaa taatgacgta
      301 tgttcccata gtaacgccaa tagggacttt ccattgacgt caatgggtgg agtatttacg
      361 gtaaactgcc cacttggcag tacatcaagt gtatcatatg ccaagtacgc cccctattga
      421 cgtcaatgac ggtaaatggc ccgcctggca ttatgcccag tacatgacct tatgggactt
      481 tcctacttgg cagtacatct acgtattagt catcgctatt accatggtcg aggtgagccc
      541 cacgttctgc ttcactctcc ccatctcccc cccctcccca cccccaattt tgtatttatt
      601 tattttttaa ttattttgtg cagcgatggg ggcggggggg gggggggggg cgcgcgccag
      661 gcggggcggg gcggggcgag gggcggggcg gggcgaggcg gagaggtgcg gcggcagcca
      721 atcagagcgg cgcgctccga aagtttcctt ttatggcgag gcggcggcgg cggcggccct
      781 ataaaaagcg aagcgcgcgg cgggcgggag tcgctgcgcg ctgccttcgc cccgtgcccc
      841 gctccgccgc cgcctcgcgc cgcccgcccc ggctctgact gaccgcgtta ctcccacagg
      901 tgagcgggcg ggacggccct tctcctccgg gctgtaatta gcgcttggtt taatgacggc
      961 ttgtttcttt tctgtggctg cgtgaaagcc ttgaggggct ccgggagggc cctttgtgcg
     1021 gggggagcgg ctcggggggt gcgtgcgtgt gtgtgtgcgt ggggagcgcc gcgtgcggct
     1081 ccgcgctgcc cggcggctgt gagcgctgcg ggcgcggcgc ggggctttgt gcgctccgca
     1141 gtgtgcgcga ggggagcgcg gccgggggcg gtgccccgcg gtgcgggggg ggctgcgagg
     1201 ggaacaaagg ctgcgtgcgg ggtgtgtgcg tgggggggtg agcagggggt gtgggcgcgt
     1261 cggtcgggct gcaacccccc ctgcaccccc ctccccgagt tgctgagcac ggcccggctt
     1321 cgggtgcggg gctccgtacg gggcgtggcg cggggctcgc cgtgccgggc ggggggtggc
     1381 ggcaggtggg ggtgccgggc ggggcggggc cgcctcgggc cggggagggc tcgggggagg
     1441 ggcgcggcgg cccccggagc gccggcggct gtcgaggcgc ggcgagccgc agccattgcc
     1501 ttttatggta atcgtgcgag agggcgcagg gacttccttt gtcccaaatc tgtgcggagc
     1561 cgaaatctgg gaggcgccgc cgcaccccct ctagcgggcg cggggcgaag cggtgcggcg
     1621 ccggcaggaa ggaaatgggc ggggagggcc ttcgtgcgtc gccgcgccgc cgtccccttc
     1681 tccctctcca gcctcggggc tgtccgcggg gggacggctg ccttcggggg ggacggggca
     1741 gggcggggtt cggcttctgg cgtgtgaccg gcggctctag agcctctgct aaccatgttc
     1801 atgccttctt ctttttccta cagctcctgg gcaacgtgct ggttattgtg ctgtctcatc
     1861 attttggcaa agaattgatt tgataccgcg ggccccgcgt ggacggtacc cttcaggtcg
     1921 acgtcgataa cttcgtatag catacattat acgaagttat ccgaatcgca ataacttcgt
     1981 ataggatact ttatacgaag ttatttggcg cgcctgactt gtacagctcg tccatgccgc
     2041 cggtggagtg gcggccctcg gagcgttcgt actgttccac cacggtgtag tcctcgttgt
     2101 gggaggtgat gtccaacttg cggtcgacgt tgtaggcgcc gggcatctgc acgggcttct
     2161 tggccttgta ggtggtcttg aagtccgcca ggtagcggcc gccgtccttc aggcgcaggg
     2221 ccatcttaat gtcgcccttc agcacgccgt cctcggggta caaccgctcg gtggacgctt
     2281 cccagcccat tgtcttcttc tgcattacgg ggccgtcagg agggaagttg gtgccgcgga
     2341 gcttcacctt gtagatcagg gtgccgtcct ccagggaggt gtcctgggtc acggtcacgg
     2401 cgccgccgtc ctcgaagttc atcacgcgct cccacttgaa gccctcgggg aaggactgct
     2461 tatagtagtc ggggatgtcg gcggggtgct tggtgaaggc cctggagccg tacatgaact
     2521 gaggggacag gatgtcccag gagaagggca gggggccacc cttggtcacc ttcagcttgg
     2581 cggtctgggt gccctcgtag gggcggccct cgccctcgcc ctcgatctcg aactcgtggc
     2641 cgttcatgga gccctccatg tgcaccttga accgcatgaa ctccttgatc actgcctcgc
     2701 ccttgctcac catgccaccg gccggccttg ataacttcgt ataatgtatg ctatacgaag
     2761 ttatccgaat cgcaataact tcgtataaag tatcctatac gaagttatcg aagatcatca
     2821 agcttatcga taatcaacct ctggattaca aaatttgtga aagattgact ggtattctta
     2881 actatgttgc tccttttacg ctatgtggat acgctgcttt aatgcctttg tatcatgcta
     2941 ttgcttcccg tatggctttc attttctcct ccttgtataa atcctggttg ctgtctcttt
     3001 atgaggagtt gtggcccgtt gtcaggcaac gtggcgtggt gtgcactgtg tttgctgacg
     3061 caacccccac tggttggggc attgccacca cctgtcagct cctttccggg actttcgctt
     3121 tccccctccc tattgccacg gcggaactca tcgccgcctg ccttgcccgc tgctggacag
     3181 gggctcggct gttgggcact gacaattccg tggtgttgtc ggggaaatca tcgtcctttc
     3241 cttggctgct cgcctatgtt gccacctgga ttctgcgcgg gacgtccttc tgctacgtcc
     3301 cttcggccct caatccagcg gaccttcctt cccgcggcct gctgccggct ctgcggcctc
     3361 ttccgcgtct tcgccttcgc cctcagacga gtcggatctc cctttgggcc gcctccccgc
     3421 atcgataccg agcgctgctc gagagatcga tctgcctcga ctgtgccttc tagttgccag
     3481 ccatctgttg tttgcccctc ccccgtgcct tccttgaccc tggaaggtgc cactcccact
     3541 gtcctttcct aataaaatga ggaaattgca tcgcattgtc tgagtaggtg tcattctatt
     3601 ctggggggtg gggtggggca ggacagcaag ggggaggatt gggaagacaa tagcaggcat
     3661 gctggggaca cgtgcggacc gagcggccgc aggaacccct agtgatggag ttggccactc
     3721 cctctctgcg cgctcgctcg ctcactgagg ccgggcgacc aaaggtcgcc cgacgcccgg
     3781 gctttgcccg ggcggcctca gtgagcgagc gagcgcgcag ctgcctgcag gggcgcctga
     3841 tgcggtattt tctccttacg catctgtgcg gtatttcaca ccgcatacgt caaagcaacc
     3901 atagtacgcg ccctgtagcg gcgcattaag cgcggcgggt gtggtggtta cgcgcagcgt
     3961 gaccgctaca cttgccagcg ccttagcgcc cgctcctttc gctttcttcc cttcctttct
     4021 cgccacgttc gccggctttc cccgtcaagc tctaaatcgg gggctccctt tagggttccg
     4081 atttagtgct ttacggcacc tcgaccccaa aaaacttgat ttgggtgatg gttcacgtag
     4141 tgggccatcg ccctgataga cggtttttcg ccctttgacg ttggagtcca cgttctttaa
     4201 tagtggactc ttgttccaaa ctggaacaac actcaactct atctcgggct attcttttga
     4261 tttataaggg attttgccga tttcggtcta ttggttaaaa aatgagctga tttaacaaaa
     4321 atttaacgcg aattttaaca aaatattaac gtttacaatt ttatggtgca ctctcagtac
     4381 aatctgctct gatgccgcat agttaagcca gccccgacac ccgccaacac ccgctgacgc
     4441 gccctgacgg gcttgtctgc tcccggcatc cgcttacaga caagctgtga ccgtctccgg
     4501 gagctgcatg tgtcagaggt tttcaccgtc atcaccgaaa cgcgcgagac gaaagggcct
     4561 cgtgatacgc ctatttttat aggttaatgt catgataata atggtttctt agacgtcagg
     4621 tggcactttt cggggaaatg tgcgcggaac ccctatttgt ttatttttct aaatacattc
     4681 aaatatgtat ccgctcatga gacaataacc ctgataaatg cttcaataat attgaaaaag
     4741 gaagagtatg agtattcaac atttccgtgt cgcccttatt cccttttttg cggcattttg
     4801 ccttcctgtt tttgctcacc cagaaacgct ggtgaaagta aaagatgctg aagatcagtt
     4861 gggtgcacga gtgggttaca tcgaactgga tctcaacagc ggtaagatcc ttgagagttt
     4921 tcgccccgaa gaacgttttc caatgatgag cacttttaaa gttctgctat gtggcgcggt
     4981 attatcccgt attgacgccg ggcaagagca actcggtcgc cgcatacact attctcagaa
     5041 tgacttggtt gagtactcac cagtcacaga aaagcatctt acggatggca tgacagtaag
     5101 agaattatgc agtgctgcca taaccatgag tgataacact gcggccaact tacttctgac
     5161 aacgatcgga ggaccgaagg agctaaccgc ttttttgcac aacatggggg atcatgtaac
     5221 tcgccttgat cgttgggaac cggagctgaa tgaagccata ccaaacgacg agcgtgacac
     5281 cacgatgcct gtagcaatgg caacaacgtt gcgcaaacta ttaactggcg aactacttac
     5341 tctagcttcc cggcaacaat taatagactg gatggaggcg gataaagttg caggaccact
     5401 tctgcgctcg gcccttccgg ctggctggtt tattgctgat aaatctggag ccggtgagcg
     5461 tgggtctcgc ggtatcattg cagcactggg gccagatggt aagccctccc gtatcgtagt
     5521 tatctacacg acggggagtc aggcaactat ggatgaacga aatagacaga tcgctgagat
     5581 aggtgcctca ctgattaagc attggtaact gtcagaccaa gtttactcat atatacttta
     5641 gattgattta aaacttcatt tttaatttaa aaggatctag gtgaagatcc tttttgataa
     5701 tctcatgacc aaaatccctt aacgtgagtt ttcgttccac tgagcgtcag accccgtaga
     5761 aaagatcaaa ggatcttctt gagatccttt ttttctgcgc gtaatctgct gcttgcaaac
     5821 aaaaaaacca ccgctaccag cggtggtttg tttgccggat caagagctac caactctttt
     5881 tccgaaggta actggcttca gcagagcgca gataccaaat actgttcttc tagtgtagcc
     5941 gtagttaggc caccacttca agaactctgt agcaccgcct acatacctcg ctctgctaat
     6001 cctgttacca gtggctgctg ccagtggcga taagtcgtgt cttaccgggt tggactcaag
     6061 acgatagtta ccggataagg cgcagcggtc gggctgaacg gggggttcgt gcacacagcc
     6121 cagcttggag cgaacgacct acaccgaact gagataccta cagcgtgagc tatgagaaag
     6181 cgccacgctt cccgaaggga gaaaggcgga caggtatccg gtaagcggca gggtcggaac
     6241 aggagagcgc acgagggagc ttccaggggg aaacgcctgg tatctttata gtcctgtcgg
     6301 gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct
     6361 atggaaaaac gccagcaacg cggccttttt acggttcctg gccttttgct ggccttttgc
     6421 tcacatgt
//
