LOCUS       sequence-263712-        7342 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  AAV plasmid encoding mouse PM20D1 with C-terminal flag tag.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7342)
  AUTHORS   Kim JT, Li VL, Terrell SM, Fischer CR, Long JZ
  TITLE     Family-wide Annotation of Enzymatic Pathways by Parallel In Vivo 
            Metabolomics.
  JOURNAL   Cell Chem Biol. 2019 Nov 21;26(11):1623-1629.e3. doi: 
            10.1016/j.chembiol.2019.09.009. Epub 2019 Oct 3.
  PUBMED    31587987
REFERENCE   2  (bases 1 to 7342)
  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: 
            "10.1016/j.chembiol.2019.09.009"; journalName: "Cell Chem Biol"; 
            date: "2019-11-21- 21"; volume: "26"; issue: "11"; pages: 
            "1623-1629.e3"
COMMENT     Sequence Label: pENN.AAV.CB7.CI.PM20D1-flag.WPRE.rBG
FEATURES             Location/Qualifiers
     source          1..7342
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..130
                     /label=AAV2 ITR
     enhancer        204..583
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        585..862
                     /label=chicken beta-actin promoter
     intron          863..1879
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     1803..1825
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1887..1906
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     regulatory      1939..1948
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             3487..3510
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             3532..3549
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     misc_feature    3565..4153
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3618..3638)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(4036..4047)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(4179..4198)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    4244..4299
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     repeat_region   4392..4521
                     /label=AAV2 ITR
     primer_bind     complement(4540..4557)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4540..4556)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4549..4571)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      4698..5153
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4785..4804)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4995..5016
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        5179..5283
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5284..5352,5353..6144)
                     /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(5502..5521)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6315..6903
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6804..6823
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     protein_bind    7191..7212
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(7227..7232,7233..7250,7251..7257)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    7265..7281
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     primer_bind     7270..7292
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     7289..7305
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     7289..7305
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
ORIGIN
        1 ctgcgcgctc gctcgctcac tgaggccgcc cgggcaaagc ccgggcgtcg ggcgaccttt
       61 ggtcgcccgg cctcagtgag cgagcgagcg cgcagagagg gagtggccaa ctccatcact
      121 aggggttcct tgtagttaat gattaacccg ccatgctact tatctaccag ggtaatgggg
      181 atcctctaga actatagcta gtcgacattg attattgact agttattaat agtaatcaat
      241 tacggggtca ttagttcata gcccatatat ggagttccgc gttacataac ttacggtaaa
      301 tggcccgcct ggctgaccgc ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt
      361 tcccatagta acgccaatag ggactttcca ttgacgtcaa tgggtggagt atttacggta
      421 aactgcccac ttggcagtac atcaagtgta tcatatgcca agtacgcccc ctattgacgt
      481 caatgacggt aaatggcccg cctggcatta tgcccagtac atgaccttat gggactttcc
      541 tacttggcag tacatctacg tattagtcat cgctattacc atggtcgagg tgagccccac
      601 gttctgcttc actctcccca tctccccccc ctccccaccc ccaattttgt atttatttat
      661 tttttaatta ttttgtgcag cgatgggggc gggggggggg ggggggcgcg cgccaggcgg
      721 ggcggggcgg ggcgaggggc ggggcggggc gaggcggaga ggtgcggcgg cagccaatca
      781 gagcggcgcg ctccgaaagt ttccttttat ggcgaggcgg cggcggcggc ggccctataa
      841 aaagcgaagc gcgcggcggg cgggagtcgc tgcgcgctgc cttcgccccg tgccccgctc
      901 cgccgccgcc tcgcgccgcc cgccccggct ctgactgacc gcgttactcc cacaggtgag
      961 cgggcgggac ggcccttctc ctccgggctg taattagcgc ttggtttaat gacggcttgt
     1021 ttcttttctg tggctgcgtg aaagccttga ggggctccgg gagggccctt tgtgcggggg
     1081 gagcggctcg gggggtgcgt gcgtgtgtgt gtgcgtgggg agcgccgcgt gcggctccgc
     1141 gctgcccggc ggctgtgagc gctgcgggcg cggcgcgggg ctttgtgcgc tccgcagtgt
     1201 gcgcgagggg agcgcggccg ggggcggtgc cccgcggtgc ggggggggct gcgaggggaa
     1261 caaaggctgc gtgcggggtg tgtgcgtggg ggggtgagca gggggtgtgg gcgcgtcggt
     1321 cgggctgcaa ccccccctgc acccccctcc ccgagttgct gagcacggcc cggcttcggg
     1381 tgcggggctc cgtacggggc gtggcgcggg gctcgccgtg ccgggcgggg ggtggcggca
     1441 ggtgggggtg ccgggcgggg cggggccgcc tcgggccggg gagggctcgg gggaggggcg
     1501 cggcggcccc cggagcgccg gcggctgtcg aggcgcggcg agccgcagcc attgcctttt
     1561 atggtaatcg tgcgagaggg cgcagggact tcctttgtcc caaatctgtg cggagccgaa
     1621 atctgggagg cgccgccgca ccccctctag cgggcgcggg gcgaagcggt gcggcgccgg
     1681 caggaaggaa atgggcgggg agggccttcg tgcgtcgccg cgccgccgtc cccttctccc
     1741 tctccagcct cggggctgtc cgcgggggga cggctgcctt cgggggggac ggggcagggc
     1801 ggggttcggc ttctggcgtg tgaccggcgg ctctagagcc tctgctaacc atgttcatgc
     1861 cttcttcttt ttcctacagc tcctgggcaa cgtgctggtt attgtgctgt ctcatcattt
     1921 tggcaaagaa ttctgcaggc caccatggct gagctacttg ctagcttgcc cgcctgggca
     1981 gctgtgctcc ttctcttttt cgctacggtc tccggatcca ctggccctag aagcagggaa
     2041 aatcgggggg cgtcccggat cccttcccag ttcagcgagg aggagcgtgt cgctataaaa
     2101 gaggcgctga aaggtgccat ccagattccc acagtgtctt tcagccacga ggaatccaac
     2161 accacagccc ttgctgagtt tggagaatat atccgcaaag ccttccctac agtgttccac
     2221 agcagccttg tccaacatga agtcgtggca aagtatagcc acctgttcac catccaaggc
     2281 tcagacccca gtttgcagcc ctacatgctg atggctcaca ttgatgtggt tcctgccccg
     2341 gaagaaggat gggaggtgcc cccgttctca ggcctggaac gcaatggctt catctatggc
     2401 cggggtgcgc tggacaacaa aaactctgtg atggcgatcc tgcatgcttt ggagctcctg
     2461 ttgatcagaa actacagccc caaaagatct ttcttcattg ctttgggcca tgatgaggag
     2521 gtgtccgggg aaaagggggc tcagaagatc tcagcactct tacaggcaag gggtgtccag
     2581 ctagccttcc ttgtggatga agggagcttt atcttggaag gcttcattcc aaacctcgag
     2641 aagccagttg ccatgatttc agtcactgag aagggtgccc ttgacctcat gctgcaagta
     2701 aacatgactc caggccactc ttcagctccc ccaaaggaga caagcattgg cattctttct
     2761 gccgctgtca gccgactgga gcagacacca atgccgaata tgtttggagg agggccattg
     2821 aagaagacaa tgaagctact ggcaaatgag ttttccttcc ctatcaatat agtcttgaga
     2881 aacctgtggc tatttcatcc cattgtgagc aggataatgg agaggaaccc cataacaaat
     2941 gcgctggtcc gaactaccac agccctcacc atgttcaatg caggaatcaa ggtgaatgtc
     3001 atccctccat tggctcaggc tacaatcaac tgccgaattc acccttcgca gacagtacat
     3061 gaggtcctag aacttgtcaa gaacaccgtg gctgatgaca gagtccagct gcatgtgttg
     3121 agatcctttg aacccctgcc catcagcccc tctgatgacc aggccatggg ctaccagctg
     3181 cttcaagaga ccatacgatc tgtcttcccg gaagtcgaca tcgtcgtccc cggtatttgt
     3241 attgccaata cggacacccg acactatgcc aacatcacca atggcatgta ccggttcaac
     3301 ccccttcccc tgaaccctca ggacttcagt ggtgtccatg gaatcaatga gaaagtttcc
     3361 gttcagaact accagaacca ggtgaagttc atctttgagt tcatccaaaa tgccgacact
     3421 tacaaagagc cagttcctca tctgcatgaa ctaacgcgtc ccagatctcc caccggtccc
     3481 ggatccgact acaaggacga tgacgacaag gagctgaaga ccgcggccgc gcatcatcac
     3541 catcaccatt aaaagcttat cgataatcaa cctctggatt acaaaatttg tgaaagattg
     3601 actggtattc ttaactatgt tgctcctttt acgctatgtg gatacgctgc tttaatgcct
     3661 ttgtatcatg ctattgcttc ccgtatggct ttcattttct cctccttgta taaatcctgg
     3721 ttgctgtctc tttatgagga gttgtggccc gttgtcaggc aacgtggcgt ggtgtgcact
     3781 gtgtttgctg acgcaacccc cactggttgg ggcattgcca ccacctgtca gctcctttcc
     3841 gggactttcg ctttccccct ccctattgcc acggcggaac tcatcgccgc ctgccttgcc
     3901 cgctgctgga caggggctcg gctgttgggc actgacaatt ccgtggtgtt gtcggggaaa
     3961 tcatcgtcct ttccttggct gctcgcctgt gttgccacct ggattctgcg cgggacgtcc
     4021 ttctgctacg tcccttcggc cctcaatcca gcggaccttc cttcccgcgg cctgctgccg
     4081 gctctgcggc ctcttccgcg tcttcgcctt cgccctcaga cgagtcggat ctccctttgg
     4141 gccgcctccc cgcatcgata ccgtcgatga ggatccgatc tttttccctc tgccaaaaat
     4201 tatggggaca tcatgaagcc ccttgagcat ctgacttctg gctaataaag gaaatttatt
     4261 ttcattgcaa tagtgtgttg gaattttttg tgtctctcac tcggaagcaa ttcgttgatc
     4321 tgaatttcga ccacccataa tacccattac cctggtagat aagtagcatg gcgggttaat
     4381 cattaactac aaggaacccc tagtgatgga gttggccact ccctctctgc gcgctcgctc
     4441 gctcactgag gccgggcgac caaaggtcgc ccgacgcccg ggctttgccc gggcggcctc
     4501 agtgagcgag cgagcgcgca gccttaatta acctaattca ctggccgtcg ttttacaacg
     4561 tcgtgactgg gaaaaccctg gcgttaccca acttaatcgc cttgcagcac atcccccttt
     4621 cgccagctgg cgtaatagcg aagaggcccg caccgatcgc ccttcccaac agttgcgcag
     4681 cctgaatggc gaatgggacg cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt
     4741 tacgcgcagc gtgaccgcta cacttgccag cgccctagcg cccgctcctt tcgctttctt
     4801 cccttccttt ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc gggggctccc
     4861 tttagggttc cgatttagtg ctttacggca cctcgacccc aaaaaacttg attagggtga
     4921 tggttcacgt agtgggccat cgccctgata gacggttttt cgccctttga cgttggagtc
     4981 cacgttcttt aatagtggac tcttgttcca aactggaaca acactcaacc ctatctcggt
     5041 ctattctttt gatttataag ggattttgcc gatttcggcc tattggttaa aaaatgagct
     5101 gatttaacaa aaatttaacg cgaattttaa caaaatatta acgcttacaa tttaggtggc
     5161 acttttcggg gaaatgtgcg cggaacccct atttgtttat ttttctaaat acattcaaat
     5221 atgtatccgc tcatgagaca ataaccctga taaatgcttc aataatattg aaaaaggaag
     5281 agtatgagta ttcaacattt ccgtgtcgcc cttattccct tttttgcggc attttgcctt
     5341 cctgtttttg ctcacccaga aacgctggtg aaagtaaaag atgctgaaga tcagttgggt
     5401 gcacgagtgg gttacatcga actggatctc aacagcggta agatccttga gagttttcgc
     5461 cccgaagaac gttttccaat gatgagcact tttaaagttc tgctatgtgg cgcggtatta
     5521 tcccgtattg acgccgggca agagcaactc ggtcgccgca tacactattc tcagaatgac
     5581 ttggttgagt actcaccagt cacagaaaag catcttacgg atggcatgac agtaagagaa
     5641 ttatgcagtg ctgccataac catgagtgat aacactgcgg ccaacttact tctgacaacg
     5701 atcggaggac cgaaggagct aaccgctttt ttgcacaaca tgggggatca tgtaactcgc
     5761 cttgatcgtt gggaaccgga gctgaatgaa gccataccaa acgacgagcg tgacaccacg
     5821 atgcctgtag caatggcaac aacgttgcgc aaactattaa ctggcgaact acttactcta
     5881 gcttcccggc aacaattaat agactggatg gaggcggata aagttgcagg accacttctg
     5941 cgctcggccc ttccggctgg ctggtttatt gctgataaat ctggagccgg tgagcgtggg
     6001 tctcgcggta tcattgcagc actggggcca gatggtaagc cctcccgtat cgtagttatc
     6061 tacacgacgg ggagtcaggc aactatggat gaacgaaata gacagatcgc tgagataggt
     6121 gcctcactga ttaagcattg gtaactgtca gaccaagttt actcatatat actttagatt
     6181 gatttaaaac ttcattttta atttaaaagg atctaggtga agatcctttt tgataatctc
     6241 atgaccaaaa tcccttaacg tgagttttcg ttccactgag cgtcagaccc cgtagaaaag
     6301 atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa
     6361 aaaccaccgc taccagcggt ggtttgtttg ccggatcaag agctaccaac tctttttccg
     6421 aaggtaactg gcttcagcag agcgcagata ccaaatactg ttcttctagt gtagccgtag
     6481 ttaggccacc acttcaagaa ctctgtagca ccgcctacat acctcgctct gctaatcctg
     6541 ttaccagtgg ctgctgccag tggcgataag tcgtgtctta ccgggttgga ctcaagacga
     6601 tagttaccgg ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac acagcccagc
     6661 ttggagcgaa cgacctacac cgaactgaga tacctacagc gtgagctatg agaaagcgcc
     6721 acgcttcccg aagggagaaa ggcggacagg tatccggtaa gcggcagggt cggaacagga
     6781 gagcgcacga gggagcttcc agggggaaac gcctggtatc tttatagtcc tgtcgggttt
     6841 cgccacctct gacttgagcg tcgatttttg tgatgctcgt caggggggcg gagcctatgg
     6901 aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct tttgctggcc ttttgctcac
     6961 atgttctttc ctgcgttatc ccctgattct gtggataacc gtattaccgc ctttgagtga
     7021 gctgataccg ctcgccgcag ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg
     7081 gaagagcgcc caatacgcaa accgcctctc cccgcgcgtt ggccgattca ttaatgcagc
     7141 tggcacgaca ggtttcccga ctggaaagcg ggcagtgagc gcaacgcaat taatgtgagt
     7201 tagctcactc attaggcacc ccaggcttta cactttatgc ttccggctcg tatgttgtgt
     7261 ggaattgtga gcggataaca atttcacaca ggaaacagct atgaccatga ttacgccaga
     7321 tttaattaag gccttaatta gg
//
