LOCUS       sequence-470755-        5540 bp    DNA     circular SYN 01-MAY-2025
DEFINITION  AiE1116m is an enhancer sequence, designed to drive AAV-mediated 
            transgene expression in specific populations of brain cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5540)
  AUTHORS   Ben-Simon Y, Hooper M, Narayan S, Daigle T, Dwivedi D, Way SW, Oster
            A, Stafford DA, Mich JK, Taormina MJ, Martinez RA, Opitz-Araya X, 
            Roth JR, Allen S, Ayala A, Bakken TE, Barcelli T, Barta S, Bendrick 
            J, Bertagnolli D, Bowlus J, Boyer G, Brouner K, Casian B, Casper T, 
            Chakka AB, Chakrabarty R, Chance RK, Chavan S, Departee M, Donadio 
            N, Dotson N, Egdorf T, Gabitto M, Garcia J, Gary A, Gasperini M, 
            Goldy J, Gore BB, Graybuck L, Greisman N, Haeseleer F, Halterman C, 
            Helback O, Hockemeyer D, Huang C, Huff S, Hunker A, Johansen N, 
            Juneau Z, Kalmbach B, Khem S, Kussick E, Kutsal R, Larsen R, Lee C, 
            Lee AY, Leibly M, Lenz GH, Liang E, Lusk N, Malone J, Mollenkopf T, 
            Morin E, Newman D, Ng L, Ngo K, Omstead V, Oyama A, Pham T, Pom CA, 
            Potekhina L, Ransford S, Rette D, Rimorin C, Rocha D, Ruiz A, 
            Sanchez REA, Sedeno-Cortes A, Sevigny JP, Shapovalova N, Shulga L, 
            Sigler AR, Siverts LA, Somasundaram S, Stewart K, Szelenyi E, Tieu 
            M, Trader C, van Velthoven CTJ, Walker M, Weed N, Wirthlin M, Wood 
            T, Wynalda B, Yao Z, Zhou T, Ariza J, Dee N, Reding M, Ronellenfitch
            K, Mufti S, Sunkin SM, Smith KA, Esposito L, Waters J, Thyagarajan 
            B, Yao S, Lein ES, Zeng H, Levi BP, Ngai J, Ting J, Tasic B
  TITLE     A suite of enhancer AAVs and transgenic mouse lines for genetic 
            access to cortical cell types.
  JOURNAL   bioRxiv [Preprint]. 2024 Sep 26:2024.06.10.597244. doi: 
            10.1101/2024.06.10.597244.
  PUBMED    38915722
REFERENCE   2  (bases 1 to 5540)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 1, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "bioRxiv 
            [Preprint]. 2024 Sep 26:2024.06.10.597244. doi: 
            10.1101/2024.06.10.597244."
COMMENT     Sequence Label: AiP14238 -
            pAAV-AiE1116m-minBG-SYFP2-P2A-3XFLA...
FEATURES             Location/Qualifiers
     source          1..5540
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      150..738
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     639..658
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     misc_feature    795..822
                     /gene="telR"
                     /label=telR
                     /note="telomeric sequence from coliphage N15"
     misc_feature    823..850
                     /gene="telL"
                     /label=telL
                     /note="telomeric sequence from coliphage N15"
     repeat_region   857..986
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     regulatory      1749..1758
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1755..1757,1758..1760,1761..2471)
                     /codon_start=1
                     /product="monomeric optimized variant of EYFP, identical to
                     mVenus but with L68V (Kremers et al., 2006)"
                     /label=SYFP2
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KLICTTGKLPVPWPTLVTTLGYGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYITADKQKNGIK
                     ANFKIRHNIEDGGVQLADHYQQNTPIGDGPVLLPDNHYLSYQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(2061..2080)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2408..2429
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             2481..2537
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     CDS             2550..2615
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             2646..3023
                     /codon_start=1
                     /gene="HIST1H2BJ"
                     /product="human histone H2B"
                     /label=H2B
                     /translation="PEPAKSAPAPKKGSKKAVTKAQKKGGKKRKRSRKESYSIYVYKVL
                     KQVHPDTGISSKAMGIMNSFVNDIFERIAGEASRLAHYNKRSTITSREIQTAVRLLLPG
                     ELAKHAVSEGTKAITKYTSAK"
     primer_bind     complement(3110..3130)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     polyA_signal    3317..3428
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   3546..3686
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   3546..3675
                     /label=AAV2 ITR
     misc_feature    3693..3720
                     /gene="telR"
                     /label=telR
                     /note="telomeric sequence from coliphage N15"
     misc_feature    3721..3748
                     /gene="telL"
                     /label=telL
                     /note="telomeric sequence from coliphage N15"
     rep_origin      3817..4272
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3904..3923)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4114..4135
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4289..4308)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4408..4430
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4468..4486)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4554..4658
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4659..4727,4728..5519)
                     /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(4877..4896)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 tatatacttt agattgattt aaaacttcat ttttaattta aaaggatcta ggtgaagatc
       61 ctttttgata atctcatgac caaaatccct taacgtgagt tttcgttcca ctgagcgtca
      121 gaccccgtag aaaagatcaa aggatcttct tgagatcctt tttttctgcg cgtaatctgc
      181 tgcttgcaaa caaaaaaacc accgctacca gcggtggttt gtttgccgga tcaagagcta
      241 ccaactcttt ttccgaaggt aactggcttc agcagagcgc agataccaaa tactgttctt
      301 ctagtgtagc cgtagttagg ccaccacttc aagaactctg tagcaccgcc tacatacctc
      361 gctctgctaa tcctgttacc agtggctgct gccagtggcg ataagtcgtg tcttaccggg
      421 ttggactcaa gacgatagtt accggataag gcgcagcggt cgggctgaac ggggggttcg
      481 tgcacacagc ccagcttgga gcgaacgacc tacaccgaac tgagatacct acagcgtgag
      541 ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc
      601 agggtcggaa caggagagcg cacgagggag cttccagggg gaaacgcctg gtatctttat
      661 agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg
      721 gggcggagcc tatggaaaaa cgccagcaac gcggcctttt tacggttcct ggccttttgc
      781 tggccttttg ctcatatcag cacacaattg cccattatac gcgcgtataa tggactattg
      841 tgtgctgata acatgtcctg caggcagctg cgcgctcgct cgctcactga ggccgcccgg
      901 gcgtcgggcg acctttggtc gcccggcctc agtgagcgag cgagcgcgca gagagggagt
      961 ggccaactcc atcactaggg gttcctgcgg ccgcacgcgt tgtgcagttc ctgaagcaga
     1021 tctgtagtat cctaccatta ttatgccata atatgaaatc tatttattca ctggcattta
     1081 aactaagcaa acgtgtaact cgtccttggc aaaatcacgt actagtgtag ttggaaaggg
     1141 cctgaatgac ataatacctt tctaactttc aagtcatttc ttgcttgttt aattgattta
     1201 atcataattc ctgtacatgt ggttttatgg agcatacaga gtacaactta gaatcaaagc
     1261 aaaacagaga ggggagaagt gtaaaaacaa ttagttattc tttgtaatat taggcactta
     1321 attgctggct caagataaaa agaattggat gtcaaagaca atggagaact ttgaactcta
     1381 aaaatagcca taatggaaat tcactgtaga aacaacttac tttacaaaag caaagcaaag
     1441 caacagcagt aagcaagagc ccatgcgttt atgggactgg tagtgtacat gccagataaa
     1501 gtattactgc atacttacca ccgtctggag cctcctttcc atttgggatt tgtatagttt
     1561 tggatgtctc tttaaattga cttcttgatt ttcattcaaa catgtactgt aaggctaaag
     1621 aataatttat gctctcataa gggagacaaa taagagagct cgggctgggc ataaaagtca
     1681 gggcagagcc atctattgct tacatttgct tctgggatcc agatctttcg aagctagcgc
     1741 taccggtcgc caccatggtc tccaagggcg aggagctgtt caccggggtg gtgcccatcc
     1801 tggtcgagct ggacggcgat gtcaacggcc acaagttcag cgtgtccggc gagggcgagg
     1861 gcgatgccac ctacggcaag ctgaccctga agctgatctg caccaccggc aagctgcccg
     1921 tgccctggcc caccctcgtg accaccctgg gctacggcgt gcagtgcttc gcccgctacc
     1981 ccgaccacat gaagcagcac gacttcttca agtccgccat gcccgaaggc tacgtccagg
     2041 agcgcaccat cttcttcaaa gacgacggca actacaagac ccgcgccgag gtgaagttcg
     2101 agggcgacac cctggtgaac cgcatcgagc tgaagggcat cgacttcaag gaggacggca
     2161 acatcctggg gcacaagctg gagtacaact acaacagcca caacgtctat atcaccgccg
     2221 acaagcagaa gaacggcatc aaggccaact tcaagatccg ccacaacatc gaggacggcg
     2281 gcgtgcagct cgccgaccac taccagcaga acacccccat cggcgacggc cccgtgctgc
     2341 tgcccgacaa ccactacctg agctaccagt ccaagctgag caaagacccc aacgagaagc
     2401 gcgatcacat ggtcctgctg gagttcgtga ccgccgccgg gatcactctc ggcatggacg
     2461 agctgtacaa gggctctggt gctaccaact tctcactgtt gaaacaggca ggggatgtag
     2521 aggagaatcc agggcctggt gctagtggag actacaaaga ccatgacgga gattataaag
     2581 atcatgacat cgattacaag gatgacgatg acaagtccgg actcagatct ggaggctccg
     2641 gaggcccaga gccagcgaag tctgctcccg ccccgaaaaa gggctccaag aaggcggtga
     2701 ctaaggcgca gaagaaaggc ggcaagaagc gcaagcgcag ccgcaaggag agctattcca
     2761 tctatgtgta taaggttctg aagcaggtcc accctgacac cggcatttcg tccaaggcca
     2821 tgggcatcat gaactcgttt gtgaacgaca ttttcgagcg catcgcaggt gaggcttccc
     2881 gcctggcgca ttacaacaag cgctcgacca tcacctccag ggagatccag acggccgtgc
     2941 gcctgctgct gcctggggag ttggccaagc acgccgtgtc cgagggtact aaggccatca
     3001 ccaagtacac cagcgctaag taatgaggcg cgccgcggcc gcgaattcga tatcataatc
     3061 aacctctgga ttacaaaatt tgtgaaagat tgactggtat tcttaactat gttgctcctt
     3121 ttacgctatg tggatacgct gctttaatgc ctttgtatca tgctattgct tcccgtatgg
     3181 ctttcatttt ctcctccttg tataaatcct ggttagttct tgccacggcg gaactcatcg
     3241 ccgcctgcct tgcccgctgc tggacagggg ctcggctgtt gggcactgac aattccgtgg
     3301 ctcgagagat cttcgactgt gccttctagt tgccagccat ctgttgtttg cccctccccc
     3361 gtgccttcct tgaccctgga aggtgccact cccactgtcc tttcctaata aaatgaggaa
     3421 attgcatcgc attgtctgag taggtgtcat tctattctgg ggggtggggt ggggcaggac
     3481 agcaaggggg aggattggga agacaatagc aggcatgaga tctcacgtgc ggaccgagcg
     3541 gccgcaggaa cccctagtga tggagttggc cactccctct ctgcgcgctc gctcgctcac
     3601 tgaggccggg cgaccaaagg tcgcccgacg cccgggcttt gcccgggcgg cctcagtgag
     3661 cgagcgagcg cgcagctgcc tgcaggggcg cctatcagca cacaattgcc cattatacgc
     3721 gcgtataatg gactattgtg tgctgatatg atgcggtatt ttctccttac gcatctgtgc
     3781 ggtatttcac accgcatacg tcaaagcaac catagtacgc gccctgtagc ggcgcattaa
     3841 gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac acttgccagc gccttagcgc
     3901 ccgctccttt cgctttcttc ccttcctttc tcgccacgtt cgccggcttt ccccgtcaag
     3961 ctctaaatcg ggggctccct ttagggttcc gatttagtgc tttacggcac ctcgacccca
     4021 aaaaacttga tttgggtgat ggttcacgta gtgggccatc gccctgatag acggtttttc
     4081 gccctttgac gttggagtcc acgttcttta atagtggact cttgttccaa actggaacaa
     4141 cactcaactc tatctcgggc tattcttttg atttataagg gattttgccg atttcggtct
     4201 attggttaaa aaatgagctg atttaacaaa aatttaacgc gaattttaac aaaatattaa
     4261 cgtttacaat tttatggtgc actctcagta caatctgctc tgatgccgca tagttaagcc
     4321 agccccgaca cccgccaaca cccgctgacg cgccctgacg ggcttgtctg ctcccggcat
     4381 ccgcttacag acaagctgtg accgtctccg ggagctgcat gtgtcagagg ttttcaccgt
     4441 catcaccgaa acgcgcgaga cgaaagggcc tcgtgatacg cctattttta taggttaatg
     4501 tcatgataat aatggtttct tagacgtcag gtggcacttt tcggggaaat gtgcgcggaa
     4561 cccctatttg tttatttttc taaatacatt caaatatgta tccgctcatg agacaataac
     4621 cctgataaat gcttcaataa tattgaaaaa ggaagagtat gagtattcaa catttccgtg
     4681 tcgcccttat tccctttttt gcggcatttt gccttcctgt ttttgctcac ccagaaacgc
     4741 tggtgaaagt aaaagatgct gaagatcagt tgggtgcacg agtgggttac atcgaactgg
     4801 atctcaacag cggtaagatc cttgagagtt ttcgccccga agaacgtttt ccaatgatga
     4861 gcacttttaa agttctgcta tgtggcgcgg tattatcccg tattgacgcc gggcaagagc
     4921 aactcggtcg ccgcatacac tattctcaga atgacttggt tgagtactca ccagtcacag
     4981 aaaagcatct tacggatggc atgacagtaa gagaattatg cagtgctgcc ataaccatga
     5041 gtgataacac tgcggccaac ttacttctga caacgatcgg aggaccgaag gagctaaccg
     5101 cttttttgca caacatgggg gatcatgtaa ctcgccttga tcgttgggaa ccggagctga
     5161 atgaagccat accaaacgac gagcgtgaca ccacgatgcc tgtagcaatg gcaacaacgt
     5221 tgcgcaaact attaactggc gaactactta ctctagcttc ccggcaacaa ttaatagact
     5281 ggatggaggc ggataaagtt gcaggaccac ttctgcgctc ggcccttccg gctggctggt
     5341 ttattgctga taaatctgga gccggtgagc gtgggtctcg cggtatcatt gcagcactgg
     5401 ggccagatgg taagccctcc cgtatcgtag ttatctacac gacggggagt caggcaacta
     5461 tggatgaacg aaatagacag atcgctgaga taggtgcctc actgattaag cattggtaac
     5521 tgtcagacca agtttactca
//
