LOCUS       sequence-470430-        4766 bp    DNA     circular SYN 30-APR-2025
DEFINITION  AiE2161m 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 4766)
  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 4766)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 30, 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: AiP1390 - pAAV-AiE2161m-minBG-SYFP2-WPRE3-BGHpA
FEATURES             Location/Qualifiers
     source          1..4766
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..130
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     CDS             complement(482..493)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     regulatory      778..787
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(784..786,787..789,790..1500)
                     /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(829..850)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1090..1109)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1437..1458
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(1593..1613)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     polyA_signal    1800..1911
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   2029..2169
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   2029..2158
                     /label=AAV2 ITR
     rep_origin      2244..2699
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2331..2350)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2541..2562
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2716..2735)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2835..2857
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(2895..2913)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        2981..3085
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3086..3154,3155..3946)
                     /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(3304..3323)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4117..4705
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4606..4625
                     /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 gcgtcaccat gtggaaagca aaagagtaat ttttacttga
      181 ttgttttgcc tactgaatcc aaccagtaaa agtaaatatt ctcccttttc cccgctggag
      241 agaggcagat agaaccagtt ctagtctggg aactacttcc tggttgcaca ggagcctgct
      301 agagcctcct cccccgactc agctgggcac aggaaggtgg cttcgtccct aaaaggtagc
      361 gcccgaggtc tggtgaggac acagtgagct ggttggccat gtgaagtgag ccagaggagg
      421 agaaggttga ggtcgctggt ccttagctct gacctgctgt tcccggtttc cttcagccaa
      481 ttctcccttc gattattttt agcatggagc agcggacaaa ggcagtgggc cagcaaggga
      541 agatagattt ctatcttaac caggccactt acacgggacc tctgtcttct agtctgtaga
      601 gtatagtaga tagtcccagc gctttaccct gtcactgtta aaaacactat gacctgcaca
      661 ccaccattat cttcatcacc tccagagctc gggctgggca taaaagtcag ggcagagcca
      721 tctattgctt acatttgctt ctgggatcca gatctttcga agctagcgct accggtcgcc
      781 accatggtga gcaagggcga ggagctgttc accggggtgg tgcccatcct ggtcgagctg
      841 gacggcgacg taaacggcca caagttcagc gtgtccggcg agggcgaggg cgatgccacc
      901 tacggcaagc tgaccctgaa gctgatctgc accaccggca agctgcccgt gccctggccc
      961 accctcgtga ccaccctggg ctacggcgtg cagtgcttcg cccgctaccc cgaccacatg
     1021 aagcagcacg acttcttcaa gtccgccatg cccgaaggct acgtccagga gcgcaccatc
     1081 ttcttcaagg acgacggcaa ctacaagacc cgcgccgagg tgaagttcga gggcgacacc
     1141 ctggtgaacc gcatcgagct gaagggcatc gacttcaagg aggacggcaa catcctgggg
     1201 cacaagctgg agtacaacta caacagccac aacgtctata tcaccgccga caagcagaag
     1261 aacggcatca aggccaactt caagatccgc cacaacatcg aggacggcgg cgtgcagctc
     1321 gccgaccact accagcagaa cacccccatc ggcgacggcc ccgtgctgct gcccgacaac
     1381 cactacctga gctaccagtc caagctgagc aaagacccca acgagaagcg cgatcacatg
     1441 gtcctgctgg agttcgtgac cgccgccggg atcactctcg gcatggacga gctgtacaag
     1501 taagtcgacg gcgcgccgcg gccgcgaatt cgatatcata atcaacctct ggattacaaa
     1561 atttgtgaaa gattgactgg tattcttaac tatgttgctc cttttacgct atgtggatac
     1621 gctgctttaa tgcctttgta tcatgctatt gcttcccgta tggctttcat tttctcctcc
     1681 ttgtataaat cctggttagt tcttgccacg gcggaactca tcgccgcctg ccttgcccgc
     1741 tgctggacag gggctcggct gttgggcact gacaattccg tggctcgaga gatcttcgac
     1801 tgtgccttct agttgccagc catctgttgt ttgcccctcc cccgtgcctt ccttgaccct
     1861 ggaaggtgcc actcccactg tcctttccta ataaaatgag gaaattgcat cgcattgtct
     1921 gagtaggtgt cattctattc tggggggtgg ggtggggcag gacagcaagg gggaggattg
     1981 ggaagacaat agcaggcatg agatctcacg tgcggaccga gcggccgcag gaacccctag
     2041 tgatggagtt ggccactccc tctctgcgcg ctcgctcgct cactgaggcc gggcgaccaa
     2101 aggtcgcccg acgcccgggc tttgcccggg cggcctcagt gagcgagcga gcgcgcagct
     2161 gcctgcaggg gcgcctgatg cggtattttc tccttacgca tctgtgcggt atttcacacc
     2221 gcatacgtca aagcaaccat agtacgcgcc ctgtagcggc gcattaagcg cggcgggtgt
     2281 ggtggttacg cgcagcgtga ccgctacact tgccagcgcc ttagcgcccg ctcctttcgc
     2341 tttcttccct tcctttctcg ccacgttcgc cggctttccc cgtcaagctc taaatcgggg
     2401 gctcccttta gggttccgat ttagtgcttt acggcacctc gaccccaaaa aacttgattt
     2461 gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt
     2521 ggagtccacg ttctttaata gtggactctt gttccaaact ggaacaacac tcaactctat
     2581 ctcgggctat tcttttgatt tataagggat tttgccgatt tcggtctatt ggttaaaaaa
     2641 tgagctgatt taacaaaaat ttaacgcgaa ttttaacaaa atattaacgt ttacaatttt
     2701 atggtgcact ctcagtacaa tctgctctga tgccgcatag ttaagccagc cccgacaccc
     2761 gccaacaccc gctgacgcgc cctgacgggc ttgtctgctc ccggcatccg cttacagaca
     2821 agctgtgacc gtctccggga gctgcatgtg tcagaggttt tcaccgtcat caccgaaacg
     2881 cgcgagacga aagggcctcg tgatacgcct atttttatag gttaatgtca tgataataat
     2941 ggtttcttag acgtcaggtg gcacttttcg gggaaatgtg cgcggaaccc ctatttgttt
     3001 atttttctaa atacattcaa atatgtatcc gctcatgaga caataaccct gataaatgct
     3061 tcaataatat tgaaaaagga agagtatgag tattcaacat ttccgtgtcg cccttattcc
     3121 cttttttgcg gcattttgcc ttcctgtttt tgctcaccca gaaacgctgg tgaaagtaaa
     3181 agatgctgaa gatcagttgg gtgcacgagt gggttacatc gaactggatc tcaacagcgg
     3241 taagatcctt gagagttttc gccccgaaga acgttttcca atgatgagca cttttaaagt
     3301 tctgctatgt ggcgcggtat tatcccgtat tgacgccggg caagagcaac tcggtcgccg
     3361 catacactat tctcagaatg acttggttga gtactcacca gtcacagaaa agcatcttac
     3421 ggatggcatg acagtaagag aattatgcag tgctgccata accatgagtg ataacactgc
     3481 ggccaactta cttctgacaa cgatcggagg accgaaggag ctaaccgctt ttttgcacaa
     3541 catgggggat catgtaactc gccttgatcg ttgggaaccg gagctgaatg aagccatacc
     3601 aaacgacgag cgtgacacca cgatgcctgt agcaatggca acaacgttgc gcaaactatt
     3661 aactggcgaa ctacttactc tagcttcccg gcaacaatta atagactgga tggaggcgga
     3721 taaagttgca ggaccacttc tgcgctcggc ccttccggct ggctggttta ttgctgataa
     3781 atctggagcc ggtgagcgtg ggtctcgcgg tatcattgca gcactggggc cagatggtaa
     3841 gccctcccgt atcgtagtta tctacacgac ggggagtcag gcaactatgg atgaacgaaa
     3901 tagacagatc gctgagatag gtgcctcact gattaagcat tggtaactgt cagaccaagt
     3961 ttactcatat atactttaga ttgatttaaa acttcatttt taatttaaaa ggatctaggt
     4021 gaagatcctt tttgataatc tcatgaccaa aatcccttaa cgtgagtttt cgttccactg
     4081 agcgtcagac cccgtagaaa agatcaaagg atcttcttga gatccttttt ttctgcgcgt
     4141 aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt tgccggatca
     4201 agagctacca actctttttc cgaaggtaac tggcttcagc agagcgcaga taccaaatac
     4261 tgttcttcta gtgtagccgt agttaggcca ccacttcaag aactctgtag caccgcctac
     4321 atacctcgct ctgctaatcc tgttaccagt ggctgctgcc agtggcgata agtcgtgtct
     4381 taccgggttg gactcaagac gatagttacc ggataaggcg cagcggtcgg gctgaacggg
     4441 gggttcgtgc acacagccca gcttggagcg aacgacctac accgaactga gatacctaca
     4501 gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca ggtatccggt
     4561 aagcggcagg gtcggaacag gagagcgcac gagggagctt ccagggggaa acgcctggta
     4621 tctttatagt cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc
     4681 gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc
     4741 cttttgctgg ccttttgctc acatgt
//
