LOCUS V1_26973 6225 bp DNA circular SYN 29-SEP-2026 DEFINITION V1_hSyn_ChR2_H134R_EYFP_baseline. ACCESSION V1_26973 VERSION V1_26973 KEYWORDS . SOURCE synthetic DNA construct ORGANISM synthetic DNA construct COMMENT Exact modular reconstruction of Addgene #26973 sequence 199313. COMMENT Display origin rotated to source base 6190. No nucleotide edits. COMMENT Original source sequence and original coordinates retained in parts manifest. FEATURES Location/Qualifiers source 1..6225 /organism="synthetic DNA construct" /mol_type="other DNA" promoter 1..448 /label="hSyn promoter - 26973" /note="Human synapsin I promoter selected for neuronal expression in the proposed cortical-organoid system. The annotation crosses the reference plasmid origin; both segments are retained. Source: Addgene #26973, pAAV-hSyn-hChR2(H134R)-EYFP, Addgene-verified sequence 199313. https://www.addgene.org/26973/sequences/ Source coordinates (1-based, inclusive): 6190..6225,1..412; 448 bp; forward strand. Exact source sequence, no optimization." misc_feature 449..491 /label="5UTR-Kozak - 26973" /note="Reference upstream leader/cloning region, including the GCCACC sequence immediately before the CDS. Operational part boundary, not a measured transcript start. The complete annotated Kozak motif extends into the CDS (source 450-459); the ATG is included only in the CDS. Source: Addgene #26973, pAAV-hSyn-hChR2(H134R)-EYFP, Addgene-verified sequence 199313. https://www.addgene.org/26973/sequences/ Source coordinates (1-based, inclusive): 413..455; 43 bp; forward strand. Exact source sequence, no optimization." CDS 492..2153 /label="ChR2(H134R)-EYFP CDS" /note="Existing Kernel part 47151a2d-d22e-4120-ba89-30354c83fce9. Reuse without creating a duplicate. Humanized ChR2(H134R)-EYFP fusion with its PAAAT linker, ATG start, and TAA stop. Source: Addgene #26973, pAAV-hSyn-hChR2(H134R)-EYFP, Addgene-verified sequence 199313. https://www.addgene.org/26973/sequences/ Source coordinates (1-based, inclusive): 456..2117; 1662 bp; forward strand. Exact source sequence, no optimization." /codon_start=1 /translation="MDYGGALSAVGRELLFVTNPVVVNGSVLVPEDQCYCAGWIESRG TNGAQTASNVLQWLAAGFSILLLMFYAYQTWKSTCGWEEIYVCAIEMVKVILEFFFEF KNPSMLYLATGHRVQWLRYAEWLLTCPVILIRLSNLTGLSNDYSRRTMGLLVSDIGTI VWGATSAMATGYVKVIFFCLGLCYGANTFFHAAKAYIEGYHTVPKGRCRQVVTGMAWL FFVSWGMFPILFILGPEGFGVLSVYGSTVGHTIIDLMSKNCWGLLGHYLRVLIHEHIL IHGDIRKTTKLNIGGTEIEVETLVEDEAEAGAVPAAATMVSKGEELFTGVVPILVELD GDVNGHKFSVSGEGEGDATYGKLTLKFICTTGKLPVPWPTLVTTFGYGLQCFARYPDH MKQHDFFKSAMPEGYVQERTIFFKDDGNYKTRAEVKFEGDTLVNRIELKGIDFKEDGN ILGHKLEYNYNSHNVYIMADKQKNGIKVNFKIRHNIEDGSVQLADHYQQNTPIGDGPV LLPDNHYLSYQSALSKDPNEKRDHMVLLEFVTAAGITLGMDELYK" misc_feature 2154..2177 /label="CDS-WPRE junction - 26973" /note="Preserved reference junction after the CDS and before WPRE. Retained to reconstruct the reference exactly; no independent regulatory function is claimed. Source: Addgene #26973, pAAV-hSyn-hChR2(H134R)-EYFP, Addgene-verified sequence 199313. https://www.addgene.org/26973/sequences/ Source coordinates (1-based, inclusive): 2118..2141; 24 bp; forward strand. Exact source sequence, no optimization." misc_feature 2178..2766 /label="WPRE - 26973" /note="Reference woodchuck hepatitis virus posttranscriptional regulatory element, retained as the baseline posttranscriptional expression-support element. It is not assigned an independent protein-coding role. Source: Addgene #26973, pAAV-hSyn-hChR2(H134R)-EYFP, Addgene-verified sequence 199313. https://www.addgene.org/26973/sequences/ Source coordinates (1-based, inclusive): 2142..2730; 589 bp; forward strand. Exact source sequence, no optimization." misc_feature 2767..2797 /label="WPRE-polyA junction - 26973" /note="Preserved reference junction between WPRE and the hGH poly(A) annotation; no independent regulatory function is claimed. Source: Addgene #26973, pAAV-hSyn-hChR2(H134R)-EYFP, Addgene-verified sequence 199313. https://www.addgene.org/26973/sequences/ Source coordinates (1-based, inclusive): 2731..2761; 31 bp; forward strand. Exact source sequence, no optimization." polyA_signal 2798..3274 /label="hGH polyA - 26973" /note="Reference human growth hormone polyadenylation region, selected to provide the baseline transcript 3-prime processing context. Source: Addgene #26973, pAAV-hSyn-hChR2(H134R)-EYFP, Addgene-verified sequence 199313. https://www.addgene.org/26973/sequences/ Source coordinates (1-based, inclusive): 2762..3238; 477 bp; forward strand. Exact source sequence, no optimization." misc_feature 3275..6225 /label="pAAV vector context - 26973" /note="Reused reference vector context, including both AAV2 ITR annotations, bacterial replication origin, AmpR selection cassette, f1 origin, and intervening sequences. This is a composite context part spanning the bacterial backbone between the two ITRs; it is not all packaged AAV payload. Source: Addgene #26973, pAAV-hSyn-hChR2(H134R)-EYFP, Addgene-verified sequence 199313. https://www.addgene.org/26973/sequences/ Source coordinates (1-based, inclusive): 3239..6189; 2951 bp; forward strand. Exact source sequence, no optimization." regulatory 486..495 /label="Kozak sequence" /note="Original annotation overlaps the 5UTR-Kozak part and the CDS start. Original coordinates: 450..459." misc_feature 492..1418 /label="ChR2(H134R) domain" /note="Source-annotated coding segment of the full fusion. Original coordinates: 456..1382." misc_feature 1419..1433 /label="PAAAT linker" /note="Existing in-frame linker between ChR2 and EYFP. Original coordinates: 1383..1397." misc_feature 1434..2153 /label="EYFP domain" /note="C-terminal fluorescent domain, including terminal stop. Original coordinates: 1398..2117." repeat_region 3314..3454 /label="AAV2 ITR downstream" /note="Source annotation; the shorter overlapping ITR call is not a separate repeat. Original coordinates: 3278..3418." rep_origin 3529..3984 /label="f1 ori" /note="Source f1 origin annotation. Original coordinates: 3493..3948." promoter 4266..4370 /label="AmpR promoter" /note="Bacterial selection-cassette promoter. Original coordinates: 4230..4334." CDS 4371..5231 /label="AmpR" /note="Source beta-lactamase coding region. Original coordinates: 4335..5195." rep_origin 5402..5990 /label="Bacterial ori" /note="Source high-copy-number ColE1/pMB1/pBR322/pUC origin annotation. Original coordinates: 5366..5954." repeat_region 6052..6181 /label="AAV2 ITR upstream (alternate)" /note="Source annotation describes a functional equivalent of wild-type AAV2 ITR; retained unchanged. Original coordinates: 6016..6145." ORIGIN 1 agtgcaagtg ggttttagga ccaggatgag gcggggtggg ggtgcctacc tgacgaccga 61 ccccgaccca ctggacaagc acccaacccc cattccccaa attgcgcatc ccctatcaga 121 gagggggagg ggaaacagga tgcggcgagg cgcgtgcgca ctgccagctt cagcaccgcg 181 gacagtgcct tcgcccccgc ctggcggcgc gcgccaccgc cgcctcagca ctgaaggcgc 241 gctgacgtca ctcgccggtc ccccgcaaac tccccttccc ggccaccttg gtcgcgtccg 301 cgccgccgcc ggcccagccg gaccgcacca cgcgaggcgc gagatagggg ggcacgggcg 361 cgaccatctg cgctgcggcg ccggcgactc agcgctgcct cagtctgcgg tgggcagcgg 421 aggagtcgtg tcgtgcctga gagcgcagtc gagaaggtac cggatcctct agagtcgact 481 ccggagccac catggactat ggcggcgctt tgtctgccgt cggacgcgaa cttttgttcg 541 ttactaatcc tgtggtggtg aacgggtccg tcctggtccc tgaggatcaa tgttactgtg 601 ccggatggat tgaatctcgc ggcacgaacg gcgctcagac cgcgtcaaat gtcctgcagt 661 ggcttgcagc aggattcagc attttgctgc tgatgttcta tgcctaccaa acctggaaat 721 ctacatgcgg ctgggaggag atctatgtgt gcgccattga aatggttaag gtgattctcg 781 agttcttttt tgagtttaag aatccctcta tgctctacct tgccacagga caccgggtgc 841 agtggctgcg ctatgcagag tggctgctca cttgtcctgt catccttatc cgcctgagca 901 acctcaccgg cctgagcaac gactacagca ggagaaccat gggactcctt gtctcagaca 961 tcgggactat cgtgtggggg gctaccagcg ccatggcaac cggctatgtt aaagtcatct 1021 tcttttgtct tggattgtgc tatggcgcga acacattttt tcacgccgcc aaagcatata 1081 tcgagggtta tcatactgtg ccaaagggtc ggtgccgcca ggtcgtgacc ggcatggcat 1141 ggctgttttt cgtgagctgg ggtatgttcc caattctctt cattttgggg cccgaaggtt 1201 ttggcgtcct gagcgtctat ggctccaccg taggtcacac gattattgat ctgatgagta 1261 aaaattgttg ggggttgttg ggacactacc tgcgcgtcct gatccacgag cacatattga 1321 ttcacggaga tatccgcaaa accaccaaac tgaacatcgg cggaacggag atcgaggtcg 1381 agactctcgt cgaagacgaa gccgaggccg gagccgtgcc agcggccgcc accatggtga 1441 gcaagggcga ggagctgttc accggggtgg tgcccatcct ggtcgagctg gacggcgacg 1501 taaacggcca caagttcagc gtgtccggcg agggcgaggg cgatgccacc tacggcaagc 1561 tgaccctgaa gttcatctgc accaccggca agctgcccgt gccctggccc accctcgtga 1621 ccaccttcgg ctacggcctg cagtgcttcg cccgctaccc cgaccacatg aagcagcacg 1681 acttcttcaa gtccgccatg cccgaaggct acgtccagga gcgcaccatc ttcttcaagg 1741 acgacggcaa ctacaagacc cgcgccgagg tgaagttcga gggcgacacc ctggtgaacc 1801 gcatcgagct gaagggcatc gacttcaagg aggacggcaa catcctgggg cacaagctgg 1861 agtacaacta caacagccac aacgtctata tcatggccga caagcagaag aacggcatca 1921 aggtgaactt caagatccgc cacaacatcg aggacggcag cgtgcagctc gccgaccact 1981 accagcagaa cacccccatc ggcgacggcc ccgtgctgct gcccgacaac cactacctga 2041 gctaccagtc cgccctgagc aaagacccca acgagaagcg cgatcacatg gtcctgctgg 2101 agttcgtgac cgccgccggg atcactctcg gcatggacga gctgtacaag taagaattcg 2161 atatcaagct tatcgataat caacctctgg attacaaaat ttgtgaaaga ttgactggta 2221 ttcttaacta tgttgctcct tttacgctat gtggatacgc tgctttaatg cctttgtatc 2281 atgctattgc ttcccgtatg gctttcattt tctcctcctt gtataaatcc tggttgctgt 2341 ctctttatga ggagttgtgg cccgttgtca ggcaacgtgg cgtggtgtgc actgtgtttg 2401 ctgacgcaac ccccactggt tggggcattg ccaccacctg tcagctcctt tccgggactt 2461 tcgctttccc cctccctatt gccacggcgg aactcatcgc cgcctgcctt gcccgctgct 2521 ggacaggggc tcggctgttg ggcactgaca attccgtggt gttgtcgggg aaatcatcgt 2581 cctttccttg gctgctcgcc tatgttgcca cctggattct gcgcgggacg tccttctgct 2641 acgtcccttc ggccctcaat ccagcggacc ttccttcccg cggcctgctg ccggctctgc 2701 ggcctcttcc gcgtcttcgc cttcgccctc agacgagtcg gatctccctt tgggccgcct 2761 ccccgcatcg ataccgagcg ctgctcgaga gatctacggg tggcatccct gtgacccctc 2821 cccagtgcct ctcctggccc tggaagttgc cactccagtg cccaccagcc ttgtcctaat 2881 aaaattaagt tgcatcattt tgtctgacta ggtgtccttc tataatatta tggggtggag 2941 gggggtggta tggagcaagg ggcaagttgg gaagacaacc tgtagggcct gcggggtcta 3001 ttgggaacca agctggagtg cagtggcaca atcttggctc actgcaatct ccgcctcctg 3061 ggttcaagcg attctcctgc ctcagcctcc cgagttgttg ggattccagg catgcatgac 3121 caggctcagc taatttttgt ttttttggta gagacggggt ttcaccatat tggccaggct 3181 ggtctccaac tcctaatctc aggtgatcta cccaccttgg cctcccaaat tgctgggatt 3241 acaggcgtga accactgctc ccttccctgt ccttctgatt ttgtaggtaa ccacgtgcgg 3301 accgagcggc cgcaggaacc cctagtgatg gagttggcca ctccctctct gcgcgctcgc 3361 tcgctcactg aggccgggcg accaaaggtc gcccgacgcc cgggctttgc ccgggcggcc 3421 tcagtgagcg agcgagcgcg cagctgcctg caggggcgcc tgatgcggta ttttctcctt 3481 acgcatctgt gcggtatttc acaccgcata cgtcaaagca accatagtac gcgccctgta 3541 gcggcgcatt aagcgcggcg ggtgtggtgg ttacgcgcag cgtgaccgct acacttgcca 3601 gcgccttagc gcccgctcct ttcgctttct tcccttcctt tctcgccacg ttcgccggct 3661 ttccccgtca agctctaaat cgggggctcc ctttagggtt ccgatttagt gctttacggc 3721 acctcgaccc caaaaaactt gatttgggtg atggttcacg tagtgggcca tcgccctgat 3781 agacggtttt tcgccctttg acgttggagt ccacgttctt taatagtgga ctcttgttcc 3841 aaactggaac aacactcaac tctatctcgg gctattcttt tgatttataa gggattttgc 3901 cgatttcggt ctattggtta aaaaatgagc tgatttaaca aaaatttaac gcgaatttta 3961 acaaaatatt aacgtttaca attttatggt gcactctcag tacaatctgc tctgatgccg 4021 catagttaag ccagccccga cacccgccaa cacccgctga cgcgccctga cgggcttgtc 4081 tgctcccggc atccgcttac agacaagctg tgaccgtctc cgggagctgc atgtgtcaga 4141 ggttttcacc gtcatcaccg aaacgcgcga gacgaaaggg cctcgtgata cgcctatttt 4201 tataggttaa tgtcatgata ataatggttt cttagacgtc aggtggcact tttcggggaa 4261 atgtgcgcgg aacccctatt tgtttatttt tctaaataca ttcaaatatg tatccgctca 4321 tgagacaata accctgataa atgcttcaat aatattgaaa aaggaagagt atgagtattc 4381 aacatttccg tgtcgccctt attccctttt ttgcggcatt ttgccttcct gtttttgctc 4441 acccagaaac gctggtgaaa gtaaaagatg ctgaagatca gttgggtgca cgagtgggtt 4501 acatcgaact ggatctcaac agcggtaaga tccttgagag ttttcgcccc gaagaacgtt 4561 ttccaatgat gagcactttt aaagttctgc tatgtggcgc ggtattatcc cgtattgacg 4621 ccgggcaaga gcaactcggt cgccgcatac actattctca gaatgacttg gttgagtact 4681 caccagtcac agaaaagcat cttacggatg gcatgacagt aagagaatta tgcagtgctg 4741 ccataaccat gagtgataac actgcggcca acttacttct gacaacgatc ggaggaccga 4801 aggagctaac cgcttttttg cacaacatgg gggatcatgt aactcgcctt gatcgttggg 4861 aaccggagct gaatgaagcc ataccaaacg acgagcgtga caccacgatg cctgtagcaa 4921 tggcaacaac gttgcgcaaa ctattaactg gcgaactact tactctagct tcccggcaac 4981 aattaataga ctggatggag gcggataaag ttgcaggacc acttctgcgc tcggcccttc 5041 cggctggctg gtttattgct gataaatctg gagccggtga gcgtgggtct cgcggtatca 5101 ttgcagcact ggggccagat ggtaagccct cccgtatcgt agttatctac acgacgggga 5161 gtcaggcaac tatggatgaa cgaaatagac agatcgctga gataggtgcc tcactgatta 5221 agcattggta actgtcagac caagtttact catatatact ttagattgat ttaaaacttc 5281 atttttaatt taaaaggatc taggtgaaga tcctttttga taatctcatg accaaaatcc 5341 cttaacgtga gttttcgttc cactgagcgt cagaccccgt agaaaagatc aaaggatctt 5401 cttgagatcc tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac 5461 cagcggtggt ttgtttgccg gatcaagagc taccaactct ttttccgaag gtaactggct 5521 tcagcagagc gcagatacca aatactgttc ttctagtgta gccgtagtta ggccaccact 5581 tcaagaactc tgtagcaccg cctacatacc tcgctctgct aatcctgtta ccagtggctg 5641 ctgccagtgg cgataagtcg tgtcttaccg ggttggactc aagacgatag ttaccggata 5701 aggcgcagcg gtcgggctga acggggggtt cgtgcacaca gcccagcttg gagcgaacga 5761 cctacaccga actgagatac ctacagcgtg agctatgaga aagcgccacg cttcccgaag 5821 ggagaaaggc ggacaggtat ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg 5881 agcttccagg gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc cacctctgac 5941 ttgagcgtcg atttttgtga tgctcgtcag gggggcggag cctatggaaa aacgccagca 6001 acgcggcctt tttacggttc ctggcctttt gctggccttt tgctcacatg tcctgcaggc 6061 agctgcgcgc tcgctcgctc actgaggccg cccgggcgtc gggcgacctt tggtcgcccg 6121 gcctcagtga gcgagcgagc gcgcagagag ggagtggcca actccatcac taggggttcc 6181 tgcggccgca cgcgtgtgtc tagactgcag agggccctgc gtatg //