LOCUS       sequence-447683-        7395 bp    DNA     circular SYN 15-APR-2025
DEFINITION  Cation channelrhodopsin ChRmine targeted to the neuronal soma and 
            proximal dendrites in a viral vector.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7395)
  AUTHORS   Adesnik H
  TITLE     Miscellaneous Optogenetic Tools
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 7395)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: 
            "Unpublished"
COMMENT     Sequence Label: pAAV-hSyn-DIO-GCaMP6m-p2A-ChRmine-ST
FEATURES             Location/Qualifiers
     source          1..7395
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    228..249
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(264..269,270..287,288..294)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    302..318
                     /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     307..329
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     326..342
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     326..342
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     repeat_region   380..509
                     /label=AAV2 ITR
     promoter        591..1037
                     /label=hSyn promoter
                     /note="human synapsin I promoter; confers neuron-specific 
                     expression (Kugler et al., 2003)"
     protein_bind    complement(1083..1116)
                     /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)."
     protein_bind    complement(1167..1200)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     CDS             complement(1212..1238)
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             complement(2430..2486)
                     /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"
     primer_bind     3058..3077
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3317..3338
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(3444..3465)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             complement(3678..3734)
                     /codon_start=1
                     /product="calmodulin-binding peptide from avian smooth 
                     muscle myosin light chain kinase"
                     /label=calmodulin-binding peptide
                     /translation="RRKWNKTGHAVRAIGRLSS"
     regulatory      3746..3755
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     protein_bind    3762..3795
                     /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)."
     protein_bind    3846..3879
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     misc_feature    3904..4492
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3957..3977)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(4375..4386)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(4495..4511)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4496..4512)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     polyA_signal    4541..4662
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4552..4571)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     4606..4625
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     repeat_region   4824..4953
                     /label=AAV2 ITR
     primer_bind     complement(4972..4989)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4972..4988)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4981..5003)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      5130..5585
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5217..5236)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5427..5448
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        5611..5715
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5716..5784,5785..6576)
                     /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(5934..5953)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6747..7335
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7236..7255
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 ttctttcctg cgttatcccc tgattctgtg gataaccgta ttaccgcctt tgagtgagct
       61 gataccgctc gccgcagccg aacgaccgag cgcagcgagt cagtgagcga ggaagcggaa
      121 gagcgcccaa tacgcaaacc gcctctcccc gcgcgttggc cgattcatta atgcagctgg
      181 cacgacaggt ttcccgactg gaaagcgggc agtgagcgca acgcaattaa tgtgagttag
      241 ctcactcatt aggcacccca ggctttacac tttatgcttc cggctcgtat gttgtgtgga
      301 attgtgagcg gataacaatt tcacacagga aacagctatg accatgatta cgccagattt
      361 aattaaggcc ttaattaggc tgcgcgctcg ctcgctcact gaggccgccc gggcaaagcc
      421 cgggcgtcgg gcgacctttg gtcgcccggc ctcagtgagc gagcgagcgc gcagagaggg
      481 agtggccaac tccatcacta ggggttcctt gtagttaatg attaacccgc catgctactt
      541 atctacgtag ccatgctcta ggaagatctc tgcagagggc cctgcgtatg agtgcaagtg
      601 ggttttagga ccaggatgag gcggggtggg ggtgcctacc tgacgaccga ccccacccac
      661 tggacaagca cccaaccccc attccccaaa ttgcgcatcc cctatcagag agggggaggg
      721 gaaacaggat gcggcgaggc gcgtgcgcac tgccagcttc agcaccgcgg acagtgcctt
      781 cgcccccgcc tggcggcgcg cgccaccgcc gcctcagcac tgaaggcgcg ctgacgtcac
      841 tcgccggtcc cccgcaaact ccccttcccg gccaccttgg tcgcgtccgc gccgccgccg
      901 gcccagccgg accgcaccac gcgaggcgcg agataggggg gcacgggcgc gaccatctgc
      961 gctgcggcgc cggcgactca gcgctgcctc agtctgcggt gggcagcgga ggagtcgtgt
     1021 cgtgcctgag agcgcagtcg aattcaagct gatagcaagg atcctctaga gtcgactccg
     1081 gaataacttc gtataggata ctttatacga agttatgcag aatggtagct ggattgtagc
     1141 tgctattagc aatatgaaac ctcttaataa cttcgtatag catacattat acgaagttat
     1201 ggcgcgcctt aagcgtaatc tggaacatcg tatgggtaga atctggtggc ttcagggaag
     1261 tccgtggcac agctgatgaa gctgtcaatg ctggacatgc tccgcatgtc gatgacgccc
     1321 tccgtgcgtg cgggcagagg ggccacgggg ctgggcatgc tgctcatctc cagctcctct
     1381 ggaggcttgc tctgcggggc catctccttg gtgttgagga tgggctggga ctgcacgttg
     1441 atgtcgatct ggtccagggg gatgtactcg ccctcgctgg tgatcctgct cttggcggcc
     1501 gcagacagtc tccgcaggcg ctcgccattg ctgctcagcc tcatatccag ctccagcttc
     1561 tggaactcat cctcgtcgtg tcccagctgc tccagcagcc gaaacagctc ctcatcatag
     1621 gtctttctga accgggccag ggcaaagccg taacaggact tggccacgat gtcgcacacg
     1681 cagtgggcca cctctgtcca gttatcgtcg atccagccga agcctctagg gcagatggcc
     1741 cacaccaggg gaaacagaat ccacagcatg gagaaggtaa acacggcgaa cttcagcacg
     1801 tgcaggctgt gctcggcgcc tgtatcctgg gccagctggg ccagccggct atactgcttg
     1861 gccacgatgc tcatcacgat agagtaggcg aagataaacc agaaacagcc aaagccatac
     1921 caggcgtagg cgccattcct cagcctaggg tcgccctctg cgtagaactc ggccagcacg
     1981 ccagacatca ggatggcaaa gatgatggcg ctacaagaca cgcgataggg ggccctcagc
     2041 tgatacagca gatcgtacac cagcattggg caggtcagca tgtagtcggc atacctgtaa
     2101 ttgccgaagc cgatgatgaa ggggcccaca tcgccgaagt tgtggcccag tgtgtcggcc
     2161 acgatcttgc cgaagtagct cagcatggca aagaaattca caaacagggc caggttctgc
     2221 tcctcctgat agccggcgta aaaggaggtt ctcttgctag ggccccgaga tgtccaggac
     2281 tcatagccca ggtagaatgt ggtgattgtc agcacgataa agcaagacca gtgggcgccg
     2341 atggccacgg agttcagctt ggtatccaga taccagccgt ccattgtgcc cacgtagaac
     2401 atctggtcgg tgcctggtgc gtgtgccatg ggaccggggt tttcttccac gtctcctgct
     2461 tgctttaaca gagagaagtt cgtggctccg gagccactag tcttcgctgt catcatttgt
     2521 acaaactctt cgtagtttac ctgaccatcc ccatcgatgt ctgcttccct gatcatttca
     2581 tcaacctctt catctgttaa cttctctcca aggtttgtca tcacgtggcg aagctctgct
     2641 gcactgatgt agccattgcc atccttatca aacacaccga acgcttctct aatttcttct
     2701 tccgtgtccc tgtagctccc ttttcttgcc atcattgtca ggaactcagg gaagtcgatt
     2761 gtgccgtcac cgtcggcatc tacttcattg atcatgtcct gcagctctgc ttctgtgggg
     2821 ttctgcccca gagaccgcat caccgtcccc agctccttgg ttgttattgt cccatccccg
     2881 tccttgtcaa atagggagaa agcctcttta aattctgcga tctgctcttc agtcagttgg
     2941 tccggcaggt tgtactccag cttgtgcccc aggatgttgc cgtcctcctt gaagtcgatg
     3001 cccttcagct cgatgcggtt caccagggtg tcgccctcga acttcacctc ggcgcgggtc
     3061 ttgtagttgc cgtcgtcctt gaagaagatg gtgcgctcct ggatgtagcc ttcgggcatg
     3121 gcggacttga agaagtcgtg ctgcttcatg tggtcggggt agcggctgaa gcactgcacg
     3181 ccgtaggtca gggtggtcac gagggtgggc cagggcacgg gcagcttgcc ggtggtgcag
     3241 atgaacttca gggtcagctt gccgtaggtg gcatcgccct caccctcgcc ggacacgctg
     3301 aacttgtggc cgtttacgtc gccgtccagc tcgaccagga tgggcaccac cccggtgaac
     3361 agctcctcgc ccttgctcac catgctccct ccggtaccgc ccttgtacag ctcgtccatg
     3421 ccgagagtga tcccggcggc ggtcacgaac tccagcagga ccatgtgatc gcgcttctcg
     3481 ttggggtctt tcgaaagttt ggactgcacg ctcaggtagt ggttgtcggg cagcagcacg
     3541 gggccgtcgc cgatgggggt gttctgctgg tagtggtagg cgagctgcac gccgccgtcc
     3601 tcgatgttgt ggcggatctt gaagttcgcc ttgatgccgt tcttctgctt gtcggccttg
     3661 atatagacgt tctcgagtga gctcagccga cctatagctc tgactgcgtg acctgtctta
     3721 ttccacttac gacgtgatga gtccaccatg gtggcgctag cataacttcg tataaagtat
     3781 cctatacgaa gttatttgcc ttaacccaga aattatcact gttattcttt agaatggtgc
     3841 aaagaataac ttcgtataat gtatgctata cgaagttatg aattcgatat caagcttatc
     3901 gataatcaac ctctggatta caaaatttgt gaaagattga ctggtattct taactatgtt
     3961 gctcctttta cgctatgtgg atacgctgct ttaatgcctt tgtatcatgc tattgcttcc
     4021 cgtatggctt tcattttctc ctccttgtat aaatcctggt tgctgtctct ttatgaggag
     4081 ttgtggcccg ttgtcaggca acgtggcgtg gtgtgcactg tgtttgctga cgcaaccccc
     4141 actggttggg gcattgccac cacctgtcag ctcctttccg ggactttcgc tttccccctc
     4201 cctattgcca cggcggaact catcgccgcc tgccttgccc gctgctggac aggggctcgg
     4261 ctgttgggca ctgacaattc cgtggtgttg tcggggaaat catcgtcctt tccttggctg
     4321 ctcgcctgtg ttgccacctg gattctgcgc gggacgtcct tctgctacgt cccttcggcc
     4381 ctcaatccag cggaccttcc ttcccgcggc ctgctgccgg ctctgcggcc tcttccgcgt
     4441 cttcgccttc gccctcagac gagtcggatc tccctttggg ccgcctcccc gcatcgatac
     4501 cgtcgacctc gacccgggcg gccgcttcga gcagacatga taagatacat tgatgagttt
     4561 ggacaaacca caactagaat gcagtgaaaa aaatgcttta tttgtgaaat ttgtgatgct
     4621 attgctttat ttgtaaccat tataagctgc aataaacaag ttaacaacaa caattgcatt
     4681 cattttatgt ttcaggttca gggggagatg tgggaggttt tttaaagcaa gtaaaacctc
     4741 tacaaatgtg gtaaaatcga taaggatctt cctagagcat ggctacgtag ataagtagca
     4801 tggcgggtta atcattaact acaaggaacc cctagtgatg gagttggcca ctccctctct
     4861 gcgcgctcgc tcgctcactg aggccgggcg accaaaggtc gcccgacgcc cgggctttgc
     4921 ccgggcggcc tcagtgagcg agcgagcgcg cagccttaat taacctaatt cactggccgt
     4981 cgttttacaa cgtcgtgact gggaaaaccc tggcgttacc caacttaatc gccttgcagc
     5041 acatccccct ttcgccagct ggcgtaatag cgaagaggcc cgcaccgatc gcccttccca
     5101 acagttgcgc agcctgaatg gcgaatggga cgcgccctgt agcggcgcat taagcgcggc
     5161 gggtgtggtg gttacgcgca gcgtgaccgc tacacttgcc agcgccctag cgcccgctcc
     5221 tttcgctttc ttcccttcct ttctcgccac gttcgccggc tttccccgtc aagctctaaa
     5281 tcgggggctc cctttagggt tccgatttag tgctttacgg cacctcgacc ccaaaaaact
     5341 tgattagggt gatggttcac gtagtgggcc atcgccctga tagacggttt ttcgcccttt
     5401 gacgttggag tccacgttct ttaatagtgg actcttgttc caaactggaa caacactcaa
     5461 ccctatctcg gtctattctt ttgatttata agggattttg ccgatttcgg cctattggtt
     5521 aaaaaatgag ctgatttaac aaaaatttaa cgcgaatttt aacaaaatat taacgcttac
     5581 aatttaggtg gcacttttcg gggaaatgtg cgcggaaccc ctatttgttt atttttctaa
     5641 atacattcaa atatgtatcc gctcatgaga caataaccct gataaatgct tcaataatat
     5701 tgaaaaagga agagtatgag tattcaacat ttccgtgtcg cccttattcc cttttttgcg
     5761 gcattttgcc ttcctgtttt tgctcaccca gaaacgctgg tgaaagtaaa agatgctgaa
     5821 gatcagttgg gtgcacgagt gggttacatc gaactggatc tcaacagcgg taagatcctt
     5881 gagagttttc gccccgaaga acgttttcca atgatgagca cttttaaagt tctgctatgt
     5941 ggcgcggtat tatcccgtat tgacgccggg caagagcaac tcggtcgccg catacactat
     6001 tctcagaatg acttggttga gtactcacca gtcacagaaa agcatcttac ggatggcatg
     6061 acagtaagag aattatgcag tgctgccata accatgagtg ataacactgc ggccaactta
     6121 cttctgacaa cgatcggagg accgaaggag ctaaccgctt ttttgcacaa catgggggat
     6181 catgtaactc gccttgatcg ttgggaaccg gagctgaatg aagccatacc aaacgacgag
     6241 cgtgacacca cgatgcctgt agcaatggca acaacgttgc gcaaactatt aactggcgaa
     6301 ctacttactc tagcttcccg gcaacaatta atagactgga tggaggcgga taaagttgca
     6361 ggaccacttc tgcgctcggc ccttccggct ggctggttta ttgctgataa atctggagcc
     6421 ggtgagcgtg ggtctcgcgg tatcattgca gcactggggc cagatggtaa gccctcccgt
     6481 atcgtagtta tctacacgac ggggagtcag gcaactatgg atgaacgaaa tagacagatc
     6541 gctgagatag gtgcctcact gattaagcat tggtaactgt cagaccaagt ttactcatat
     6601 atactttaga ttgatttaaa acttcatttt taatttaaaa ggatctaggt gaagatcctt
     6661 tttgataatc tcatgaccaa aatcccttaa cgtgagtttt cgttccactg agcgtcagac
     6721 cccgtagaaa agatcaaagg atcttcttga gatccttttt ttctgcgcgt aatctgctgc
     6781 ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt tgccggatca agagctacca
     6841 actctttttc cgaaggtaac tggcttcagc agagcgcaga taccaaatac tgttcttcta
     6901 gtgtagccgt agttaggcca ccacttcaag aactctgtag caccgcctac atacctcgct
     6961 ctgctaatcc tgttaccagt ggctgctgcc agtggcgata agtcgtgtct taccgggttg
     7021 gactcaagac gatagttacc ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc
     7081 acacagccca gcttggagcg aacgacctac accgaactga gatacctaca gcgtgagcta
     7141 tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca ggtatccggt aagcggcagg
     7201 gtcggaacag gagagcgcac gagggagctt ccagggggaa acgcctggta tctttatagt
     7261 cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg
     7321 cggagcctat ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc cttttgctgg
     7381 ccttttgctc acatg
//
