LOCUS       sequence-97014-e        7607 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Co-expresses hM4Dnrxn and a red fluorescent label from a 
            Cre-dependent virus.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7607)
  AUTHORS   Stachniak TJ, Ghosh A, Sternson SM
  TITLE     Chemogenetic synaptic silencing of neural circuits localizes a 
            hypothalamus-->midbrain pathway for feeding behavior.
  JOURNAL   Neuron. 2014 May 21;82(4):797-808. doi: 
            10.1016/j.neuron.2014.04.008. Epub 2014 Apr 24.
  PUBMED    24768300
REFERENCE   2  (bases 1 to 7607)
  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.neuron.2014.04.008"; journalName: "Neuron"; date: 
            "2014-05-21- 21"; volume: "82"; issue: "4"; pages: "797-808"
COMMENT     Sequence Label: AA0250-mCherry-2a-hM4D-nrxn1a rev
FEATURES             Location/Qualifiers
     source          1..7607
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      complement(182..770)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(262..281)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(941..1732,1733..1801))
                     /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     1564..1583
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1802..1906)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(1932..2387)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2069..2090)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2281..2300
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     repeat_region   2459..2588
                     /label=AAV2 ITR
     promoter        2901..3178
                     /label=chicken beta-actin promoter
     primer_bind     3458..3480
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     protein_bind    3596..3629
                     /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    3692..3725
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     CDS             complement(4014..4040)
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             complement(4041..4067)
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             complement(5505..5558)
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     regulatory      6275..6284
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     protein_bind    complement(6330..6363)
                     /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(6434..6467)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     misc_feature    6509..7097
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(6562..6582)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(6980..6991)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    7140..7261
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(7151..7170)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     7205..7224
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     repeat_region   7423..7552
                     /label=AAV2 ITR
ORIGIN
        1 cttccgcttc ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat
       61 cagctcactc aaaggcggta atacggttat ccacagaatc aggggataac gcaggaaaga
      121 acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt
      181 ttttccatag gctccgcccc cctgacgagc atcacaaaaa tcgacgctca agtcagaggt
      241 ggcgaaaccc gacaggacta taaagatacc aggcgtttcc ccctggaagc tccctcgtgc
      301 gctctcctgt tccgaccctg ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa
      361 gcgtggcgct ttctcatagc tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct
      421 ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc ttatccggta
      481 actatcgtct tgagtccaac ccggtaagac acgacttatc gccactggca gcagccactg
      541 gtaacaggat tagcagagcg aggtatgtag gcggtgctac agagttcttg aagtggtggc
      601 ctaactacgg ctacactaga agaacagtat ttggtatctg cgctctgctg aagccagtta
      661 ccttcggaaa aagagttggt agctcttgat ccggcaaaca aaccaccgct ggtagcggtg
      721 gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa gaagatcctt
      781 tgatcttttc tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg
      841 tcatgagatt atcaaaaagg atcttcacct agatcctttt aaattaaaaa tgaagtttta
      901 aatcaatcta aagtatatat gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg
      961 aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga ctccccgtcg
     1021 tgtagataac tacgatacgg gagggcttac catctggccc cagtgctgca atgataccgc
     1081 gagacccacg ctcaccggct ccagatttat cagcaataaa ccagccagcc ggaagggccg
     1141 agcgcagaag tggtcctgca actttatccg cctccatcca gtctattaat tgttgccggg
     1201 aagctagagt aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc attgctacag
     1261 gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt cagctccggt tcccaacgat
     1321 caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc
     1381 cgatcgttgt cagaagtaag ttggccgcag tgttatcact catggttatg gcagcactgc
     1441 ataattctct tactgtcatg ccatccgtaa gatgcttttc tgtgactggt gagtactcaa
     1501 ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac
     1561 gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga aaacgttctt
     1621 cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg taacccactc
     1681 gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg tgagcaaaaa
     1741 caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt tgaatactca
     1801 tactcttcct ttttcaatat tattgaagca tttatcaggg ttattgtctc atgagcggat
     1861 acatatttga atgtatttag aaaaataaac aaataggggt tccgcgcaca tttccccgaa
     1921 aagtgccacc taaattgtaa gcgttaatat tttgttaaaa ttcgcgttaa atttttgtta
     1981 aatcagctca ttttttaacc aataggccga aatcggcaaa atcccttata aatcaaaaga
     2041 atagaccgag atagggttga gtgttgttcc agtttggaac aagagtccac tattaaagaa
     2101 cgtggactcc aacgtcaaag ggcgaaaaac cgtctatcag ggcgatggcc cactacgtga
     2161 accatcaccc taatcaagtt ttttggggtc gaggtgccgt aaagcactaa atcggaaccc
     2221 taaagggagc ccccgattta gagcttgacg gggaaagccg gcgaacgtgg cgagaaagga
     2281 agggaagaaa gcgaaaggag cgggcgctag ggcgctggca agtgtagcgg tcacgctgcg
     2341 cgtaaccacc acacccgccg cgcttaatgc gccgctacag ggcgcgtccc attcgccatt
     2401 caggctgcgc aactgttggg aagggcgatc ggtgcgggcc tcttcgctat tacgccagct
     2461 gcgcgctcgc tcgctcactg aggccgcccg ggcaaagccc gggcgtcggg cgacctttgg
     2521 tcgcccggcc tcagtgagcg agcgagcgcg cagagaggga gtggccaact ccatcactag
     2581 gggttccttg tagttaatga ttaacccgcc atgctactta tctacgtagc catgctctag
     2641 gaagatcgta ccattgacgt caataatgac gtatgttccc atagtaacgc caatagggac
     2701 tttccattga cgtcaatggg tggagtattt acggtaaact gcccacttgg cagtacatca
     2761 agtgtatcat atgccaagta cgccccctat tgacgtcaat gacggtaaat ggcccgcctg
     2821 gcattatgcc cagtacatga ccttatggga ctttcctact tggcagtaca tctacgtatt
     2881 agtcatcgct attaccatgg tcgaggtgag ccccacgttc tgcttcactc tccccatctc
     2941 ccccccctcc ccacccccaa ttttgtattt atttattttt taattatttt gtgcagcgat
     3001 gggggcgggg gggggggggg ggcgcgcgcc aggcggggcg gggcggggcg aggggcgggg
     3061 cggggcgagg cggagaggtg cggcggcagc caatcagagc ggcgcgctcc gaaagtttcc
     3121 ttttatggcg aggcggcggc ggcggcggcc ctataaaaag cgaagcgcgc ggcgggcggg
     3181 agtcgctgcg acgctgcctt cgccccgtgc cccgctccgc cgccgcctcg cgccgcccgc
     3241 cccggctctg actgaccgcg ttactcccac aggtgagcgg gcgggacggc ccttctcctc
     3301 cgggctgtaa ttagcgcttg gtttaatgac ggcttgtttc ttttctgtgg ctgcgtgaaa
     3361 gccttgaggg gctccgggag ggccctttgt gcggggggag cggctcgggg ctgtccgcgg
     3421 ggggacggct gccttcgggg gggacggggc agggcggggt tcggcttctg gcgtgtgacc
     3481 ggcggctcta gagcctctgc taaccatgtt catgccttct tctttttcct acagctcctg
     3541 ggcaacgtgc tggttattgt gctgtctcat cattttggca aagaattgga tccccataac
     3601 ttcgtataaa gtatcctata cgaagttata tcaaaatagg aagaccaatg cttcaccatc
     3661 gacccgaatt gccaagcatc accatcgacc cataacttcg tataatgtat gctatacgaa
     3721 gttatactag agcctctgct aaccatgttc atgccttctt ctttttccta cagctcctgg
     3781 gcaacgtgct ggttattgtg ctgtctcatc attttggcaa agaattcggt acctcgagca
     3841 ccggttcaca catagtactc tttgtcctta ttcttcttgt tcttattcgc tgacttcgcc
     3901 gatgagggct gcttttcctt caccaccgct ccgtttgact gggcggagtt cgagatatag
     3961 ttccgtgact cgtccacatg atacgagcct tcgtcccggt tccgatactt gtaagcgtaa
     4021 tccgggacgt catagggata ggcgtaatcg gggacatcat atgggtatct cgctgtcccg
     4081 atatttcggt actggcacaa caggagatgg cggaacgttt tcttgaacgt ggcgttacac
     4141 aatgcgtagc atgccggatt gatagtggag ttgacgtagc agagccaata cccgatcgac
     4201 cagaccgtgt ctgggatgca cgattgacaa aatgtattca ccaacaccat gacattatac
     4261 ggagtccacg tcaagataaa cgccaggagg atggcgaaaa tcgttcttgt cacctttctc
     4321 tctctcgcgg ccatttgtct tttcttccgc acctgattcc gcgcgattga cgcgaacttt
     4381 ctggcgacgt tggcggctgg cctcatacca gcgggcgttg cgggtacgat ctcgatggct
     4441 gttacgcact cattacccgt ttgtttcgtc acaatctgaa tctttgacca tcgcgacgcg
     4501 gggttcagcg ccctgggctg aagaggcggt gccggcattg ccggcgtcgt cgcttctgtt
     4561 gtgctcaact cggtagcagg tcgctctttc gtattctggg tcgcgctccc cgacgacgac
     4621 tcgttggaag tgtccttatc cgccacgggt cgcggtggag gaggcagcgc cggaggtggc
     4681 gcctcttcca gtttaccatt cctcagctcc tcgcgcgctg cttcaccggg aggaggcttt
     4741 ttcacggact gtttcataag cggtgatttg agaaaagcca atgtcttggc tttcttctcc
     4801 ttaggcccct cgggcctgtg cttgtggacg cggcttcttg acgcaagcga gatgtgaatg
     4861 tacaacacgg tcataatgac gaccggaagg tagaatcccg caatcgcagt cccgaaagtc
     4921 acagcggggt tcgaaaggaa ttggatgaaa cattggttgt ccgggactgt gcgtttaccc
     4981 acgacaaact gccagaacag aatggcagga gcccaaagta cgaacgagag gacccatgca
     5041 gccgcgatca tcagccctgc cattttggtc gttctccttg cggggtaggt caagggtttg
     5101 gttacacaga aatagcgatc aaatgagata atcagaaggt tcatgactga agcattgctc
     5161 accacacagt caagtgcaag ccagaggtca cacacgacgg ctccgagagg ccagtaccct
     5221 ttgatgatgt atactgtata gaggttcatc gagaaagccc caatgatcag atccgcgcaa
     5281 gccagggaga acaagaaata gttgtttacg gtctggagtt gcctgttgac cttaatcgag
     5341 agcatgacca gaatgtttcc cacgacagtc acaagtgaaa ggcttccagt tactgtggcg
     5401 atgaatacca tttctactgt ttcgtagcga ttgtgcgagg acgacgtcac gagccgtacc
     5461 gactgattac cggagctccc attgacaggc gtgaagttcg ccataggtcc cggattctct
     5521 tcgacatccc cacacgtcaa aaggctcccg cgtccttctg ctctgctagc cttatacaac
     5581 tcatccattc ccccggtcga atgtcgcccc tccgcccttt cgtactgttc cacgatcgta
     5641 taatcctcgt tatggcttgt aatgtcgagt ttgatattca cattgtaggc acccggcaac
     5701 tgcaccggct tcttcgcctt atacgttgtc ttgacttcgg cgtcgtagtg tccaccgtct
     5761 ttcaacttca atctctgttt gatctcccct ttgagtgctc catcttcggg atacatccgc
     5821 tccgacgacg cctcccaacc catcgtcttt ttctgcatga ctggaccgtc cgaggggaaa
     5881 ttagtgcctc tgagtttcac cttgtagata aactcgccgt cttggaggga cgagtcttgc
     5941 gtcactgtca ctacccctcc atcctcaaaa ttcatcactc gctcccattt gaagccctcg
     6001 ggaaacgaca gtttaagata gtctggaatg tccgcgggat gcttcacata cgctttggac
     6061 ccgtacataa actgcggtga aaggatgtcc cacgcgaaag gcaacggtcc tcctttggtg
     6121 actttcagct tggctgtctg tgtcccctcg taggggcgtc cctcaccctc tccctcgatc
     6181 tcaaactcgt gcccgttgac gctgccttcc atgtgcacct taaaccgcat gaactccttg
     6241 ataattgcca tgttatcttc ttcccccttt gataccatgg tggcggccgc atgcaggtgg
     6301 cttccggtgg atcccggtgc tcgactagta taacttcgta taggatactt tatacgaagt
     6361 tatcattggg attcttccta ttttgatcca agcatcacca tcgaccctct agtccagatc
     6421 tcaccatcga cccataactt cgtatagcat acattatacg aagttatgtc cctcgaagag
     6481 gttcgaattc gatatcaagc ttatcgataa tcaacctctg gattacaaaa tttgtgaaag
     6541 attgactggt attcttaact atgttgctcc ttttacgcta tgtggatacg ctgctttaat
     6601 gcctttgtat catgctattg cttcccgtat ggctttcatt ttctcctcct tgtataaatc
     6661 ctggttgctg tctctttatg aggagttgtg gcccgttgtc aggcaacgtg gcgtggtgtg
     6721 cactgtgttt gctgacgcaa cccccactgg ttggggcatt gccaccacct gtcagctcct
     6781 ttccgggact ttcgctttcc ccctccctat tgccacggcg gaactcatcg ccgcctgcct
     6841 tgcccgctgc tggacagggg ctcggctgtt gggcactgac aattccgtgg tgttgtcggg
     6901 gaaatcatcg tcctttcctt ggctgctcgc ctgtgttgcc acctggattc tgcgcgggac
     6961 gtccttctgc tacgtccctt cggccctcaa tccagcggac cttccttccc gcggcctgct
     7021 gccggctctg cggcctcttc cgcgtcttcg ccttcgccct cagacgagtc ggatctccct
     7081 ttgggccgcc tccccgcatc gataccgtcg acccgggcgg ccgcttcgag cagacatgat
     7141 aagatacatt gatgagtttg gacaaaccac aactagaatg cagtgaaaaa aatgctttat
     7201 ttgtgaaatt tgtgatgcta ttgctttatt tgtaaccatt ataagctgca ataaacaagt
     7261 taacaacaac aattgcattc attttatgtt tcaggttcag ggggagatgt gggaggtttt
     7321 ttaaagcaag taaaacctct acaaatgtgg taaaatcgat aaggatcttc ctagagcatg
     7381 gctacgtaga taagtagcat ggcgggttaa tcattaacta caaggaaccc ctagtgatgg
     7441 agttggccac tccctctctg cgcgctcgct cgctcactga ggccgggcga ccaaaggtcg
     7501 cccgacgccc gggctttgcc cgggcggcct cagtgagcga gcgagcgcgc agctgcatta
     7561 atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt gggcgct
//
