LOCUS       sequence-316153-        3770 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Gateway entry clone containing HyPep presynaptic component (Hydra 
            RFamide) for HySyn.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3770)
  AUTHORS   Hawk JD, Wisdom EM, Sengupta T, Kashlan ZD, Colon-Ramos DA
  TITLE     A genetically encoded tool for reconstituting synthetic modulatory 
            neurotransmission and reconnect neural circuits in vivo.
  JOURNAL   Nat Commun. 2021 Aug 9;12(1):4795. doi: 10.1038/s41467-021-24690-9.
  PUBMED    34373460
REFERENCE   2  (bases 1 to 3770)
  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"; journalName: "Nat 
            Commun."; date: "2021-08-9"; volume: "12(1):4795. doi"; pages: " 
            10.1038/s41467-021-24690-9"
COMMENT     Sequence Label: pJH120_pE12_HyPep_emGFP
FEATURES             Location/Qualifiers
     source          1..3770
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     1..17
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    33..132
                     /gene="mutant version of attL"
                     /label=attL1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     CDS             join(612..614,615..1328)
                     /codon_start=1
                     /product="Emerald GFP"
                     /label=EmGFP
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHKVYITADKQKNGIKV
                     NFKTRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     complement(654..675)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(915..934)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1262..1283
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     protein_bind    complement(1359..1458)
                     /gene="mutant version of attL"
                     /label=attL2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(1475..1494)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1476..1494)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1499..1515)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1499..1515)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1617..1636)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             1628..2437
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYGKP
                     DAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGKTA
                     FQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDASD
                     FDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGIAD
                     RYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     complement(1695..1714)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      2584..3172
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3073..3092
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3326..3343
                     /label=L4440
                     /note="L4440 vector, forward primer"
     terminator      complement(3502..3529)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     terminator      complement(3621..3707)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     join(3756..3770,1..8)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     join(3770,1..17)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
ORIGIN
        1 gtaaaacgac ggccagtctt aagctcgggc cccaaataat gattttattt tgactgatag
       61 tgacctgttc gttgcaacaa attgatgagc aatgcttttt tataatgcca actttgtaca
      121 aaaaagcagg cttagccacc atgctgggaa acaaaagaat gggactgtgt ggactgaccc
      181 tggccctgtc tctgctggtg tgtctgggaa ttttggctga gggatatcca tctccagata
      241 atccaggaga ggacgcccca gctgaagatg agacaggagc tgaagatctg ggagaggagc
      301 acgcttggaa ggctagccat agacagaaaa gacagtggct gggaggaaga ttcggaaaaa
      361 gatctggaga agaggaggat catgatatga tgaagagaca gtggctcgga ggaagatttg
      421 gaaagagaag cggagatgag gaggccgaga ctagtcacag acagaagaga cagtggttta
      481 acggaagatt cggcaaaaga tctggcgagg aggaagatca cgatatgatg aaaagacagt
      541 ggttcaacgg aagatttggc aaaagatccg gagatgagga agcctctaga ggatcccaca
      601 gacaaaaaag agtgagcaag ggcgaggagc tgttcaccgg ggtggtgccc atcctggtcg
      661 agctggacgg cgacgtaaac ggccacaagt tcagcgtgtc cggcgagggc gagggcgatg
      721 ccacctacgg caagctgacc ctgaagttca tctgcaccac cggcaagctg cccgtgccct
      781 ggcccaccct cgtgaccacc ttgacctacg gcgtgcagtg cttcgcccgc taccccgacc
      841 acatgaaaca gcacgacttc ttcaagtccg ccatgcccga aggctacgtc caggagcgca
      901 ccatcttctt caaggacgac ggcaactaca agacccgcgc cgaggtgaag ttcgagggcg
      961 acaccctggt gaaccgcatc gagctgaagg gcatcgactt caaggaggac ggcaacatcc
     1021 tggggcacaa gctggagtac aactacaaca gccacaaggt ctatatcacc gccgacaagc
     1081 agaagaacgg catcaaggtg aacttcaaga cccgccacaa catcgaggac ggcagcgtgc
     1141 agctcgccga ccactaccag cagaacaccc ccatcggcga cggccccgtg ctgctgcccg
     1201 acaaccacta cctgagcacc cagtccgccc tgagcaaaga ccccaacgag aagcgcgatc
     1261 acatggtcct gctggagttc gtgaccgccg ccgggatcac tctcggcatg gacgagctgt
     1321 acaagtaaaa gagatctgga gatgaagaat gatcatgaac ccagctttct tgtacaaagt
     1381 tggcattata agaaagcatt gcttatcaat ttgttgcaac gaacaggtca ctatcagtca
     1441 aaataaaatc attatttgcc atccagctga tatcccctat agtgagtcgt attacatggt
     1501 catagctgtt tcctggcagc tctggcccgt gtctcaaaat ctctgatgtt acattgcaca
     1561 agataaaaat atatcatcat gaacaataaa actgtctgct tacataaaca gtaatacaag
     1621 gggtgttatg agccatattc aacgggaaac gtcgaggccg cgattaaatt ccaacatgga
     1681 tgctgattta tatgggtata aatgggctcg cgataatgtc gggcaatcag gtgcgacaat
     1741 ctatcgcttg tatgggaagc ccgatgcgcc agagttgttt ctgaaacatg gcaaaggtag
     1801 cgttgccaat gatgttacag atgagatggt cagactaaac tggctgacgg aatttatgcc
     1861 tcttccgacc atcaagcatt ttatccgtac tcctgatgat gcatggttac tcaccactgc
     1921 gatccccgga aaaacagcat tccaggtatt agaagaatat cctgattcag gtgaaaatat
     1981 tgttgatgcg ctggcagtgt tcctgcgccg gttgcattcg attcctgttt gtaattgtcc
     2041 ttttaacagc gatcgcgtat ttcgtctcgc tcaggcgcaa tcacgaatga ataacggttt
     2101 ggttgatgcg agtgattttg atgacgagcg taatggctgg cctgttgaac aagtctggaa
     2161 agaaatgcat aaacttttgc cattctcacc ggattcagtc gtcactcatg gtgatttctc
     2221 acttgataac cttatttttg acgaggggaa attaataggt tgtattgatg ttggacgagt
     2281 cggaatcgca gaccgatacc aggatcttgc catcctatgg aactgcctcg gtgagttttc
     2341 tccttcatta cagaaacggc tttttcaaaa atatggtatt gataatcctg atatgaataa
     2401 attgcagttt catttgatgc tcgatgagtt tttctaatca gaattggtta attggttgta
     2461 acactggcag agcattacgc tgacttgacg ggacggcgca agctcatgac caaaatccct
     2521 taacgtgagt tacgcgtcgt tccactgagc gtcagacccc gtagaaaaga tcaaaggatc
     2581 ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct
     2641 accagcggtg gtttgtttgc cggatcaaga gctaccaact ctttttccga aggtaactgg
     2701 cttcagcaga gcgcagatac caaatactgt tcttctagtg tagccgtagt taggccacca
     2761 cttcaagaac tctgtagcac cgcctacata cctcgctctg ctaatcctgt taccagtggc
     2821 tgctgccagt ggcgataagt cgtgtcttac cgggttggac tcaagacgat agttaccgga
     2881 taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca cagcccagct tggagcgaac
     2941 gacctacacc gaactgagat acctacagcg tgagctatga gaaagcgcca cgcttcccga
     3001 agggagaaag gcggacaggt atccggtaag cggcagggtc ggaacaggag agcgcacgag
     3061 ggagcttcca gggggaaacg cctggtatct ttatagtcct gtcgggtttc gccacctctg
     3121 acttgagcgt cgatttttgt gatgctcgtc aggggggcgg agcctatgga aaaacgccag
     3181 caacgcggcc tttttacggt tcctggcctt ttgctggcct tttgctcaca tgttctttcc
     3241 tgcgttatcc cctgattctg tggataaccg tattaccgcc tttgagtgag ctgataccgc
     3301 tcgccgcagc cgaacgaccg agcgcagcga gtcagtgagc gaggaagcgg aagagcgccc
     3361 aatacgcaaa ccgcctctcc ccgcgcgttg gccgattcat taatgcagct ggcacgacag
     3421 gtttcccgac tggaaagcgg gcagtgagcg caacgcaatt aatacgcgta ccgctagcca
     3481 ggaagagttt gtagaaacgc aaaaaggcca tccgtcagga tggccttctg cttagtttga
     3541 tgcctggcag tttatggcgg gcgtcctgcc cgccaccctc cgggccgttg cttcacaacg
     3601 ttcaaatccg ctcccggcgg atttgtccta ctcaggagag cgttcaccga caaacaacag
     3661 ataaaacgaa aggcccagtc ttccgactga gcctttcgtt ttatttgatg cctggcagtt
     3721 ccctactctc gcgttaacgc tagcatggat gttttcccag tcacgacgtt
//
