LOCUS       sequence-478917-        6836 bp    DNA     circular SYN 17-JUL-2025
DEFINITION  Expresses TeNT in the mammalian cytosol.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6836)
  AUTHORS   Roh H, Kim D, Kim B, Jeon Y, Kim Y, Jacko M, Xu F, Lin C, Won Um J, 
            Ting AY
  TITLE     Light-activated tetanus neurotoxin for conditional proteolysis and 
            inducible synaptic inhibition in vivo
  JOURNAL   bioRxiv 2025.01.27.635161; doi: 10.1101/2025.01.27.635161
REFERENCE   2  (bases 1 to 6836)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Jul 17, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "bioRxiv 
            2025.01.27.635161; doi: 10.1101/2025.01.27.635161"
COMMENT     Sequence Label: pcDNA3-TeNT
FEATURES             Location/Qualifiers
     source          1..6836
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     863..882
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      901..910
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             910..951
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     promoter        complement(2389..2407)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(2390..2407)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(2427..2444)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2433..2657
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2703..3131
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2790..2809)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3000..3021
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3140..3160)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3145..3474
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3325..3460
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3387..3406
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3541..4335
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(3595..3614)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4205..4224
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4509..4630
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4546..4565)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4600..4619
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4679..4695)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4679..4695)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4692..4714)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4703..4719
                     /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)."
     promoter        complement(join(4727..4733,4734..4751,4752..4757))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4772..4793
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4910..4927)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5081..5669)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5161..5180)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5840..6631,6632..6700))
                     /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     6463..6482
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6701..6805)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gcttgcggcc
      901 gccaccatgg gcaagcccat ccccaacccc ctgctgggcc tggacagcac caagggctcg
      961 ggctcgacct cgggctcggg caccggtccg atcaccatca acaacttccg ttactccgac
     1021 ccggttaaca acgacaccat catcatgatg gaaccgccgt actgcaaagg tctagacatc
     1081 tactacaaag ccttcaaaat caccgaccgt atctggatcg ttccggaacg ttacgaattt
     1141 ggtaccaaac cggaagactt caacccgccg tcctccctga tcgaaggtgc ttccgaatac
     1201 tacgacccga actacctgcg taccgactcc gacaaagacc gtttcctgca gaccatggtt
     1261 aaactgttca accgtatcaa aaacaacgtt gctggtgaag ctctgctgga caaaatcatc
     1321 aacgctatcc cgtacctggg taactcctac tccctgctgg acaaattcga caccaactcc
     1381 aactccgttt ccttcaacct gctggaacag gatccgtccg gtgctaccac caaatccgct
     1441 atgctgacca acctgatcat cttcggtccg ggtccggttc tgaacaaaaa cgaagttcgt
     1501 ggtatcgttc tgcgtgttga caacaaaaac tacttcccgt gccgtgacgg tttcggttcc
     1561 atcatgcaga tggctttctg cccggaatac gtaccgacct tcgacaacgt tatcgaaaac
     1621 atcacctccc tgaccatcgg taaatccaaa tacttccagg acccggctct gctgctgatg
     1681 cacgaactga tccacgttct gcacggtctg tacggcatgc aggtttcctc ccacgaaatc
     1741 atcccgtcca aacaggaaat ctacatgcag cacacctacc cgatctccgc tgaagagctc
     1801 ttcaccttcg gtggtcagga cgctaacctg atctccatcg atatcaaaaa cgacctgtac
     1861 gaaaaaaccc tgaacgacta caaagctatc gctaacaaac tgtcccaggt tacctcctgc
     1921 aacgacccga acatcgacat cgactcctac aaacagatct atcagcagaa gtatcagttt
     1981 gataaggatt ccaacggcca gtatatagtg aacgaggata agtttcagat actgtataac
     2041 tccataatgt atggctttac tgagatagag ctgggcaaga agtttaacat aaagactcgc
     2101 ttaagctatt tttccatgaa ccacgatcct gtgaagatac ctaacctgct ggatgatact
     2161 atatataacg atactgaggg ctttaacata gagtccaagg atctgaagtc cgagtataag
     2221 ggccagaaca tgagggtgaa cactaacgcc tttaggaacg tcgacggttc cggtctggtt
     2281 tccaaactga tcggtctgtg caaaaaaatc atcccgccga ccaacatccg tgaaaacctg
     2341 tacaaccgta ccgcttaata actcgagcat gcatctagag ggccctattc tatagtgtca
     2401 cctaaatgct agagctcgct gatcagcctc gactgtgcct tctagttgcc agccatctgt
     2461 tgtttgcccc tcccccgtgc cttccttgac cctggaaggt gccactccca ctgtcctttc
     2521 ctaataaaat gaggaaattg catcgcattg tctgagtagg tgtcattcta ttctgggggg
     2581 tggggtgggg caggacagca agggggagga ttgggaagac aatagcaggc atgctgggga
     2641 tgcggtgggc tctatggctt ctgaggcgga aagaaccagc tggggctcta gggggtatcc
     2701 ccacgcgccc tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc gcagcgtgac
     2761 cgctacactt gccagcgccc tagcgcccgc tcctttcgct ttcttccctt cctttctcgc
     2821 cacgttcgcc ggctttcccc gtcaagctct aaatcggggg ctccctttag ggttccgatt
     2881 tagtgcttta cggcacctcg accccaaaaa acttgattag ggtgatggtt cacgtagtgg
     2941 gccatcgccc tgatagacgg tttttcgccc tttgacgttg gagtccacgt tctttaatag
     3001 tggactcttg ttccaaactg gaacaacact caaccctatc tcggtctatt cttttgattt
     3061 ataagggatt ttgccgattt cggcctattg gttaaaaaat gagctgattt aacaaaaatt
     3121 taacgcgaat taattctgtg gaatgtgtgt cagttagggt gtggaaagtc cccaggctcc
     3181 ccagcaggca gaagtatgca aagcatgcat ctcaattagt cagcaaccag gtgtggaaag
     3241 tccccaggct ccccagcagg cagaagtatg caaagcatgc atctcaatta gtcagcaacc
     3301 atagtcccgc ccctaactcc gcccatcccg cccctaactc cgcccagttc cgcccattct
     3361 ccgccccatg gctgactaat tttttttatt tatgcagagg ccgaggccgc ctctgcctct
     3421 gagctattcc agaagtagtg aggaggcttt tttggaggcc taggcttttg caaaaagctc
     3481 ccgggagctt gtatatccat tttcggatct gatcaagaga caggatgagg atcgtttcgc
     3541 atgattgaac aagatggatt gcacgcaggt tctccggccg cttgggtgga gaggctattc
     3601 ggctatgact gggcacaaca gacaatcggc tgctctgatg ccgccgtgtt ccggctgtca
     3661 gcgcaggggc gcccggttct ttttgtcaag accgacctgt ccggtgccct gaatgaactg
     3721 caggacgagg cagcgcggct atcgtggctg gccacgacgg gcgttccttg cgcagctgtg
     3781 ctcgacgttg tcactgaagc gggaagggac tggctgctat tgggcgaagt gccggggcag
     3841 gatctcctgt catctcacct tgctcctgcc gagaaagtat ccatcatggc tgatgcaatg
     3901 cggcggctgc atacgcttga tccggctacc tgcccattcg accaccaagc gaaacatcgc
     3961 atcgagcgag cacgtactcg gatggaagcc ggtcttgtcg atcaggatga tctggacgaa
     4021 gagcatcagg ggctcgcgcc agccgaactg ttcgccaggc tcaaggcgcg catgcccgac
     4081 ggcgaggatc tcgtcgtgac ccatggcgat gcctgcttgc cgaatatcat ggtggaaaat
     4141 ggccgctttt ctggattcat cgactgtggc cggctgggtg tggcggaccg ctatcaggac
     4201 atagcgttgg ctacccgtga tattgctgaa gagcttggcg gcgaatgggc tgaccgcttc
     4261 ctcgtgcttt acggtatcgc cgctcccgat tcgcagcgca tcgccttcta tcgccttctt
     4321 gacgagttct tctgagcggg actctggggt tcgaaatgac cgaccaagcg acgcccaacc
     4381 tgccatcacg agatttcgat tccaccgccg ccttctatga aaggttgggc ttcggaatcg
     4441 ttttccggga cgccggctgg atgatcctcc agcgcgggga tctcatgctg gagttcttcg
     4501 cccaccccaa cttgtttatt gcagcttata atggttacaa ataaagcaat agcatcacaa
     4561 atttcacaaa taaagcattt ttttcactgc attctagttg tggtttgtcc aaactcatca
     4621 atgtatctta tcatgtctgt ataccgtcga cctctagcta gagcttggcg taatcatggt
     4681 catagctgtt tcctgtgtga aattgttatc cgctcacaat tccacacaac atacgagccg
     4741 gaagcataaa gtgtaaagcc tggggtgcct aatgagtgag ctaactcaca ttaattgcgt
     4801 tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat taatgaatcg
     4861 gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc tcgctcactg
     4921 actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc agctcactca aaggcggtaa
     4981 tacggttatc cacagaatca ggggataacg caggaaagaa catgtgagca aaaggccagc
     5041 aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg ctccgccccc
     5101 ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg acaggactat
     5161 aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc
     5221 cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt tctcatagct
     5281 cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc tgtgtgcaca
     5341 aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc
     5401 cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt agcagagcga
     5461 ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc tacactagaa
     5521 gaacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa agagttggta
     5581 gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt tgcaagcagc
     5641 agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct acggggtctg
     5701 acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgagatta tcaaaaagga
     5761 tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa agtatatatg
     5821 agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc tcagcgatct
     5881 gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact acgatacggg
     5941 agggcttacc atctggcccc agtgctgcaa tgataccgcg agacccacgc tcaccggctc
     6001 cagatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt ggtcctgcaa
     6061 ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta agtagttcgc
     6121 cagttaatag tttgcgcaac gttgttgcca ttgctacagg catcgtggtg tcacgctcgt
     6181 cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt acatgatccc
     6241 ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc agaagtaagt
     6301 tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt actgtcatgc
     6361 catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc tgagaatagt
     6421 gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg ggataatacc gcgccacata
     6481 gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa ctctcaagga
     6541 tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac tgatcttcag
     6601 catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa
     6661 aaaagggaat aagggcgaca cggaaatgtt gaatactcat actcttcctt tttcaatatt
     6721 attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa tgtatttaga
     6781 aaaataaaca aataggggtt ccgcgcacat ttccccgaaa agtgccacct gacgtc
//
