LOCUS       sequence-207249-        4884 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses Fn crRNA and mCherry in mammalian cells..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4884)
  AUTHORS   Toth E, Czene BC, Kulcsar PI, Krausz SL, Talas A, Nyeste A, Varga E,
            Huszar K, Weinhardt N, Ligeti Z, Borsy AE, Fodor E, Welker E
  TITLE     Mb- and FnCpf1 nucleases are active in mammalian cells: activities 
            and PAM preferences of four wild-type Cpf1 nucleases and of their 
            altered PAM specificity variants.
  JOURNAL   Nucleic Acids Res. 2018 Sep 20. pii: 5103950. doi: 
            10.1093/nar/gky815.
  PUBMED    30239882
REFERENCE   2  (bases 1 to 4884)
  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: "Nucleic 
            Acids Res. 2018 Sep 20. pii: 5103950. doi: 10.1093/nar/gky815."
COMMENT     Sequence Label: pTE4938
FEATURES             Location/Qualifiers
     source          1..4884
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        7..255
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     7..27
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     178..197
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     enhancer        308..593
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer; 
                     contains an 18-bp deletion relative to the standard CMV 
                     enhancer"
     promoter        595..872
                     /label=chicken beta-actin promoter
     intron          873..1100
                     /label=hybrid intron
                     /note="hybrid between chicken beta-actin (CBA) and minute 
                     virus of mice (MMV) introns (Gray et al., 2011)"
     regulatory      1112..1121
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1118..1828
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(1263..1281)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     polyA_signal    1957..2078
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1994..2013)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2048..2067
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     2208..2227
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2341..2445
                     /gene="bla"
                     /label=AmpR promoter
     promoter        2447..2804
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2447..2638
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2470..2490)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      2655..2790
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             2839..3633
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             2839..3633
                     /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
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(2893..2912)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3503..3522
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(3821..3840)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    3865..3912
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4241..4829
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4730..4749
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 acatgtgagg gcctatttcc catgattcct tcatatttgc atatacgata caaggctgtt
       61 agagagataa ttggaattaa tttgactgta aacacaaaga tattagtaca aaatacgtga
      121 cgtagaaagt aataatttct tgggtagttt gcagttttaa aattatgttt taaaatggac
      181 tatcatatgc ttaccgtaac ttgaaagtat ttcgatttct tggctttata tatcttgtgg
      241 aaaggacgaa acaccgtaat ttctactgtt gtagatggag acgttcgaac gtctcctttt
      301 ttgtacccgt tacataactt acggtaaatg gcccgcctgg ctgaccgccc aacgaccccc
      361 gcccattgac gtcaatagta acgccaatag ggactttcca ttgacgtcaa tgggtggagt
      421 atttacggta aactgcccac ttggcagtac atcaagtgta tcatatgcca agtacgcccc
      481 ctattgacgt caatgacggt aaatggcccg cctggcattg tgcccagtac atgaccttat
      541 gggactttcc tacttggcag tacatctacg tattagtcat cgctattacc atggtcgagg
      601 tgagccccac gttctgcttc actctcccca tctccccccc ctccccaccc ccaattttgt
      661 atttatttat tttttaatta ttttgtgcag cgatgggggc gggggggggg ggggggcgcg
      721 cgccaggcgg ggcggggcgg ggcgaggggc ggggcggggc gaggcggaga ggtgcggcgg
      781 cagccaatca gagcggcgcg ctccgaaagt ttccttttat ggcgaggcgg cggcggcggc
      841 ggccctataa aaagcgaagc gcgcggcggg cgggagtcgc tgcgcgctgc cttcgccccg
      901 tgccccgctc cgccgccgcc tcgcgccgcc cgccccggct ctgactgacc gcgttactcc
      961 cacaggtgag cgggcgggac ggcccttctc ctccgggctg taattagctg agcaagaggt
     1021 aagggtttaa gggatggttg gttggtgggg tattaatgtt taattacctg gagcacctgc
     1081 ctgaaatcac tttttttcag gttggaccgg tgccaccatg gtgagcaagg gcgaggagga
     1141 taacatggcc atcatcaagg agttcatgcg cttcaaggtg cacatggagg gctccgtgaa
     1201 cggccacgag ttcgagatcg agggcgaggg cgagggccgc ccctacgagg gcacccagac
     1261 cgccaagctg aaggtgacca agggtggccc cctgcccttc gcctgggaca tcctgtcccc
     1321 tcagttcatg tacggctcca aggcctacgt gaagcacccc gccgacatcc ccgactactt
     1381 gaagctgtcc ttccccgagg gcttcaagtg ggagcgcgtg atgaacttcg aggacggcgg
     1441 cgtggtgacc gtgacccagg actcctccct gcaggacggc gagttcatct acaaggtgaa
     1501 gctgcgcggc accaacttcc cctccgacgg ccccgtaatg cagaaaaaga ccatgggctg
     1561 ggaggcctcc tccgagcgga tgtaccccga ggacggcgcc ctgaagggcg agatcaagca
     1621 gaggctgaag ctgaaggacg gcggccacta cgacgctgag gtcaagacca cctacaaggc
     1681 caagaagccc gtgcagctgc ccggcgccta caacgtcaac atcaagttgg acatcacctc
     1741 ccacaacgag gactacacca tcgtggaaca gtacgaacgc gccgagggcc gccactccac
     1801 cggcggcatg gacgagctgt acaagtagga tccaccggat ctagataact gatcataatc
     1861 agccatacca catttgtaga ggttttactt gctttaaaaa acctcccaca cctccccctg
     1921 aacctgaaac ataaaatgaa tgcaattgtt gttgttaact tgtttattgc agcttataat
     1981 ggttacaaat aaagcaatag catcacaaat ttcacaaata aagcattttt ttcactgcat
     2041 tctagttgtg gtttgtccaa actcatcaat gtatcttaac gcggtgaacc atcaccctaa
     2101 tcaagttttt tggggtcgag gtgccgtaaa gcactaaatc ggaaccctaa agggagcccc
     2161 cgatttagag cttgacgggg aaagccggcg aacgtggcga gaaaggaagg gaagaaagcg
     2221 aaaggagcgg gcgctagggc gctggcaagt gtagcggtca cgctgcgcgt aaccaccaca
     2281 cccgccgcgc ttaatgcgcc gctacagggc gcgtcaggtg gcacttttcg gggaaatgtg
     2341 cgcggaaccc ctatttgttt atttttctaa atacattcaa atatgtatcc gctcatgaga
     2401 caataaccct gataaatgct tcaataatat tgaaaaagga agagtcctga ggcggaaaga
     2461 accagctgtg gaatgtgtgt cagttagggt gtggaaagtc cccaggctcc ccagcaggca
     2521 gaagtatgca aagcatgcat ctcaattagt cagcaaccag gtgtggaaag tccccaggct
     2581 ccccagcagg cagaagtatg caaagcatgc atctcaatta gtcagcaacc atagtcccgc
     2641 ccctaactcc gcccatcccg cccctaactc cgcccagttc cgcccattct ccgccccatg
     2701 gctgactaat tttttttatt tatgcagagg ccgaggccgc ctcggcctct gagctattcc
     2761 agaagtagtg aggaggcttt tttggaggcc taggcttttg caaagatcga tcaagagaca
     2821 ggatgaggat cgtttcgcat gattgaacaa gatggattgc acgcaggttc tccggccgct
     2881 tgggtggaga ggctattcgg ctatgactgg gcacaacaga caatcggctg ctctgatgcc
     2941 gccgtgttcc ggctgtcagc gcaggggcgc ccggttcttt ttgtcaagac cgacctgtcc
     3001 ggtgccctga atgaactgca agacgaggca gcgcggctat cgtggctggc cacgacgggc
     3061 gttccttgcg cagctgtgct cgacgttgtc actgaagcgg gaagggactg gctgctattg
     3121 ggcgaagtgc cggggcagga tctcctgtca tctcaccttg ctcctgccga gaaagtatcc
     3181 atcatggctg atgcaatgcg gcggctgcat acgcttgatc cggctacctg cccattcgac
     3241 caccaagcga aacatcgcat cgagcgagca cgtactcgga tggaagccgg tcttgtcgat
     3301 caggatgatc tggacgaaga gcatcagggg ctcgcgccag ccgaactgtt cgccaggctc
     3361 aaggcgagca tgcccgacgg cgaggatctc gtcgtgaccc atggcgatgc ctgcttgccg
     3421 aatatcatgg tggaaaatgg ccgcttttct ggattcatcg actgtggccg gctgggtgtg
     3481 gcggaccgct atcaggacat agcgttggct acccgtgata ttgctgaaga gcttggcggc
     3541 gaatgggctg accgcttcct cgtgctttac ggtatcgccg ctcccgattc gcagcgcatc
     3601 gccttctatc gccttcttga cgagttcttc tgagcgggac tctggggttc gaaatgaccg
     3661 accaagcgac gcccaacctg ccatcacgag atttcgattc caccgccgcc ttctatgaaa
     3721 ggttgggctt cggaatcgtt ttccgggacg ccggctggat gatcctccag cgcggggatc
     3781 tcatgctgga gttcttcgcc caccctaggg ggaggctaac tgaaacacgg aaggagacaa
     3841 taccggaagg aacccgcgct atgacggcaa taaaaagaca gaataaaacg cacggtgttg
     3901 ggtcgtttgt tcataaacgc ggggttcggt cccagggctg gcactctgtc gataccccac
     3961 cgagacccca ttggggccaa tacgcccgcg tttcttcctt ttccccaccc caccccccaa
     4021 gttcgggtga aggcccaggg ctcgcagcca acgtcggggc ggcaggccct gccatagcct
     4081 caggttactc atatatactt tagattgatt taaaacttca tttttaattt aaaaggatct
     4141 aggtgaagat cctttttgat aatctcatga ccaaaatccc ttaacgtgag ttttcgttcc
     4201 actgagcgtc agaccccgta gaaaagatca aaggatcttc ttgagatcct ttttttctgc
     4261 gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt tgtttgccgg
     4321 atcaagagct accaactctt tttccgaagg taactggctt cagcagagcg cagataccaa
     4381 atactgttct tctagtgtag ccgtagttag gccaccactt caagaactct gtagcaccgc
     4441 ctacatacct cgctctgcta atcctgttac cagtggctgc tgccagtggc gataagtcgt
     4501 gtcttaccgg gttggactca agacgatagt taccggataa ggcgcagcgg tcgggctgaa
     4561 cggggggttc gtgcacacag cccagcttgg agcgaacgac ctacaccgaa ctgagatacc
     4621 tacagcgtga gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc
     4681 cggtaagcgg cagggtcgga acaggagagc gcacgaggga gcttccaggg ggaaacgcct
     4741 ggtatcttta tagtcctgtc gggtttcgcc acctctgact tgagcgtcga tttttgtgat
     4801 gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa cgcggccttt ttacggttcc
     4861 tggccttttg ctggcctttt gctc
//
