LOCUS       sequence-475755-        5947 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Express LgBiT-NCK1 Fusion Protein in Mammalian Cells under a CMV 
            promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5947)
  AUTHORS   Promega
  TITLE     Promega Plasmid Collection
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 5947)
  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: 
            "Unpublished"
COMMENT     Sequence Label: LgBiT-NCK1 Fusion Vector
FEATURES             Location/Qualifiers
     source          1..5947
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     polyA_signal    17..98
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     42..61
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     promoter        complement(2001..2019)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(2002..2019)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     enhancer        2139..2442
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     primer_bind     2597..2617
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     promoter        2650..2697
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             2729..3523
                     /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(2783..2802)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3393..3412
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     rep_origin      complement(3681..4269)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3761..3780)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     polyA_signal    4395..4443
                     /note="synthetic polyadenylation signal"
     CDS             complement(4467..5504)
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     EVGRV"
     RBS             4480..4488
                     /label=Shine-Dalgarno sequence
                     /note="full consensus sequence for ribosome-binding sites 
                     upstream of start codons in E. coli; complementary to a 
                     region in the 3' end of the 16S rRNA (Chen et al., 1994)"
     promoter        complement(5535..5892)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      5549..5684
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     enhancer        5701..5892
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     5849..5869
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
ORIGIN
        1 aagacgcgtg acgtcggcag tgaaaaaaat gctttatttg tgaaatttgt gatgctattg
       61 ctttatttgt aaccattata agctgcaata aacaagttaa caacaacaat tgcattcatt
      121 ttatgtttca ggttcagggg gaggtgtggg aggtttttta aagcaagtaa aacctctaca
      181 aatgtggtat ggctgattat gatccgcggc cgcaatacgt cgacgttatc agctgacttc
      241 gtacgagagc ctaggtgttt ctagaggatc cccgggtacc gagcccgaat tcgtttaaac
      301 tgataaatgc ttgacaagat ataatttttc tccttgttca cttgtaaaaa ttggtgcctt
      361 tttgtaatgt tctacaagtt cttccatggt gctgaattta cgctgcccaa tgcagtagac
      421 agtctctttt agttggactt taaaatgctt gtttttccct tgtgctttta gtgatactga
      481 gaaatcattt ggcgaagatt cactatcacg aatgaggaaa tccccttcat gtcctctttc
      541 atttaatgcc atttctgctt gatgcctggt gactttgcca taataccaag gattgccagc
      601 aaactttcca gtgagtgaag gcctaatgta atcacactgt ggaggtgatg gttccaaacc
      661 tgaagttaat ggattattct gcataacggt aacatagttt tttggtacta gaccaaccat
      721 accattgatc ttcctgcatt tccaccactc tgggtcattt tcaggttttt caataacatc
      781 cattacatct cctttctcga aattaagttc ttcatcatta gatgagctga atgggtaaag
      841 agcctgtacc acatgcaaca cttgcccagt atttaggtta ttgacgactg ctgctaattt
      901 ctctgacaga gaacccacat ggtcacccaa aggactgtca ccttcttcag ttacatagtt
      961 tgaagggaac catccaactt gtccattgta gctaccacgc caccacccat cactgcattt
     1021 ctccatgacg atcacctttg tcccctttat caatgataat tcatcctctc tctcagccat
     1081 gtagttaaat ttcacataag cgggcatgtt gaggtcatag agacgttccc ctgggtcaac
     1141 aaaactatca tcagcaggag atgcagaatc tggcacacta ggttttcttt tcacttttcc
     1201 aatgcctaag gtatccttta ggtttttcac aatagatgct ttccgagcac tgtttttcct
     1261 ttccacatag ttagaaggca caaaacctgt tttattcatg gaatttcgaa ctcgccacca
     1321 ggacttagaa tcatccagaa gccataatct ctcattcttc ttgatgtcca actcttgttc
     1381 ttgttgggcc acataatcaa atttggctac taccaccact tcttctgcca tggcgatcgc
     1441 accgctcgag cctccacctc cgctcccgcc accaccggag ctcccactgt tgatggttac
     1501 tcggaacagc atggagccgt cgggggtgat caggcgctcg tcgataattt tgttgccgtt
     1561 ccacagggtc cctgttacag tgatcttttt gccgtcgaac acggcgatgc cttcatacgg
     1621 ccgtccgaaa tagttcagca tgttcggcgt aaccccgtcg attaccagtg tgccataggg
     1681 caggatcacc ttaaagtgat gatcatccac agggtacacc accttaaaca cctcttcgat
     1741 ctgggccatt tggtcggcgc tcagaccttc atacgggatg atgacatgga tgtcgatctt
     1801 cagggcattt tcaccgctcc ggacaatcct ttggatcgga gttacggaca cggcgagatt
     1861 ctgcagcaaa ctggacacac ctccctgttc aaggacttgg tccaggttgt aggcggctgt
     1921 ctgttcccag tccccaacga aatcttcgag tgtgaagacc atggtgagcg ctagctgcaa
     1981 aaagaacaag tagcttgtat tctatagtgt cacctaaatg ggcgatctga cggttcacta
     2041 aaccagctct gcttatatag gcctccctgg cgcgccacta gttattaata gtaatcaatt
     2101 acggggtcat tagttcatag cccatatatg gagttccgcg ttacataact tacggtaaat
     2161 ggcccgcctg gctgaccgcc caacgacccc cgcccattga cgtcaataat gacgtatgtt
     2221 cccatagtaa cgccaatagg gactttccat tgacgtcaat gggtggagta tttacggtaa
     2281 actgcccact tggcagtaca tcaagtgtat catatgccaa gtccgccccc tattgacgtc
     2341 aatgacggta aatggcccgc ctggcattat gcccagtaca tgaccttacg ggactttcct
     2401 acttggcagt acatctacgt attagtcatc gctattacca tggtgatgcg gttttggcag
     2461 tacaccaatg ggcgtggata gcggtttgac tcacggggat ttccaagtct ccaccccatt
     2521 gacgtcaatg ggagtttgtt ttggcaccaa aatcaacggg actttccaaa atgtcgtaat
     2581 aaccccgccc cgttgacgca aatgggcggt aggcgtgtac ggtgggaggt caaatcgaat
     2641 tctaatactg ttgacaatta atcatcggca tagtatatcg gcatagtata atacgacaag
     2701 gtgaggaact aaacccagga ggcagatcat gattgaacaa gatggattgc acgcaggttc
     2761 tccggccgct tgggtggaga ggctattcgg ctatgactgg gcacaacaga caatcggctg
     2821 ctctgatgcc gccgtgttcc ggctgtcagc gcaggggcgc ccggttcttt ttgtcaagac
     2881 cgacctgtcc ggtgccctga atgaactgca ggacgaggca gcgcggctat cgtggctggc
     2941 cacgacgggc gttccttgcg cagctgtgct cgacgttgtc actgaagcgg gaagggactg
     3001 gctgctattg ggcgaagtgc cggggcagga tctcctgtca tctcaccttg ctcctgccga
     3061 gaaagtatcc atcatggctg atgcaatgcg gcggctgcat acgcttgatc cggctacctg
     3121 cccattcgac caccaagcga aacatcgcat cgagcgagca cgtactcgga tggaagccgg
     3181 tcttgtcgat caggatgatc tggacgaaga gcatcagggg ctcgcgccag ccgaactgtt
     3241 cgccaggctc aaggcgcgca tgcccgacgg cgaggatctc gtcgtgaccc atggcgatgc
     3301 ctgcttgccg aatatcatgg tggaaaatgg ccgcttttct ggattcatcg actgtggccg
     3361 gctgggtgtg gcggaccgct atcaggacat agcgttggct acccgtgata ttgctgaaga
     3421 gcttggcggc gaatgggctg accgcttcct cgtgctttac ggtatcgccg ctcccgattc
     3481 gcagcgcatc gccttctatc gccttcttga cgagttcttc tgattcgaaa tgaccgacca
     3541 agcgacgccc aaccggtatc agctcactca aaggcggtaa tacggttatc cacagaatca
     3601 ggggataacg caggaaagaa catgtgagca aaaggccagc aaaaggccag gaaccgtaaa
     3661 aaggccgcgt tgctggcgtt tttccatagg ctccgccccc ctgacgagca tcacaaaaat
     3721 cgacgctcaa gtcagaggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc
     3781 cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc
     3841 gcctttctcc cttcgggaag cgtggcgctt tctcatagct cacgctgtag gtatctcagt
     3901 tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac
     3961 cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgacttatcg
     4021 ccactggcag cagccactgg taacaggatt agcagagcga ggtatgtagg cggtgctaca
     4081 gagttcttga agtggtggcc taactacggc tacactagaa ggacagtatt tggtatctgc
     4141 gctctgctga agccagttac cttcggaaaa agagttggta gctcttgatc cggcaaacaa
     4201 accaccgctg gtagcggtgg tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa
     4261 ggatttcaag aagatccttt gatcttttct acggggtctg acgctcagtg gaacgaaaac
     4321 tcacgttaag ggattttggt catgagatta tcaaaaagga tcttcaccta gatcctttta
     4381 tagtccggaa tcgtcacaca aaaaaccaac acacagatgt aatgaaaata aagatatttt
     4441 attgcggcca tcgtgatggc tagagtctaa actcgaccta cctccttagc gcgcggtcgt
     4501 gtactgggcc ggcggttgcc gctgtcggcc agcacctcga cgcagccgtc ggtccatacg
     4561 gctgcgctcc ggcgagcgat ttgtgtgcga ccgacggtgc cggccccgct gcggacgatg
     4621 gcatcgcagc ggccttgagc ccaggcagca tcgtcgaagt tgccgtccac gaggctctgg
     4681 tagagctgat ccaggccgat gcgcagcatg taggctcgca gacgagggct gccggccagc
     4741 tcgggatgcc ggcgctcgaa gtagcgagtc tgctgctcca tgcaagccag ccagggccgc
     4801 cagaagaaga tgttggccac ctcgtactga ctgtccccga acatagcttc ggaccagtcg
     4861 attacggcgg tgatgcggcc gttgtcggtc aggacgttgt tgctgccgaa gtcggcatgg
     4921 acgaggtggc gcacctcggg acagtcttcg gcccacagca tcagttcgtc cagggcttga
     4981 gctacgctgg cggacacggt gtcgtccatc acggtctgcc agtggtagac atggggatca
     5041 gcaatggcgc aaatgaaatc ccgccaggtt gtgtactggc cgatgccttg gggcccgaat
     5101 gggccgaagc cgctggtttg actgaggtcg gcggcggcaa tagcatccat agcttcggcg
     5161 acaggctgta acacagcagg cagctctgtt tcggggaggt cttggagagt aacgccttgg
     5221 gcgcgtctac tgatgcagta tgtcaggctc tcgctgaact cgccgatgtc caacacttcg
     5281 gggatgggta gtgcagcgct ggcgaagtgg cggtacacgt agcggtcttt gtagaagcca
     5341 tcagcgcagc tattcacccg cagtacatag ccgcgtccgc cgacatcgaa gctgaaggct
     5401 cggctctctt cgccctccga caactgcatc aggtcgctca cactgtcgaa cttctcgatg
     5461 agaaattttt caacgctggt agcggtgagt tcgggcttct tcatggtggc gctagtgtca
     5521 gaagaatcga gctttttgca aaagcctagg cctccaaaaa agcctcctca ctacttctgg
     5581 aatagctcag aggccgaggc ggcctcggcc tctgcataaa taaaaaaaat tagtcagcca
     5641 tggggcggag aatgggcgga actgggcgga gttaggggcg ggatgggcgg agttaggggc
     5701 gggactatgg ttgctgacta attgagatgc atgctttgca tacttctgcc tgctggggag
     5761 cctggggact ttccacacct ggttgctgac taattgagat gcatgctttg catacttctg
     5821 cctgctgggg agcctgggga ctttccacac cctaactgac acacattcca cagctggttc
     5881 tttccgcctc agaaggtacc taaccaagtt cctctttcag aggttatttc aggccatggt
     5941 gctgcgc
//
