LOCUS       sequence-356621-        6688 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses FLAG-tagged Acanthoeca spectabilis NF-kB3 protein in 
            mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6688)
  AUTHORS   Williams LM, Sridhar S, Samaroo J, Peart J, Adindu EK, Addanki A, 
            DiRusso CJ, Aguirre Carrion PJ, Rodriguez-Sastre N, Siggers T, 
            Gilmore TD
  TITLE     Comparison of NF-kappaB from the protists Capsaspora owczarzaki and 
            Acanthoeca spectabilis reveals extensive evolutionary 
            diversification of this transcription factor.
  JOURNAL   Commun Biol. 2021 Dec 16;4(1):1404. doi: 10.1038/s42003-021-02924-2.
  PUBMED    34916615
REFERENCE   2  (bases 1 to 6688)
  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: "Commun 
            Biol."; date: "2021-12-16"; volume: "4(1):1404. doi"; pages: " 
            10.1038/s42003-021-02924-2"
COMMENT     Sequence Label: pcDNA-FLAG-As-NF-kB3
FEATURES             Location/Qualifiers
     source          1..6688
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        70..449
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        450..653
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     604..624
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     698..717
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        698..716
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             739..762
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     promoter        complement(2076..2094)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(2077..2094)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(2114..2131)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2120..2344
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2390..2818
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2477..2496)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2687..2708
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2827..2847)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2832..3161
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3012..3147
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3074..3093
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3228..4022
                     /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(3282..3301)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3892..3911
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4196..4317
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4233..4252)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4287..4306
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4366..4382)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4366..4382)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4379..4401)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4390..4406
                     /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(4414..4420,4421..4438,4439..4444))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4459..4480
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4597..4614)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4768..5356)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4848..4867)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5527..6318,6319..6387))
                     /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     6150..6169
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6388..6492)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(6567..6586)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 gcatgaagaa tctgcttagg gttaggcgtt ttgcgctgct tcgcgatgta cgggccagat
       61 atacgcgttg acattgatta ttgactagtt attaatagta atcaattacg gggtcattag
      121 ttcatagccc atatatggag ttccgcgtta cataacttac ggtaaatggc ccgcctggct
      181 gaccgcccaa cgacccccgc ccattgacgt caataatgac gtatgttccc atagtaacgc
      241 caatagggac tttccattga cgtcaatggg tggagtattt acggtaaact gcccacttgg
      301 cagtacatca agtgtatcat atgccaagta cgccccctat tgacgtcaat gacggtaaat
      361 ggcccgcctg gcattatgcc cagtacatga ccttatggga ctttcctact tggcagtaca
      421 tctacgtatt agtcatcgct attaccatgg tgatgcggtt ttggcagtac atcaatgggc
      481 gtggatagcg gtttgactca cggggatttc caagtctcca ccccattgac gtcaatggga
      541 gtttgttttg gcaccaaaat caacgggact ttccaaaatg tcgtaacaac tccgccccat
      601 tgacgcaaat gggcggtagg cgtgtacggt gggaggtcta tataagcaga gctctctggc
      661 taactagaga acccactgct tactggctta tcgaaattaa tacgactcac tatagggaga
      721 cccaagcttg gtaccatgga ctacaaggac gacgatgaca agttaacacg tgctgcagat
      781 gcgaattctc caaatgattt cgaggctgtt ctgacgtctg ggggtgggtc tcatgcttgc
      841 ggaccatctg ccgccaagag agctagggtg gacttgagtg atgcctccga aaacgcctat
      901 aaccaggcat ggagaccctc cgcatctgtg gcttgtggag ctacctttca gccaccaagg
      961 acacccatga ctgcatggga cctgggcatt cttcctccta caactcatac aggccacgga
     1021 ggattcccac cgcgagcata ccccattgag tctttcccta tgcatcgagg tttccttcag
     1081 gttagtggag ctgcagcagc cccgccaata ccgagattga tggggggcca ccatcagtac
     1141 caccctacac acgccatgta tcgcgatctg cgggaagtct tggaaattac cgagcagccg
     1201 actgaaagag ggcggttcag atacgcaaag gaaaaaagac gaaccccact cccggggaga
     1261 agggatggag gatttcccac ggttagggtt gctgacagat atcgagacgt cttgccagat
     1321 ggcactcaca tacaggcatc cgtggtgacg agacaggacg gagaagacgg gatcccacgc
     1381 cctcactggc acagactcga gggaagggaa ggagagagcg ttagccagcc tctcagcctg
     1441 ggagccgcta cattctccaa cctcgtggtg atccggagcg acagaaatgc cgcagaaaat
     1501 cactggggcc ccaggcctac cgaagaccaa caggtgatcc gaattatgtt ttccgtgctt
     1561 ttccggttgc ctaccggtga aatggcccgg tcttgggtgg cttccgaacc tatctacggg
     1621 tgcgacctga aaattcagaa catgtctcac acagaagtcc cgatgctctc cggtgctgag
     1681 gtgaccctgc tgactagtaa gatcaggaaa cagtccatcg ccctgatgtt ggttgatgaa
     1741 tttacagatc atgacttcgc tcccggaaca aagattgacc cagcaaatgg gtgggagtgt
     1801 gatcaggacg ggaaaatctg ttgcactatt cagcccagct acgtgcacca tcagtacgct
     1861 ctcgtggcgc agataccaca gtattgggac ctgaccttgg agtctgacag acaaattagc
     1921 gccaggatta tcgataagga tcagaagatg gagtccaact gttctcaact cacatatgtc
     1981 cccggatctg aggccagaag gcgcagacca ccacccgaat gagaattcag ggatccctcg
     2041 agcggccgct cgagcatgca tctagagggc cctattctat agtgtcacct aaatgctaga
     2101 gctcgctgat cagcctcgac tgtgccttct agttgccagc catctgttgt ttgcccctcc
     2161 cccgtgcctt ccttgaccct ggaaggtgcc actcccactg tcctttccta ataaaatgag
     2221 gaaattgcat cgcattgtct gagtaggtgt cattctattc tggggggtgg ggtggggcag
     2281 gacagcaagg gggaggattg ggaagacaat agcaggcatg ctggggatgc ggtgggctct
     2341 atggcttctg aggcggaaag aaccagctgg ggctctaggg ggtatcccca cgcgccctgt
     2401 agcggcgcat taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc tacacttgcc
     2461 agcgccctag cgcccgctcc tttcgctttc ttcccttcct ttctcgccac gttcgccggc
     2521 tttccccgtc aagctctaaa tcgggggctc cctttagggt tccgatttag tgctttacgg
     2581 cacctcgacc ccaaaaaact tgattagggt gatggttcac gtagtgggcc atcgccctga
     2641 tagacggttt ttcgcccttt gacgttggag tccacgttct ttaatagtgg actcttgttc
     2701 caaactggaa caacactcaa ccctatctcg gtctattctt ttgatttata agggattttg
     2761 ccgatttcgg cctattggtt aaaaaatgag ctgatttaac aaaaatttaa cgcgaattaa
     2821 ttctgtggaa tgtgtgtcag ttagggtgtg gaaagtcccc aggctcccca gcaggcagaa
     2881 gtatgcaaag catgcatctc aattagtcag caaccaggtg tggaaagtcc ccaggctccc
     2941 cagcaggcag aagtatgcaa agcatgcatc tcaattagtc agcaaccata gtcccgcccc
     3001 taactccgcc catcccgccc ctaactccgc ccagttccgc ccattctccg ccccatggct
     3061 gactaatttt ttttatttat gcagaggccg aggccgcctc tgcctctgag ctattccaga
     3121 agtagtgagg aggctttttt ggaggcctag gcttttgcaa aaagctcccg ggagcttgta
     3181 tatccatttt cggatctgat caagagacag gatgaggatc gtttcgcatg attgaacaag
     3241 atggattgca cgcaggttct ccggccgctt gggtggagag gctattcggc tatgactggg
     3301 cacaacagac aatcggctgc tctgatgccg ccgtgttccg gctgtcagcg caggggcgcc
     3361 cggttctttt tgtcaagacc gacctgtccg gtgccctgaa tgaactgcag gacgaggcag
     3421 cgcggctatc gtggctggcc acgacgggcg ttccttgcgc agctgtgctc gacgttgtca
     3481 ctgaagcggg aagggactgg ctgctattgg gcgaagtgcc ggggcaggat ctcctgtcat
     3541 ctcaccttgc tcctgccgag aaagtatcca tcatggctga tgcaatgcgg cggctgcata
     3601 cgcttgatcc ggctacctgc ccattcgacc accaagcgaa acatcgcatc gagcgagcac
     3661 gtactcggat ggaagccggt cttgtcgatc aggatgatct ggacgaagag catcaggggc
     3721 tcgcgccagc cgaactgttc gccaggctca aggcgcgcat gcccgacggc gaggatctcg
     3781 tcgtgaccca tggcgatgcc tgcttgccga atatcatggt ggaaaatggc cgcttttctg
     3841 gattcatcga ctgtggccgg ctgggtgtgg cggaccgcta tcaggacata gcgttggcta
     3901 cccgtgatat tgctgaagag cttggcggcg aatgggctga ccgcttcctc gtgctttacg
     3961 gtatcgccgc tcccgattcg cagcgcatcg ccttctatcg ccttcttgac gagttcttct
     4021 gagcgggact ctggggttcg aaatgaccga ccaagcgacg cccaacctgc catcacgaga
     4081 tttcgattcc accgccgcct tctatgaaag gttgggcttc ggaatcgttt tccgggacgc
     4141 cggctggatg atcctccagc gcggggatct catgctggag ttcttcgccc accccaactt
     4201 gtttattgca gcttataatg gttacaaata aagcaatagc atcacaaatt tcacaaataa
     4261 agcatttttt tcactgcatt ctagttgtgg tttgtccaaa ctcatcaatg tatcttatca
     4321 tgtctgtata ccgtcgacct ctagctagag cttggcgtaa tcatggtcat agctgtttcc
     4381 tgtgtgaaat tgttatccgc tcacaattcc acacaacata cgagccggaa gcataaagtg
     4441 taaagcctgg ggtgcctaat gagtgagcta actcacatta attgcgttgc gctcactgcc
     4501 cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg
     4561 gagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc
     4621 ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag gcggtaatac ggttatccac
     4681 agaatcaggg gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa
     4741 ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc cgcccccctg acgagcatca
     4801 caaaaatcga cgctcaagtc agaggtggcg aaacccgaca ggactataaa gataccaggc
     4861 gtttccccct ggaagctccc tcgtgcgctc tcctgttccg accctgccgc ttaccggata
     4921 cctgtccgcc tttctccctt cgggaagcgt ggcgctttct catagctcac gctgtaggta
     4981 tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac cccccgttca
     5041 gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag tccaacccgg taagacacga
     5101 cttatcgcca ctggcagcag ccactggtaa caggattagc agagcgaggt atgtaggcgg
     5161 tgctacagag ttcttgaagt ggtggcctaa ctacggctac actagaagaa cagtatttgg
     5221 tatctgcgct ctgctgaagc cagttacctt cggaaaaaga gttggtagct cttgatccgg
     5281 caaacaaacc accgctggta gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag
     5341 aaaaaaagga tctcaagaag atcctttgat cttttctacg gggtctgacg ctcagtggaa
     5401 cgaaaactca cgttaaggga ttttggtcat gagattatca aaaaggatct tcacctagat
     5461 ccttttaaat taaaaatgaa gttttaaatc aatctaaagt atatatgagt aaacttggtc
     5521 tgacagttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc tatttcgttc
     5581 atccatagtt gcctgactcc ccgtcgtgta gataactacg atacgggagg gcttaccatc
     5641 tggccccagt gctgcaatga taccgcgaga cccacgctca ccggctccag atttatcagc
     5701 aataaaccag ccagccggaa gggccgagcg cagaagtggt cctgcaactt tatccgcctc
     5761 catccagtct attaattgtt gccgggaagc tagagtaagt agttcgccag ttaatagttt
     5821 gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc
     5881 ttcattcagc tccggttccc aacgatcaag gcgagttaca tgatccccca tgttgtgcaa
     5941 aaaagcggtt agctccttcg gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt
     6001 atcactcatg gttatggcag cactgcataa ttctcttact gtcatgccat ccgtaagatg
     6061 cttttctgtg actggtgagt actcaaccaa gtcattctga gaatagtgta tgcggcgacc
     6121 gagttgctct tgcccggcgt caatacggga taataccgcg ccacatagca gaactttaaa
     6181 agtgctcatc attggaaaac gttcttcggg gcgaaaactc tcaaggatct taccgctgtt
     6241 gagatccagt tcgatgtaac ccactcgtgc acccaactga tcttcagcat cttttacttt
     6301 caccagcgtt tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa agggaataag
     6361 ggcgacacgg aaatgttgaa tactcatact cttccttttt caatattatt gaagcattta
     6421 tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa ataaacaaat
     6481 aggggttccg cgcacatttc cccgaaaagt gccacctgac gtcgacggat cgggagatct
     6541 cccgatcccc tatggtgcac tctcagtaca atctgctctg atgccgcata gttaagccag
     6601 tatctgctcc ctgcttgtgt gttggaggtc gctgagtagt gcgcgagcaa aatttaagct
     6661 acaacaaggc aaggcttgac cgacaatt
//
