LOCUS       sequence-59322-e        6690 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6690)
  AUTHORS   Lamothe B, Besse A, Campos AD, Webster WK, Wu H, Darnay BG
  TITLE     Site-specific Lys-63-linked tumor necrosis factor 
            receptor-associated factor 6 auto-ubiquitination is a critical 
            determinant of I kappa B kinase activation.
  JOURNAL   J Biol Chem. 2007 Feb 9;282(6):4102-12. Epub 2006 Nov 29.
  PUBMED    17135271
REFERENCE   2  (bases 1 to 6690)
  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: "J Biol 
            Chem."; date: "2007-02-9"; volume: "282(6)"; pages: "4102-12. Epub 
            2006 Nov 29"
COMMENT     Sequence Label: pCR3-FLAG-TRAF6-C70A
FEATURES             Location/Qualifiers
     source          1..6690
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        10..389
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        390..593
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     544..564
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     638..657
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        638..656
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             722..745
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             755..778
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     promoter        complement(2405..2423)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(2406..2423)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(2443..2460)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2449..2673
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      complement(2805..3393)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2885..2904)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     polyA_signal    3722..3769
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     primer_bind     3794..3813
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     CDS             complement(4001..4789)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDQEHQGL
                     APAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIALA
                     TRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(4112..4131)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     4716..4735
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     promoter        complement(4824..5181)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4838..4973
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     enhancer        4990..5181
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     5138..5158
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     CDS             complement(join(5241..6032,6033..6101))
                     /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     5864..5883
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6102..6206)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      6233..6688
                     /direction=RIGHT
                     /label=M13 ori
                     /note="M13 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6320..6339)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     6530..6551
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
ORIGIN
        1 gcgcgcgttg acattgatta ttgactagtt attaatagta atcaattacg gggtcattag
       61 ttcatagccc atatatggag ttccgcgtta cataacttac ggtaaatggc ccgcctggct
      121 gaccgcccaa cgacccccgc ccattgacgt caataatgac gtatgttccc atagtaacgc
      181 caatagggac tttccattga cgtcaatggg tggactattt acggtaaact gcccacttgg
      241 cagtacatca agtgtatcat atgccaagta cgccccctat tgacgtcaat gacggtaaat
      301 ggcccgcctg gcattatgcc cagtacatga ccttatggga ctttcctact tggcagtaca
      361 tctacgtatt agtcatcgct attaccatgg tgatgcggtt ttggcagtac atcaatgggc
      421 gtggatagcg gtttgactca cggggatttc caagtctcca ccccattgac gtcaatggga
      481 gtttgttttg gcaccaaaat caacgggact ttccaaaatg tcgtaacaac tccgccccat
      541 tgacgcaaat gggcggtagg cgtgtacggt gggaggtcta tataagcaga gctctctggc
      601 taactagaga acccactgct tactggctta tcgaaattaa tacgactcac tatagggaga
      661 cccaagcttg gtaccgagct cggatccact agtaacggcc gccagtgtgc tggaattgat
      721 ggactacaag gacgacgatg acaaggaatt gatggactac aaggacgacg atgacaagga
      781 attcatgagt ctcttaaact gtgagaacag ctgcgggtcc agccagtcgt ccagtgactg
      841 ctgcgctgcc atggccgcct cctgcagcgc tgcagtgaaa gatgacagcg tgagtggctc
      901 tgccagcacc gggaacctct ccagctcctt catggaggag atccagggct acgatgtgga
      961 gtttgaccca cctctggaga gcaagtatga ggctcccatc tgcttgatgg ctttacggga
     1021 agcagtgcaa acaccatgtg gccacaggtt ctgcaaagcc tgcatcatca aatccataag
     1081 ggatgcaggg cacaagtgcc cagttgacaa tgaaatactg ctggaaaatc aactgtttcc
     1141 cgacaatttt gcaaagcgag agattctttc cctgacggta aagtgcccaa ataaaggctg
     1201 tttgcaaaag atggaactga gacatctcga ggatcatcaa gtacattgtg aatttgctct
     1261 agtgaattgt ccccagtgcc aacgtccttt ccagaagtgc caggttaata cacacattat
     1321 tgaggattgt cccaggaggc aggtttcttg tgtaaactgt gctgtgtcca tggcatatga
     1381 agagaaagag atccatgatc aaagctgtcc tctggcaaat atcatctgtg aatactgtgg
     1441 tacaatcctc atcagagaac agatgcctaa tcattatgat ctggactgcc caacagctcc
     1501 aatcccttgc acattcagtg tttttggctg tcatgaaaag atgcagagga atcacttggc
     1561 acgacacttg caagagaata cccagttgca catgagactg ttggcccagg ctgttcataa
     1621 tgttaacctt gctttgcgtc cgtgcgatgc cgcctctcca tcccggggat gtcgtccaga
     1681 ggacccaaat tatgaggaaa ctatcaaaca gttggagagt cgcctagtaa gacaggacca
     1741 tcagatccgg gagctgactg ccaaaatgga aactcagagt atgtacgtgg gcgagctcaa
     1801 acggaccatt cggaccctgg aggacaaggt tgccgaaatg gaagcacagc agtgtaacgg
     1861 gatctacatt tggaagattg gcaactttgg gatgcacttg aaatcccaag aagaggaaag
     1921 acctgttgtc atccatagcc ctggattcta cacaggcaga cctgggtaca agctgtgcat
     1981 gcgcctgcat cttcagttac cgacagctca gcgctgtgca aactatatat ccctttttgt
     2041 ccacacaatg caaggagaat atgacagcca cctcccctgg cccttccagg gtacaatacg
     2101 ccttacaatt ctcgaccagt ctgaagcact tataaggcaa aaccacgaag aggtcatgga
     2161 cgccaaacca gaactgcttg cctttcagcg acccacaatc ccacggaacc ccaaaggttt
     2221 tggctatgta acatttatgc acctggaagc cttaagacag ggaaccttca ttaaggatga
     2281 tacattacta gtgcgctgtg aagtctctac ccgctttgac atgggtggcc ttcggaagga
     2341 gggtttccag ccacgaagta ctgatgcggg ggtgtaggtc gagcatgcat ctagagggcc
     2401 ctattctata gtgtcaccta aatgctagag ctcgctgatc agcctcgact gtgccttcta
     2461 gttgccagcc atctgttgtt tgcccctccc ccgtgccttc cttgaccctg gaaggtgcca
     2521 ctcccactgt cctttcctaa taaaatgagg aaattgcatc gcattgtctg agtaggtgtc
     2581 attctattct ggggggtggg gtggggcagg acagcaaggg ggaggattgg gaagacaata
     2641 gcaggcatgc tggggatgcg gtgggctcta tggcttctga ggcggaaaga accagtggcg
     2701 gtaatacggt tatccacaga atcaggggat aacgcaggaa agaacatgtg agcaaaaggc
     2761 cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca taggctccgc
     2821 ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga
     2881 ctataaagat accaggcgtt tccccctgga agctccctcg tgcgctctcc tgttccgacc
     2941 ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc gctttctcat
     3001 agctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct gggctgtgtg
     3061 cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg tcttgagtcc
     3121 aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag gattagcaga
     3181 gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta cggctacact
     3241 agaaggacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt
     3301 ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt tgtttgcaag
     3361 cagcagatta cgcgcagaaa aaaaggatct caagaagatc ctttgatctt ttctacgggg
     3421 tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgag attatcaaaa
     3481 aggatcttca cctagatcct tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata
     3541 tatgagtaac ctgaggctat ggcagggcct gccgccccga cgttggctgc gagccctggg
     3601 ccttcacccg aacttggggg gtggggtggg gaaaaggaag aaacgcgggc gtattggccc
     3661 caatggggtc tcggtggggt atcgacagag tgccagccct gggaccgaac cccgcgttta
     3721 tgaacaaacg acccaacacc gtgcgtttta ttctgtcttt ttattgccgt catagcgcgg
     3781 gttccttccg gtattgtctc cttccgtgtt tcagttagcc tccccctagg gtgggcgaag
     3841 aactccagca tgagatcccc gcgctggagg atcatccagc cggcgtcccg gaaaacgatt
     3901 ccgaagccca acctttcata gaaggcggcg gtggaatcga aatctcgtga tggcaggttg
     3961 ggcgtcgctt ggtcggtcat ttcgaacccc agagtcccgc tcagaagaac tcgtcaagaa
     4021 ggcgatagaa ggcgatgcgc tgcgaatcgg gagcggcgat accgtaaagc acgaggaagc
     4081 ggtcagccca ttcgccgcca agctcttcag caatatcacg ggtagccaac gctatgtcct
     4141 gatagcggtc cgccacaccc agccggccac agtcgatgaa tccagaaaag cggccatttt
     4201 ccaccatgat attcggcaag caggcatcgc catgggtcac gacgagatcc tcgccgtcgg
     4261 gcatgctcgc cttgagcctg gcgaacagtt cggctggcgc gagcccctga tgctcttgat
     4321 catcctgatc gacaagaccg gcttccatcc gagtacgtgc tcgctcgatg cgatgtttcg
     4381 cttggtggtc gaatgggcag gtagccggat caagcgtatg cagccgccgc attgcatcag
     4441 ccatgatgga tactttctcg gcaggagcaa ggtgagatga caggagatcc tgccccggca
     4501 cttcgcccaa tagcagccag tcccttcccg cttcagtgac aacgtcgagc acagctgcgc
     4561 aaggaacgcc cgtcgtggcc agccacgata gccgcgctgc ctcgtcttgc agttcattca
     4621 gggcaccgga caggtcggtc ttgacaaaaa gaaccgggcg cccctgcgct gacagccgga
     4681 acacggcggc atcagagcag ccgattgtct gttgtgccca gtcatagccg aatagcctct
     4741 ccacccaagc ggccggagaa cctgcgtgca atccatcttg ttcaatcatg cgaaacgatc
     4801 ctcatcctgt ctcttgatcg atctttgcaa aagcctaggc ctccaaaaaa gcctcctcac
     4861 tacttctgga atagctcaga ggccgaggcg gcctcggcct ctgcataaat aaaaaaaatt
     4921 agtcagccat ggggcggaga atgggcggaa ctgggcggag ttaggggcgg gatgggcgga
     4981 gttaggggcg ggactatggt tgctgactaa ttgagatgca tgctttgcat acttctgcct
     5041 gctggggagc ctggggactt tccacacctg gttgctgact aattgagatg catgctttgc
     5101 atacttctgc ctgctgggga gcctggggac tttccacacc ctaactgaca cacattccac
     5161 agctggttct ttccgcctca ggactcttcc tttttcaata aatcaatcta aagtatatat
     5221 gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc
     5281 tgtctatttc gttcatccat agttgcctga ctccccgtcg tgtagataac tacgatacgg
     5341 gagggcttac catctggccc cagtgctgca atgataccgc gagacccacg ctcaccggct
     5401 ccagatttat cagcaataaa ccagccagcc ggaagggccg agcgcagaag tggtcctgca
     5461 actttatccg cctccatcca gtctattaat tgttgccggg aagctagagt aagtagttcg
     5521 ccagttaata gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt gtcacgctcg
     5581 tcgtttggta tggcttcatt cagctccggt tcccaacgat caaggcgagt tacatgatcc
     5641 cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag
     5701 ttggccgcag tgttatcact catggttatg gcagcactgc ataattctct tactgtcatg
     5761 ccatccgtaa gatgcttttc tgtgactggt gagtactcaa ccaagtcatt ctgagaatag
     5821 tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac gggataatac cgcgccacat
     5881 agcagaactt taaaagtgct catcattgga aaacgttctt cggggcgaaa actctcaagg
     5941 atcttaccgc tgttgagatc cagttcgatg taacccactc gtgcacccaa ctgatcttca
     6001 gcatctttta ctttcaccag cgtttctggg tgagcaaaaa caggaaggca aaatgccgca
     6061 aaaaagggaa taagggcgac acggaaatgt tgaatactca tactcttcct ttttcaatat
     6121 tattgaagca tttatcaggg ttattgtctc atgagcggat acatatttga atgtatttag
     6181 aaaaataaac aaataggggt tccgcgcaca tttccccgaa aagtgccacc tgacgcgccc
     6241 tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt
     6301 gccagcgccc tagcgcccgc tcctttcgct ttcttccctt cctttctcgc cacgttcgcc
     6361 ggctttcccc gtcaagctct aaatcggggg ctccctttag ggttccgatt tagtgcttta
     6421 cggcacctcg accccaaaaa acttgattag ggtgatggtt cacgtagtgg gccatcgccc
     6481 tgatagacgg tttttcgccc tttgacgttg gagtccacgt tctttaatag tggactcttg
     6541 ttccaaactg gaacaacact caaccctatc tcggtctatt cttttgattt ataagggatt
     6601 ttgccgattt cggcctattg gttaaaaaat gagctgattt aacaaaaatt taacgcgaat
     6661 tttaacaaaa tattaacgct tacaatttac
//
