LOCUS       sequence-44716-e        5678 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 5678)
  AUTHORS   Maine GN, Li H, Zaidi IW, Basrur V, Elenitoba-Johnson KS, Burstein E
  TITLE     A bimolecular affinity purification method under denaturing 
            conditions for rapid isolation of a ubiquitinated protein for mass 
            spectrometry analysis.
  JOURNAL   Nat Protoc. 2010 Aug;5(8):1447-59. Epub 2010 Jul 22.
  PUBMED    20671728
REFERENCE   2  (bases 1 to 5678)
  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: "Nat 
            Protoc."; date: "2010-08"; volume: "5(8)"; pages: "1447-59. Epub 
            2010 Jul 22"
COMMENT     Sequence Label: pEBB-TB
FEATURES             Location/Qualifiers
     source          1..5678
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    107..128
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(143..148,149..166,167..173)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    181..197
                     /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)."
     primer_bind     186..208
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     205..221
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     205..221
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     promoter        252..462
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     promoter        274..470
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      321..456
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     promoter        557..1739
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          788..1730
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     1687..1707
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     CDS             1841..1861
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     primer_bind     complement(2812..2829)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2812..2828)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2821..2843)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      3041..3496
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3128..3147)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3338..3359
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3513..3532)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3632..3654
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3692..3710)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3778..3882
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3883..3951,3952..4743)
                     /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     complement(4101..4120)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4914..5502
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5403..5422
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5656..5673
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 agcgcccaat acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagctggc
       61 acgacaggtt tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat gtgagttagc
      121 tcactcatta ggcaccccag gctttacact ttatgcttcc ggctcgtatg ttgtgtggaa
      181 ttgtgagcgg ataacaattt cacacaggaa acagctatga ccatgattac gccaagcttc
      241 caggctcccc agcaggcaga agtatgcaaa gcatgcatct caattagtca gcaaccatag
      301 tcccgcccct aactccgccc atcccgcccc taactccgcc cagttccgcc cattctccgc
      361 cccatggctg actaattttt tttatttatg cagaggccga ggccgcctcg gcctctgagc
      421 tattccagaa gtagtgagga ggcttttttg gaggcctagg cttttgcaaa aagctttgca
      481 aagatggata aagttttaaa cagagaggaa tctttgcagc taatggacct tctaggtctt
      541 gaaaggagtg ggaattggct ccggtgcccg tcagtgggca gagcgcacat cgcccacagt
      601 ccccgagaag ttggggggag gggtcggcaa ttgaaccggt gcctagagaa ggtggcgcgg
      661 ggtaaactgg gaaagtgatg tcgtgtactg gctccgcctt tttcccgagg gtgggggaga
      721 accgtatata agtgcagtag tcgccgtgaa cgttcttttt cgcaacgggt ttgccgccag
      781 aacacaggta agtgccgtgt gtggttcccg cgggcctggc ctctttacgg gttatggccc
      841 ttgcgtgcct tgaattactt ccacgcccct ggctgcagta cgtgattctt gatcccgagc
      901 ttcgggttgg aagtgggtgg gagagttcga ggccttgcgc ttaaggagcc ccttcgcctc
      961 gtgcttgagt tgaggcctgg cctgggcgct ggggccgccg cgtgcgaatc tggtggcacc
     1021 ttcgcgcctg tctcgctgct ttcgataagt ctctagccat ttaaaatttt tgatgacctg
     1081 ctgcgacgct ttttttctgg caagatagtc ttgtaaatgc gggccaagat ctgcacactg
     1141 gtatttcggt ttttggggcc gcgggcggcg acggggcccg tgcgtcccag cgcacatgtt
     1201 cggcgaggcg gggcctgcga gcgcggccac cgagaatcgg acgggggtag tctcaagctg
     1261 gccggcctgc tctggtgcct ggcctcgcgc cgccgtgtat cgccccgccc tgggcggcaa
     1321 ggctggcccg gtcggcacca gttgcgtgag cggaaagatg gccgcttccc ggccctgctg
     1381 cagggagctc aaaatggagg acgcggcgct cgggagagcg ggcgggtgag tcacccacac
     1441 aaaggaaaag ggcctttccg tcctcagccg tcgcttcatg tgactccacg gagtaccggg
     1501 cgccgtccag gcacctcgat tagttctcga gcttttggag tacgtcgtct ttaggttggg
     1561 gggaggggtt ttatgcgatg gagtttcccc acactgagtg ggtggagact gaagttaggc
     1621 cagcttggca cttgatgtaa ttctccttgg aatttgccct ttttgagttt ggatcttggt
     1681 tcattctcaa gcctcagaca gtggttcaaa gtttttttct tccatttcag gtgtcgtgag
     1741 gaattctcta gagatccctc gacctcgaga tccattgtgc tggatccact agtggtacca
     1801 tcgatgcggc cgcgactacg atatacccac aaccgccagc gagaatctgt actttcaatc
     1861 agagttaaaa accgccgcac tagctcaaca tgacgaggct gctggaaagg ccggtgaagg
     1921 tgaaatccct gcccctcttg ctggtaccgt ttctaagata ctggtaaaag aaggtgacac
     1981 tgttaaagct ggtcaaacag ttctggtgct ggaggctatg aaaatggaga cagaaattaa
     2041 cgctcctact gacggaaaag ttgaaaaggt gttagttaag gaaagagatg ctgttcaagg
     2101 tggtcaaggt ctaatcaaga tcggcgttta agggccgcga ctctagagtg agggtcccca
     2161 cctgggaccc ttgagagtat caggtctccc acgtgggaga caagaaatcc ctgtttaata
     2221 tttaaacagc agtgttcccc atctgggtcc ttgcacccct cactctggcc tcagccgact
     2281 gcacagcggc ccctgcatcc ccttggctgt gaggcccctg gacaagcaga ggtggccaga
     2341 gctgggaggc atggccctgg ggtcccacga atttgctggg gaatctcgtt tttcttctta
     2401 agacttttgg gacatggttt gactcccgaa catcaccgac gcgtctcctg tttttctggg
     2461 tggcctcggg acacctgccc tgcccccacg agggtcagga ctgtgactct ttttagggcc
     2521 aggcaggtgc ctggacattt gccttgctgg acggggactg gggatgtggg agggagcaga
     2581 caggaggaat catgtcaggc ctgtgtgtga aaggaagctc cactgtcacc ctccacctct
     2641 tcacccccca ctcaccagtg tcccctccac tgtcacattg taactgaact tcaggataat
     2701 aaagtgtttg cctccaaaaa aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa
     2761 aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa aaaaagaatt cactggccgt
     2821 cgttttacaa cgtcgtgact gggaaaaccc tggcgttacc caacttaatc gccttgcagc
     2881 acatccccct ttcgccagct ggcgtaatag cgaagaggcc cgcaccgatc gcccttccca
     2941 acagttgcgc agcctgaatg gcgaatggcg cctgatgcgg tattttctcc ttacgcatct
     3001 gtgcggtatt tcacaccgca tacgtcaaag caaccatagt acgcgccctg tagcggcgca
     3061 ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc cagcgcccta
     3121 gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg ctttccccgt
     3181 caagctctaa atcgggggct ccctttaggg ttccgattta gtgctttacg gcacctcgac
     3241 cccaaaaaac ttgatttggg tgatggttca cgtagtgggc catcgccctg atagacggtt
     3301 tttcgccctt tgacgttgga gtccacgttc tttaatagtg gactcttgtt ccaaactgga
     3361 acaacactca accctatctc gggctattct tttgatttat aagggatttt gccgatttcg
     3421 gcctattggt taaaaaatga gctgatttaa caaaaattta acgcgaattt taacaaaata
     3481 ttaacgttta caattttatg gtgcactctc agtacaatct gctctgatgc cgcatagtta
     3541 agccagcccc gacacccgcc aacacccgct gacgcgccct gacgggcttg tctgctcccg
     3601 gcatccgctt acagacaagc tgtgaccgtc tccgggagct gcatgtgtca gaggttttca
     3661 ccgtcatcac cgaaacgcgc gagacgaaag ggcctcgtga tacgcctatt tttataggtt
     3721 aatgtcatga taataatggt ttcttagacg tcaggtggca cttttcgggg aaatgtgcgc
     3781 ggaaccccta tttgtttatt tttctaaata cattcaaata tgtatccgct catgagacaa
     3841 taaccctgat aaatgcttca ataatattga aaaaggaaga gtatgagtat tcaacatttc
     3901 cgtgtcgccc ttattccctt ttttgcggca ttttgccttc ctgtttttgc tcacccagaa
     3961 acgctggtga aagtaaaaga tgctgaagat cagttgggtg cacgagtggg ttacatcgaa
     4021 ctggatctca acagcggtaa gatccttgag agttttcgcc ccgaagaacg ttttccaatg
     4081 atgagcactt ttaaagttct gctatgtggc gcggtattat cccgtattga cgccgggcaa
     4141 gagcaactcg gtcgccgcat acactattct cagaatgact tggttgagta ctcaccagtc
     4201 acagaaaagc atcttacgga tggcatgaca gtaagagaat tatgcagtgc tgccataacc
     4261 atgagtgata acactgcggc caacttactt ctgacaacga tcggaggacc gaaggagcta
     4321 accgcttttt tgcacaacat gggggatcat gtaactcgcc ttgatcgttg ggaaccggag
     4381 ctgaatgaag ccataccaaa cgacgagcgt gacaccacga tgcctgtagc aatggcaaca
     4441 acgttgcgca aactattaac tggcgaacta cttactctag cttcccggca acaattaata
     4501 gactggatgg aggcggataa agttgcagga ccacttctgc gctcggccct tccggctggc
     4561 tggtttattg ctgataaatc tggagccggt gagcgtgggt ctcgcggtat cattgcagca
     4621 ctggggccag atggtaagcc ctcccgtatc gtagttatct acacgacggg gagtcaggca
     4681 actatggatg aacgaaatag acagatcgct gagataggtg cctcactgat taagcattgg
     4741 taactgtcag accaagttta ctcatatata ctttagattg atttaaaact tcatttttaa
     4801 tttaaaagga tctaggtgaa gatccttttt gataatctca tgaccaaaat cccttaacgt
     4861 gagttttcgt tccactgagc gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat
     4921 cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct accagcggtg
     4981 gtttgtttgc cggatcaaga gctaccaact ctttttccga aggtaactgg cttcagcaga
     5041 gcgcagatac caaatactgt ccttctagtg tagccgtagt taggccacca cttcaagaac
     5101 tctgtagcac cgcctacata cctcgctctg ctaatcctgt taccagtggc tgctgccagt
     5161 ggcgataagt cgtgtcttac cgggttggac tcaagacgat agttaccgga taaggcgcag
     5221 cggtcgggct gaacgggggg ttcgtgcaca cagcccagct tggagcgaac gacctacacc
     5281 gaactgagat acctacagcg tgagctatga gaaagcgcca cgcttcccga agggagaaag
     5341 gcggacaggt atccggtaag cggcagggtc ggaacaggag agcgcacgag ggagcttcca
     5401 gggggaaacg cctggtatct ttatagtcct gtcgggtttc gccacctctg acttgagcgt
     5461 cgatttttgt gatgctcgtc aggggggcgg agcctatgga aaaacgccag caacgcggcc
     5521 tttttacggt tcctggcctt ttgctggcct tttgctcaca tgttctttcc tgcgttatcc
     5581 cctgattctg tggataaccg tattaccgcc tttgagtgag ctgataccgc tcgccgcagc
     5641 cgaacgaccg agcgcagcga gtcagtgagc gaggaagc
//
