LOCUS       sequence-435059-        3110 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  golden gate cloning.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3110)
  AUTHORS   Delhaye L, Moschonas GD, Fijalkowska D, Verhee A, De Sutter D, Van 
            de Steene T, De Meyer M, Grzesik H, Van Moortel L, De Bosscher K, 
            Jacobs T, Eyckerman S
  TITLE     Leveraging a self-cleaving peptide for tailored control in proximity
            labeling proteomics.
  JOURNAL   Cell Rep Methods. 2024 Jul 15;4(7):100818. doi: 
            10.1016/j.crmeth.2024.100818. Epub 2024 Jul 9.
  PUBMED    38986614
REFERENCE   2  (bases 1 to 3110)
  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: "Cell Rep 
            Methods."; date: "2024-07-15"; volume: "4(7):100818. doi"; pages: " 
            10.1016/j.crmeth.2024.100818. Epub 2024 Jul 9"
COMMENT     Sequence Label: pGGA-pTRE-B
FEATURES             Location/Qualifiers
     source          1..3110
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    85..106
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(121..126,127..144,145..151)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    159..175
                     /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        189..207
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     189..206
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     protein_bind    join(253..271,272..294,295..313,314..336,337..355,356..378,
                     379..397,398..420,421..439,440..462,463..481,482..504,
                     505..523)
                     /label=tetracycline response element
                     /bound_moiety="tetracycline repressor TetR"
                     /note="contains seven copies of the tetracycline operator 
                     tetO"
     primer_bind     550..574
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     primer_bind     complement(673..692)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(674..692)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(702..724)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     complement(918..937)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     1037..1059
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(1097..1115)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        1183..1287
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1288..1356,1357..2148)
                     /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(1506..1525)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2319..2907
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2808..2827
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3061..3078
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 tctccccgcg cgttggccga ttcattaatg cagctggcac gacaggtttc ccgactggaa
       61 agcgggcagt gagcgcaacg caattaatgt gagttagctc actcattagg caccccaggc
      121 tttacacttt atgcttccgg ctcgtatgtt gtgtggaatt gtgagcggat aacaatttca
      181 cacgagctat ttaggtgaca ctatagaagt gaagcttggt ctcaacctgg taccgagctc
      241 gactttcact tttctctatc actgataggg agtggtaaac tcgactttca cttttctcta
      301 tcactgatag ggagtggtaa actcgacttt cacttttctc tatcactgat agggagtggt
      361 aaactcgact ttcacttttc tctatcactg atagggagtg gtaaactcga ctttcacttt
      421 tctctatcac tgatagggag tggtaaactc gactttcact tttctctatc actgataggg
      481 agtggtaaac tcgactttca cttttctcta tcactgatag ggagtggtaa actcgaccta
      541 tataagcaga gctcgtttag tgaaccgtca gatcgcctgc agacgccatc cacgctgttt
      601 tgacctccat agaagacacc gggaccgatc cagcctccgc ggccccgaat tgaacatgag
      661 accgaattct cgccctatag tgagtcgtat tacctggccg tcgttttaca acgtcgtgac
      721 tgggaaaacc ctggcgttac ccaacttaat cgccttgcag cacatccccc tttcgccagc
      781 tggcgtaata gcgaagaggc ccgcaccgat cgcccttccc aacagttgcg cagcctgaat
      841 ggcgaatggc gcctgatgcg gtattttctc cttacgcatc tgtgcggtat ttcacaccgc
      901 atatggtgca ctctcagtac aatctgctct gatgccgcat agttaagcca gccccgacac
      961 ccgccaacac ccgctgacgc gccctgacgg gcttgtctgc tcccggcatc cgcttacaga
     1021 caagctgtga ccgtctccgg gagctgcatg tgtcagaggt tttcaccgtc atcaccgaaa
     1081 cgcgcgagac gaaagggcct cgtgatacgc ctatttttat aggttaatgt catgataata
     1141 atggtttctt agacgtcagg tggcactttt cggggaaatg tgcgcggaac ccctatttgt
     1201 ttatttttct aaatacattc aaatatgtat ccgctcatga gacaataacc ctgataaatg
     1261 cttcaataat attgaaaaag gaagagtatg agtattcaac atttccgtgt cgcccttatt
     1321 cccttttttg cggcattttg ccttcctgtt tttgctcacc cagaaacgct ggtgaaagta
     1381 aaagatgctg aagatcagtt gggtgcacga gtgggttaca tcgaactgga tctcaacagc
     1441 ggtaagatcc ttgagagttt tcgccccgaa gaacgttttc caatgatgag cacttttaaa
     1501 gttctgctat gtggcgcggt attatcccgt attgacgccg ggcaagagca actcggtcgc
     1561 cgcatacact attctcagaa tgacttggtt gagtactcac cagtcacaga aaagcatctt
     1621 acggatggca tgacagtaag agaattatgc agtgctgcca taaccatgag tgataacact
     1681 gcggccaact tacttctgac aacgatcgga ggaccgaagg agctaaccgc ttttttgcac
     1741 aacatggggg atcatgtaac tcgccttgat cgttgggaac cggagctgaa tgaagccata
     1801 ccaaacgacg agcgtgacac cacgatgcct gtagcaatgg caacaacgtt gcgcaaacta
     1861 ttaactggcg aactacttac tctagcttcc cggcaacaat taatagactg gatggaggcg
     1921 gataaagttg caggaccact tctgcgctcg gcccttccgg ctggctggtt tattgctgat
     1981 aaatctggag ccggtgagcg tggctctcga ggtatcattg cagcactggg gccagatggt
     2041 aagccctccc gtatcgtagt tatctacacg acggggagtc aggcaactat ggatgaacga
     2101 aatagacaga tcgctgagat aggtgcctca ctgattaagc attggtaact gtcagaccaa
     2161 gtttactcat atatacttta gattgattta aaacttcatt tttaatttaa aaggatctag
     2221 gtgaagatcc tttttgataa tctcatgacc aaaatccctt aacgtgagtt ttcgttccac
     2281 tgagcgtcag accccgtaga aaagatcaaa ggatcttctt gagatccttt ttttctgcgc
     2341 gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg tttgccggat
     2401 caagagctac caactctttt tccgaaggta actggcttca gcagagcgca gataccaaat
     2461 actgttcttc tagtgtagcc gtagttaggc caccacttca agaactctgt agcaccgcct
     2521 acatacctcg ctctgctaat cctgttacca gtggctgctg ccagtggcga taagtcgtgt
     2581 cttaccgggt tggactcaag acgatagtta ccggataagg cgcagcggtc gggctgaacg
     2641 gggggttcgt gcacacagcc cagcttggag cgaacgacct acaccgaact gagataccta
     2701 cagcgtgagc tatgagaaag cgccacgctt cccgaaggga gaaaggcgga caggtatccg
     2761 gtaagcggca gggtcggaac aggagagcgc acgagggagc ttccaggggg aaacgcctgg
     2821 tatctttata gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc
     2881 tcgtcagggg ggcggagcct atggaaaaac gccagcaacg cggccttttt acggttcctg
     2941 gccttttgct ggccttttgc tcacatgttc tttcctgcgt tatcccctga ttctgtggat
     3001 aaccgtatta ccgcctttga gtgagctgat accgctcgcc gcagccgaac gaccgagcgc
     3061 agcgagtcag tgagcgagga agcggaagag cgcccaatac gcaaaccgcc
//
