LOCUS       sequence-198797-        4027 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses NRI protein in E. coli. The expression is controlled by a 
            pLac promoter..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4027)
  AUTHORS   Guo S, Yeung E, Murray RM
  TITLE     Implementation and System Identification of a Phosphorylation-Based 
            Insulator in a Cell-Free Transcription-Translation System
  JOURNAL   bioRxiv 122606
REFERENCE   2  (bases 1 to 4027)
  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: "bioRxiv 
            122606"
COMMENT     Sequence Label: pSG06-pLac-NRI
FEATURES             Location/Qualifiers
     source          1..4027
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     RBS             7..29
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     rep_origin      1732..2277
                     /direction=RIGHT
                     /label=p15A ori
                     /note="Plasmids containing the medium-copy-number p15A 
                     origin of replication can be propagated in E. coli cells 
                     that contain a second plasmid with the ColE1 origin.
                     "
     primer_bind     2178..2197
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2394..2411
                     /label=L4440
                     /note="L4440 vector, forward primer"
     CDS             complement(join(2644..3435,3436..3504))
                     /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     3267..3286
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3505..3609)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     3677..3695
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     protein_bind    3992..4008
                     /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)."
ORIGIN
        1 aataattttg tttaacttta agaaggagat ataccatgca acgagggata gtctgggtag
       61 tcgatgacga tagttccatc cgttgggtgc ttgaacgtgc gctcgctggg gcaggtttaa
      121 cctgtacgac gtttgagaac ggcgcagaag tgctggaggc gctggcgagc aaaacgccgg
      181 atgtgctgct ttcagatatc cgtatgccgg gaatggacgg gctggcgctg ctcaagcaga
      241 ttaaacagcg ccatccaatg cttccggtca tcattatgac cgcacattcc gatctggatg
      301 ctgccgtcag cgcctatcaa caaggggcgt ttgattatct gcccaaaccg tttgatatcg
      361 acgaagcagt ggcgctggtt gagcgcgcta tcagtcatta ccaggaacag cagcagccgc
      421 gtaatgttca gcttaacggc ccaacgaccg atatcatcgg cgaagcgcca gccatgcagg
      481 acgtgttccg tattatcggt cggctttcgc gttcttctat tagcgtgctg attaacggcg
      541 aatccggcac cggtaaagaa ctggtcgctc atgccctgca tcgccacagt ccgcgcgcca
      601 aagcgccgtt tatcgcgctg aatatggcag ctatcccaaa agatttgatc gaatcagaac
      661 tgtttggcca cgagaaaggc gcgtttactg gcgcgaatac cattcgtcag gggcgttttg
      721 aacaggccga tggcggtaca ttattcctcg acgaaattgg tgatatgccg ctggatgtgc
      781 agacgcgttt gctgcgcgtg ctggcagacg gtcagtttta ccgcgttggc ggctatgcgc
      841 cggtgaaagt ggatgtgcgg attatcgctg ccactcacca gaatctcgaa cagcgagtgc
      901 aggaaggtaa gttccgtgag gatctgttcc accgcctgaa cgttatccgc gttcatctgc
      961 cgccgctgcg cgaacgtcgg gaagatattc cccgtctggc gcgccatttt ttacaggttg
     1021 ccgcgcgcga actgggcgta gaagcgaagt tactgcatcc ggaaaccgaa gctgctctga
     1081 cgcgtctggc gtggccaggc aacgtgcgcc agctggaaaa cacctgccgc tggctaacgg
     1141 tgatggccgc cgggcaggaa gtgttgattc aggatttgcc cggcgaactg tttgaatcaa
     1201 cggttgcgga gagtacttcg caaatgcaac cggacagctg ggcgacgctt cttgcgcagt
     1261 gggcagacag agcgctgcgt tccggtcatc aaaatctgct ttccgaagcg cagccagagc
     1321 tggagcggac gttactgacg accgcgttgc gacatacgca ggggcataaa caggaagcgg
     1381 cgcggctact cggctggggc cgcaacaccc tgacgcgtaa gttaaaagag ctggggatgg
     1441 agtgactcga gcaaagcccg ccgaaaggcg ggcttttctg tgtcgaccga tgcccttgag
     1501 agccttcaac ccagtcagct ccttccggtg ggcgcggggc atgactatcg tcgccgcact
     1561 tatgactgtc ttctttatca tgcaactcgt aggacaggtg ccggcatgat aagctgtcaa
     1621 acatgagaat tgcaacttat atcgtatggg gctgacttca ggtgctacat ttgaagagat
     1681 aaattgcact gaaatctaga aatattttat ctgattaata agatgatctt cttgagatcg
     1741 ttttggtctg cgcgtaatct cttgctctga aaacgaaaaa accgccttgc agggcggttt
     1801 ttcgaaggtt ctctgagcta ccaactcttt gaaccgaggt aactggcttg gaggagcgca
     1861 gtcaccaaaa cttgtccttt cagtttagcc ttaaccggcg catgacttca agactaactc
     1921 ctctaaatca attaccagtg gctgctgcca gtggtgcttt tgcatgtctt tccgggttgg
     1981 actcaagacg atagttaccg gataaggcgc agcggtcgga ctgaacgggg ggttcgtgca
     2041 tacagtccag cttggagcga actgcctacc cggaactgag tgtcaggcgt ggaatgagac
     2101 aaacgcggcc ataacagcgg aatgacaccg gtaaaccgaa aggcaggaac aggagagcgc
     2161 acgagggagc cgccaggggg aaacgcctgg tatctttata gtcctgtcgg gtttcgccac
     2221 cactgatttg agcgtcagat ttcgtgatgc ttgtcagggg ggcggagcct atggaaaaac
     2281 ggctttgccg cggccctctc acttccctgt taagtatctt cctggcatct tccaggaaat
     2341 ctccgccccg ttcgtaagcc atttccgctc gccgcagtcg aacgaccgag cgtagcgagt
     2401 cagtgagcga ggaagcggaa tatatcctgt atcacatatt ctgctgacgc accggtgcag
     2461 ccttttttct cctgccacat gaagcacttc actgacaccc tcatcagtgc caacatagta
     2521 agccagtata cactccgcta gggtcatgag attatcaaaa aggatcttca cctagatcct
     2581 tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa cttggtctga
     2641 cagttaccaa tgcttaatca gtgaggcacc tatctcagcg atctgtctat ttcgttcatc
     2701 catagttgcc tgactccccg tcgtgtagat aactacgata cgggagggct taccatctgg
     2761 ccccagtgct gcaatgatac cgcgagaccc acgctcaccg gctccagatt tatcagcaat
     2821 aaaccagcca gccggaaggg ccgagcgcag aagtggtcct gcaactttat ccgcctccat
     2881 ccagtctatt aattgttgcc gggaagctag agtaagtagt tcgccagtta atagtttgcg
     2941 caacgttgtt gccattgcta caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc
     3001 attcagctcc ggttcccaac gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa
     3061 agcggttagc tccttcggtc ctccgatcgt tgtcagaagt aagttggccg cagtgttatc
     3121 actcatggtt atggcagcac tgcataattc tcttactgtc atgccatccg taagatgctt
     3181 ttctgtgact ggtgagtact caaccaagtc attctgagaa tagtgtatgc ggcgaccgag
     3241 ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca catagcagaa ctttaaaagt
     3301 gctcatcatt ggaaaacgtt cttcggggcg aaaactctca aggatcttac cgctgttgag
     3361 atccagttcg atgtaaccca ctcgtgcacc caactgatct tcagcatctt ttactttcac
     3421 cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc
     3481 gacacggaaa tgttgaatac tcatactctt cctttttcaa tattattgaa gcatttatca
     3541 gggttattgt ctcatgagcg gatacatatt tgaatgtatt tagaaaaata aacaaatagg
     3601 ggttccgcgc acatttcccc gaaaagtgcc acctgacgtc taagaaacca ttattatcat
     3661 gacattaacc tataaaaata ggcgtatcac gaggcccttt cgtcttcaag aattctggcg
     3721 aatcctctga ccagccagaa aacgaccttt ctgtggtgaa accggatgct gcaattcaga
     3781 gcgccagcaa gtgggggaca gcagaagacc tgaccgccgc agagtggatg tttgacatgg
     3841 tgaagactat cgcaccatca gccagaaaac cgaattttgc tgggtgggct aacgatatcc
     3901 gcctgatgcg tgaacgtgac ggacgtaacc accgcgacat gtgtgtgctg ttccgctggg
     3961 catgcataaa tgtgagcgga taacattgac attgtgagcg gataacaaga tactgagcac
     4021 agctagc
//
