LOCUS       sequence-339470-        4368 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Crowding FRET sensor with mCerulean3 and mCitrine for E. coli 
            expression. Linker comprises 18x GSG.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4368)
  AUTHORS   Liu B, Aberg C, van Eerden FJ, Marrink SJ, Poolman B, Boersma AJ
  TITLE     Design and Properties of Genetically Encoded Probes for Sensing 
            Macromolecular Crowding.
  JOURNAL   Biophys J. 2017 May 9;112(9):1929-1939. doi: 
            10.1016/j.bpj.2017.04.004.
  PUBMED    28494963
REFERENCE   2  (bases 1 to 4368)
  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"; doi: 
            "10.1016/j.bpj.2017.04"; journalName: "Biophys J"; date: "2017-05-9-
            9"; volume: "112"; issue: "9"; pages: "1929-1939"
COMMENT     Sequence Label: pRSETA_crG18
FEATURES             Location/Qualifiers
     source          1..4368
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     20..39
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        20..38
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     RBS             70..92
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             103..120
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             1305..1316
                     /codon_start=1
                     /product="WELQut protease recognition and cleavage site"
                     /label=WELQut site
                     /translation="WELQ"
     primer_bind     complement(1769..1787)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1783..1830
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      1927..2382
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2014..2033)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2224..2245
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        2408..2512
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2513..2581,2582..3373)
                     /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(2731..2750)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3544..4132
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4033..4052
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4286..4303
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 gatctcgatc ccgcgaaatt aatacgactc actataggga gaccacaacg gtttccctct
       61 agaaataatt ttgtttaact ttaagaagga gatatacata tgcatcatca tcaccatcat
      121 gtgagcaaag gtgaagagct ctttaccggt gttgttccga ttctggttga actggatggt
      181 gacgttaatg gtcacaaatt ttcagttagc ggtgaaggcg aaggtgatgc aacctatggt
      241 aaactgaccc tgaaatttat ctgtaccacc ggcaaactgc cggttccgtg gccgaccctg
      301 gttaccaccc tgagctgggg tgttcagtgt tttgcacgtt atccggatca catgaaacag
      361 cacgattttt tcaaaagcgc aatgccggaa ggttatgttc aagaacgtac catcttcttc
      421 aaagatgacg gcaactataa aacccgtgcc gaagttaaat ttgaaggtga taccctggtg
      481 aatcgcattg aactgaaagg catcgatttt aaagaggatg gtaatatcct gggccacaaa
      541 ctggaatata atgccattca tggcaacgtg tatatcaccg cagataaaca gaaaaacggc
      601 atcaaagcaa attttggcct gaactgcaat attgaagatg gttcagttca gctggcagat
      661 cattatcagc agaatacccc gattggtgat ggtccggttc tgctgccgga taatcattat
      721 ctgagcaccc agagcaaact gagcaaagat ccgaatgaaa aacgtgatca catggtgctg
      781 ctggaatttg ttaccgcagc aggtattacc ctgggtatgg atgaactgta taaaggtagc
      841 ggtggtagtg gtggttcagg tggctctggt ggcagcggtg gatccggtgg tagcggtggt
      901 tcaggtggta gtggcggtag tggtggctca gggggttctg gtggctccgg tggctctggc
      961 ggtagcggag gcagtggtgg aagtggtggt tccatggtgt caaaaggtga ggaactgttt
     1021 acaggcgtgg tgccgatcct ggtagagctg gacggggatg tgaatggcca taaattcagc
     1081 gtttcaggtg aaggtgaggg cgacgccacg tacggaaaac tgacactgaa attcatttgc
     1141 acaacaggta aactgcctgt gccttggcct acactggtga ccacctttgg ttatggtctg
     1201 atgtgctttg ctcgctatcc tgaccacatg aaacaacatg atttctttaa atctgccatg
     1261 cctgaaggct acgtgcaaga gcgcaccatt tttttcaaag acgatgggaa ttacaaaaca
     1321 cgtgcggagg tgaaattcga gggcgataca ctggttaacc gtatcgagct gaaaggtatc
     1381 gacttcaaag aggacggaaa cattctgggt cataaactgg aatacaacta caacagccat
     1441 aacgtgtaca tcatggccga caaacaaaaa aacgggatta aagtgaactt caaaatccgc
     1501 cacaacatcg aagatggcag cgtgcagctg gccgaccact atcaacaaaa cacaccgatc
     1561 ggcgacggtc ctgtactgct gcctgacaac cactatctgt catatcagag cgcactgtca
     1621 aaagatccta acgagaaacg cgaccacatg gttctgctgg aattcgtgac agccgctggc
     1681 attacactgg gcatggacga gctgtacaaa taaaagcttg atccggctgc taacaaagcc
     1741 cgaaaggaag ctgagttggc tgctgccacc gctgagcaat aactagcata accccttggg
     1801 gcctctaaac gggtcttgag gggttttttg ctgaaaggag gaactatatc cggatctggc
     1861 gtaatagcga agaggcccgc accgatcgcc cttcccaaca gttgcgcagc ctgaatggcg
     1921 aatgggacgc gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg
     1981 tgaccgctac acttgccagc gccctagcgc ccgctccttt cgctttcttc ccttcctttc
     2041 tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg ggggctccct ttagggttcc
     2101 gatttagtgc tttacggcac ctcgacccca aaaaacttga ttagggtgat ggttcacgta
     2161 gtgggccatc gccctgatag acggtttttc gccctttgac gttggagtcc acgttcttta
     2221 atagtggact cttgttccaa actggaacaa cactcaaccc tatctcggtc tattcttttg
     2281 atttataagg gattttgccg atttcggcct attggttaaa aaatgagctg atttaacaaa
     2341 aatttaacgc gaattttaac aaaatattaa cgcttacaat ttaggtggca cttttcgggg
     2401 aaatgtgcgc ggaaccccta tttgtttatt tttctaaata cattcaaata tgtatccgct
     2461 catgagacaa taaccctgat aaatgcttca ataatattga aaaaggaaga gtatgagtat
     2521 tcaacatttc cgtgtcgccc ttattccctt ttttgcggca ttttgccttc ctgtttttgc
     2581 tcacccagaa acgctggtga aagtaaaaga tgctgaagat cagttgggtg cacgagtggg
     2641 ttacatcgaa ctggatctca acagcggtaa gatccttgag agttttcgcc ccgaagaacg
     2701 ttttccaatg atgagcactt ttaaagttct gctatgtggc gcggtattat cccgtattga
     2761 cgccgggcaa gagcaactcg gtcgccgcat acactattct cagaatgact tggttgagta
     2821 ctcaccagtc acagaaaagc atcttacgga tggcatgaca gtaagagaat tatgcagtgc
     2881 tgccataacc atgagtgata acactgcggc caacttactt ctgacaacga tcggaggacc
     2941 gaaggagcta accgcttttt tgcacaacat gggggatcat gtaactcgcc ttgatcgttg
     3001 ggaaccggag ctgaatgaag ccataccaaa cgacgagcgt gacaccacga tgcctgtagc
     3061 aatggcaaca acgttgcgca aactattaac tggcgaacta cttactctag cttcccggca
     3121 acaattaata gactggatgg aggcggataa agttgcagga ccacttctgc gctcggccct
     3181 tccggctggc tggtttattg ctgataaatc tggagccggt gagcgtgggt ctcgcggtat
     3241 cattgcagca ctggggccag atggtaagcc ctcccgtatc gtagttatct acacgacggg
     3301 gagtcaggca actatggatg aacgaaatag acagatcgct gagataggtg cctcactgat
     3361 taagcattgg taactgtcag accaagttta ctcatatata ctttagattg atttaaaact
     3421 tcatttttaa tttaaaagga tctaggtgaa gatccttttt gataatctca tgaccaaaat
     3481 cccttaacgt gagttttcgt tccactgagc gtcagacccc gtagaaaaga tcaaaggatc
     3541 ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct
     3601 accagcggtg gtttgtttgc cggatcaaga gctaccaact ctttttccga aggtaactgg
     3661 cttcagcaga gcgcagatac caaatactgt tcttctagtg tagccgtagt taggccacca
     3721 cttcaagaac tctgtagcac cgcctacata cctcgctctg ctaatcctgt taccagtggc
     3781 tgctgccagt ggcgataagt cgtgtcttac cgggttggac tcaagacgat agttaccgga
     3841 taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca cagcccagct tggagcgaac
     3901 gacctacacc gaactgagat acctacagcg tgagctatga gaaagcgcca cgcttcccga
     3961 agggagaaag gcggacaggt atccggtaag cggcagggtc ggaacaggag agcgcacgag
     4021 ggagcttcca gggggaaacg cctggtatct ttatagtcct gtcgggtttc gccacctctg
     4081 acttgagcgt cgatttttgt gatgctcgtc aggggggcgg agcctatgga aaaacgccag
     4141 caacgcggcc tttttacggt tcctggcctt ttgctggcct tttgctcaca tgttctttcc
     4201 tgcgttatcc cctgattctg tggataaccg tattaccgcc tttgagtgag ctgataccgc
     4261 tcgccgcagc cgaacgaccg agcgcagcga gtcagtgagc gaggaagcgg aagagcgccc
     4321 aatacgcaaa ccgcctctcc ccgcgcgttg gccgattcat taatgcag
//
