LOCUS       sequence-339474-        4419 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Crowding FRET sensor with mCerulean3 and mCitrine for E. coli 
            expression. Linker comprises 2 helices, and a 6AA GSG domain.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4419)
  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 4419)
  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_crE6G2
FEATURES             Location/Qualifiers
     source          1..4419
                     /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             1356..1367
                     /codon_start=1
                     /product="WELQut protease recognition and cleavage site"
                     /label=WELQut site
                     /translation="WELQ"
     primer_bind     complement(1820..1838)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1834..1881
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      1978..2433
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2065..2084)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2275..2296
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        2459..2563
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2564..2632,2633..3424)
                     /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(2782..2801)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3595..4183
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4084..4103
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4337..4354
                     /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 catggttctg
      781 ctcgagtttg ttaccgcagc aggtattacc ctgggtatgg atgaactgta taaagcagaa
      841 gcagcagcaa aagaagccgc tgccaaagaa gcggcagcga aagaggctgc cgcaaaagag
      901 gcagcagcga aagaagcagc ggctaaagca ggtagcggtg gtagtggtgc cgaggcagcc
      961 gctaaagaag ctgcggcaaa agaagcagcg gcaaaagagg ctgcagctaa agaagccgca
     1021 gcaaaagagg cagcagcaaa agccggtggg agtaaaggtg aggaactgtt tacaggcgtg
     1081 gtgccgatcc tggtagagct ggacggggat gtgaatggcc ataaattcag cgtttcaggt
     1141 gaaggtgagg gcgacgccac atacggaaaa ctgacactga aattcatttg cacaacaggt
     1201 aaactgcctg tgccttggcc tacactggtg accacctttg gttatggtct gatgtgcttt
     1261 gctcgctatc ctgaccacat gaaacaacat gatttcttta aatctgccat gcctgaaggc
     1321 tacgtgcaag agcgcaccat ttttttcaaa gacgatggga attacaaaac acgtgcggag
     1381 gtgaaattcg agggcgatac actggttaac cgtatcgagc tgaaaggtat cgacttcaaa
     1441 gaggacggaa acattctggg tcataaactg gaatacaact acaacagcca taacgtgtac
     1501 atcatggccg acaaacaaaa aaacgggatt aaagtgaact tcaaaatccg ccacaacatc
     1561 gaagatggca gcgtgcagct ggccgaccac tatcaacaaa acacaccgat cggcgacggt
     1621 cctgtactgc tgcctgacaa ccactatctg tcatatcaga gcgcactgtc aaaagatcct
     1681 aacgagaaac gcgaccacat ggttctgctg gaattcgtga cagccgctgg cattacactg
     1741 ggcatggacg agctgtacaa ataaaagctt gatccggctg ctaacaaagc ccgaaaggaa
     1801 gctgagttgg ctgctgccac cgctgagcaa taactagcat aaccccttgg ggcctctaaa
     1861 cgggtcttga ggggtttttt gctgaaagga ggaactatat ccggatctgg cgtaatagcg
     1921 aagaggcccg caccgatcgc ccttcccaac agttgcgcag cctgaatggc gaatgggacg
     1981 cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta
     2041 cacttgccag cgccctagcg cccgctcctt tcgctttctt cccttccttt ctcgccacgt
     2101 tcgccggctt tccccgtcaa gctctaaatc gggggctccc tttagggttc cgatttagtg
     2161 ctttacggca cctcgacccc aaaaaacttg attagggtga tggttcacgt agtgggccat
     2221 cgccctgata gacggttttt cgccctttga cgttggagtc cacgttcttt aatagtggac
     2281 tcttgttcca aactggaaca acactcaacc ctatctcggt ctattctttt gatttataag
     2341 ggattttgcc gatttcggcc tattggttaa aaaatgagct gatttaacaa aaatttaacg
     2401 cgaattttaa caaaatatta acgcttacaa tttaggtggc acttttcggg gaaatgtgcg
     2461 cggaacccct atttgtttat ttttctaaat acattcaaat atgtatccgc tcatgagaca
     2521 ataaccctga taaatgcttc aataatattg aaaaaggaag agtatgagta ttcaacattt
     2581 ccgtgtcgcc cttattccct tttttgcggc attttgcctt cctgtttttg ctcacccaga
     2641 aacgctggtg aaagtaaaag atgctgaaga tcagttgggt gcacgagtgg gttacatcga
     2701 actggatctc aacagcggta agatccttga gagttttcgc cccgaagaac gttttccaat
     2761 gatgagcact tttaaagttc tgctatgtgg cgcggtatta tcccgtattg acgccgggca
     2821 agagcaactc ggtcgccgca tacactattc tcagaatgac ttggttgagt actcaccagt
     2881 cacagaaaag catcttacgg atggcatgac agtaagagaa ttatgcagtg ctgccataac
     2941 catgagtgat aacactgcgg ccaacttact tctgacaacg atcggaggac cgaaggagct
     3001 aaccgctttt ttgcacaaca tgggggatca tgtaactcgc cttgatcgtt gggaaccgga
     3061 gctgaatgaa gccataccaa acgacgagcg tgacaccacg atgcctgtag caatggcaac
     3121 aacgttgcgc aaactattaa ctggcgaact acttactcta gcttcccggc aacaattaat
     3181 agactggatg gaggcggata aagttgcagg accacttctg cgctcggccc ttccggctgg
     3241 ctggtttatt gctgataaat ctggagccgg tgagcgtggg tctcgcggta tcattgcagc
     3301 actggggcca gatggtaagc cctcccgtat cgtagttatc tacacgacgg ggagtcaggc
     3361 aactatggat gaacgaaata gacagatcgc tgagataggt gcctcactga ttaagcattg
     3421 gtaactgtca gaccaagttt actcatatat actttagatt gatttaaaac ttcattttta
     3481 atttaaaagg atctaggtga agatcctttt tgataatctc atgaccaaaa tcccttaacg
     3541 tgagttttcg ttccactgag cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga
     3601 tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt
     3661 ggtttgtttg ccggatcaag agctaccaac tctttttccg aaggtaactg gcttcagcag
     3721 agcgcagata ccaaatactg ttcttctagt gtagccgtag ttaggccacc acttcaagaa
     3781 ctctgtagca ccgcctacat acctcgctct gctaatcctg ttaccagtgg ctgctgccag
     3841 tggcgataag tcgtgtctta ccgggttgga ctcaagacga tagttaccgg ataaggcgca
     3901 gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa cgacctacac
     3961 cgaactgaga tacctacagc gtgagctatg agaaagcgcc acgcttcccg aagggagaaa
     4021 ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga gggagcttcc
     4081 agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct gacttgagcg
     4141 tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaaaacgcca gcaacgcggc
     4201 ctttttacgg ttcctggcct tttgctggcc ttttgctcac atgttctttc ctgcgttatc
     4261 ccctgattct gtggataacc gtattaccgc ctttgagtga gctgataccg ctcgccgcag
     4321 ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg gaagagcgcc caatacgcaa
     4381 accgcctctc cccgcgcgtt ggccgattca ttaatgcag
//
