LOCUS       sequence-180919-        3580 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bacterial expression of a fluorescent protein, Citrine .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3580)
  AUTHORS   Griesbeck O, Baird GS, Campbell RE, Zacharias DA, Tsien RY
  TITLE     Reducing the environmental sensitivity of yellow fluorescent 
            protein. Mechanism and applications.
  JOURNAL   J Biol Chem. 2001 Aug 3;276(31):29188-94. Epub 2001 May 31.
  PUBMED    11387331
REFERENCE   2  (bases 1 to 3580)
  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: "J Biol 
            Chem."; date: "2001-08-3"; volume: "276(31)"; pages: "29188-94. Epub
            2001 May 31"
COMMENT     Sequence Label: pNCS Citrine
FEATURES             Location/Qualifiers
     source          1..3580
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     1..18
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     103..122
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        103..121
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     RBS             153..175
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             195..212
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             216..248
                     /codon_start=1
                     /product="leader peptide from bacteriophage T7 gene 10"
                     /label=T7 tag (gene 10 leader)
                     /note="promotes efficient translation in E. coli"
                     /translation="MASMTGGQQMG"
     CDS             join(252..260,261..275)
                     /codon_start=1
                     /product="Xpress(TM) epitope tag, including an enterokinase
                     recognition and cleavage site"
                     /label=Xpress(TM) tag
                     /translation="DLYDDDDK"
     CDS             join(282..284,285..287,288..998)
                     /codon_start=1
                     /product="enhanced variant of YFP (Heikal et al., 2001)"
                     /label=Citrine
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFGYGLMCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(327..348)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(588..607)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     935..956
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(1064..1082)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1078..1125
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      1222..1677
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1309..1328)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1519..1540
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        1703..1807
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1808..1876,1877..2668)
                     /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(2026..2045)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2839..3427
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3328..3347
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 agcgagtcag tgagcgagga agcggaagag cgcccaatac gcaaaccgcc tctccccgcg
       61 cgttggccga ttcattaatg caggatctcg atcccgcgaa attaatacga ctcactatag
      121 ggagaccaca acggtttccc tctagaaata attttgttta actttaagaa ggagatatac
      181 atatgcgggg ttctcatcat catcatcatc atggtatggc tagcatgact ggtggacagc
      241 aaatgggtcg ggatctgtac gacgatgacg ataaggatcc catggtgagc aagggcgagg
      301 agctgttcac cggggtggtg cccatcctgg tcgagctgga cggcgacgta aacggccaca
      361 agttcagcgt gtccggcgag ggcgagggcg atgccaccta cggcaagctg accctgaagt
      421 tcatctgcac caccggcaag ctgcccgtgc cctggcccac cctcgtgacc accttcggct
      481 acggcctgat gtgcttcgcc cgctaccccg accacatgaa gcagcacgac ttcttcaagt
      541 ccgccatgcc cgaaggctac gtccaggagc gcaccatctt cttcaaggac gacggcaact
      601 acaagacccg cgccgaggtg aagttcgagg gcgacaccct ggtgaaccgc atcgagctga
      661 agggcatcga cttcaaggag gacggcaaca tcctggggca caagctggag tacaactaca
      721 acagccacaa cgtctatatc atggccgaca agcagaagaa cggcatcaag gtgaacttca
      781 agatccgcca caacatcgag gacggcagcg tgcagctcgc cgaccactac cagcagaaca
      841 cccccatcgg cgacggcccc gtgctgctgc ccgacaacca ctacctgagc taccagtccg
      901 ccctgagcaa agaccccaac gagaagcgcg atcacatggt cctgctggag ttcgtgaccg
      961 ccgccgggat cactctcggc atggacgagc tatacaagta agaattcgaa gcttgatccg
     1021 gctgctaaca aagcccgaaa ggaagctgag ttggctgctg ccaccgctga gcaataacta
     1081 gcataacccc ttggggcctc taaacgggtc ttgaggggtt ttttgctgaa aggaggaact
     1141 atatccggat ctggcgtaat agcgaagagg cccgcaccga tcgcccttcc caacagttgc
     1201 gcagcctgaa tggcgaatgg gacgcgccct gtagcggcgc attaagcgcg gcgggtgtgg
     1261 tggttacgcg cagcgtgacc gctacacttg ccagcgccct agcgcccgct cctttcgctt
     1321 tcttcccttc ctttctcgcc acgttcgccg gctttccccg tcaagctcta aatcgggggc
     1381 tccctttagg gttccgattt agtgctttac ggcacctcga ccccaaaaaa cttgattagg
     1441 gtgatggttc acgtagtggg ccatcgccct gatagacggt ttttcgccct ttgacgttgg
     1501 agtccacgtt ctttaatagt ggactcttgt tccaaactgg aacaacactc aaccctatct
     1561 cggtctattc ttttgattta taagggattt tgccgatttc ggcctattgg ttaaaaaatg
     1621 agctgattta acaaaaattt aacgcgaatt ttaacaaaat attaacgctt acaatttagg
     1681 tggcactttt cggggaaatg tgcgcggaac ccctatttgt ttatttttct aaatacattc
     1741 aaatatgtat ccgctcatga gacaataacc ctgataaatg cttcaataat attgaaaaag
     1801 gaagagtatg agtattcaac atttccgtgt cgcccttatt cccttttttg cggcattttg
     1861 ccttcctgtt tttgctcacc cagaaacgct ggtgaaagta aaagatgctg aagatcagtt
     1921 gggtgcacga gtgggttaca tcgaactgga tctcaacagc ggtaagatcc ttgagagttt
     1981 tcgccccgaa gaacgttttc caatgatgag cacttttaaa gttctgctat gtggcgcggt
     2041 attatcccgt attgacgccg ggcaagagca actcggtcgc cgcatacact attctcagaa
     2101 tgacttggtt gagtactcac cagtcacaga aaagcatctt acggatggca tgacagtaag
     2161 agaattatgc agtgctgcca taaccatgag tgataacact gcggccaact tacttctgac
     2221 aacgatcgga ggaccgaagg agctaaccgc ttttttgcac aacatggggg atcatgtaac
     2281 tcgccttgat cgttgggaac cggagctgaa tgaagccata ccaaacgacg agcgtgacac
     2341 cacgatgcct gtagcaatgg caacaacgtt gcgcaaacta ttaactggcg aactacttac
     2401 tctagcttcc cggcaacaat taatagactg gatggaggcg gataaagttg caggaccact
     2461 tctgcgctcg gcccttccgg ctggctggtt tattgctgat aaatctggag ccggtgagcg
     2521 tgggtctcgc ggtatcattg cagcactggg gccagatggt aagccctccc gtatcgtagt
     2581 tatctacacg acggggagtc aggcaactat ggatgaacga aatagacaga tcgctgagat
     2641 aggtgcctca ctgattaagc attggtaact gtcagaccaa gtttactcat atatacttta
     2701 gattgattta aaacttcatt tttaatttaa aaggatctag gtgaagatcc tttttgataa
     2761 tctcatgacc aaaatccctt aacgtgagtt ttcgttccac tgagcgtcag accccgtaga
     2821 aaagatcaaa ggatcttctt gagatccttt ttttctgcgc gtaatctgct gcttgcaaac
     2881 aaaaaaacca ccgctaccag cggtggtttg tttgccggat caagagctac caactctttt
     2941 tccgaaggta actggcttca gcagagcgca gataccaaat actgttcttc tagtgtagcc
     3001 gtagttaggc caccacttca agaactctgt agcaccgcct acatacctcg ctctgctaat
     3061 cctgttacca gtggctgctg ccagtggcga taagtcgtgt cttaccgggt tggactcaag
     3121 acgatagtta ccggataagg cgcagcggtc gggctgaacg gggggttcgt gcacacagcc
     3181 cagcttggag cgaacgacct acaccgaact gagataccta cagcgtgagc tatgagaaag
     3241 cgccacgctt cccgaaggga gaaaggcgga caggtatccg gtaagcggca gggtcggaac
     3301 aggagagcgc acgagggagc ttccaggggg aaacgcctgg tatctttata gtcctgtcgg
     3361 gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct
     3421 atggaaaaac gccagcaacg cggccttttt acggttcctg gccttttgct ggccttttgc
     3481 tcacatgttc tttcctgcgt tatcccctga ttctgtggat aaccgtatta ccgcctttga
     3541 gtgagctgat accgctcgcc gcagccgaac gaccgagcgc
//
