LOCUS       sequence-231191-        6299 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  fluorescent protein expression with an N-terminal His-tag.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6299)
  AUTHORS   Holzapfel HY, Stern AD, Bouhaddou M, Anglin CM, Putur D, Comer S, 
            Birtwistle MR
  TITLE     Fluorescence Multiplexing with Spectral Imaging and Combinatorics.
  JOURNAL   ACS Comb Sci. 2018 Nov 12;20(11):653-659. doi: 
            10.1021/acscombsci.8b00101. Epub 2018 Oct 30.
  PUBMED    30339749
REFERENCE   2  (bases 1 to 6299)
  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: 
            "10.1021/acscombsci.8b00101"; journalName: "ACS Comb Sci"; date: 
            "2018-11-12- 12"; volume: "20"; issue: "11"; pages: "653-659"
COMMENT     Sequence Label: pQLinkHD_mTFP1-mVenus
FEATURES             Location/Qualifiers
     source          1..6299
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(36..50,51..56,57..73,74..80)
                     /label=T5 promoter
                     /note="bacteriophage T5 promoter for E. coli RNA 
                     polymerase, with embedded lac operator"
     protein_bind    56..72
                     /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)."
     protein_bind    88..104
                     /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)."
     RBS             123..134
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             147..167
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=7xHis
                     /translation="HHHHHHH"
     protein_bind    180..204
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             207..914
                     /codon_start=1
                     /product="monomeric teal (cyan) fluorescent protein (Ai et 
                     al., 2006)"
                     /label=mTFP1
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEETTMGVIKPDMKIKLKMEGNVNGHAFVIEGEGEGKPYDG
                     TNTINLEVKEGAPLPFSYDILTTAFAYGNRAFTKYPDDIPNYFKQSFPEGYSWERTMTF
                     EDKGIVKVKSDISMEEDSFIYEIHLKGENFPPNGPVMQKKTTGWDASTERMYVRDGVLK
                     GDVKHKLLLEGGGHHRVDFKTIYRAKKAVKLPDYHFVDHRIEILNHDKDYNKVTVYESA
                     VARNSTDGMDELYK"
     CDS             join(942..944,945..947,948..1460)
                     /codon_start=1
                     /product="N-terminal fragment of mVenus for use in 
                     bimolecular fluorescence complementation (BiFC) (Kodama and
                     Hu, 2010)"
                     /label=VN173
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KLICTTGKLPVPWPTLVTTLGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYITADKQKNGIK
                     ANFKIRHNIE"
     primer_bind     complement(987..1008)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1248..1267)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1595..1616
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     protein_bind    complement(1660..1684)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     terminator      1706..1800
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             1894..2553
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(1960..1979)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     terminator      2618..2704
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     CDS             complement(2797..3879)
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /label=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)."
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     3841..3860
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(3880..3957)
                     /gene="lacI (mutant)"
                     /label=lacIq promoter
                     /note="In the lacIq allele, a single base change in the 
                     promoter boosts expression of the lacI gene about 10-fold."
     primer_bind     complement(4041..4063)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    4149..4289
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     4204..4223
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(4304..4321)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4475..5063)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4555..4574)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5234..6025,6026..6094))
                     /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     5857..5876
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6095..6199)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6267..6285
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 ctcgagctta attaacaaca ccatttgtcg agaaatcata aaaaatttat ttgctttgtg
       61 agcggataac aattataata gattcaattg tgagcggata acaatttcac acagaattca
      121 ttaaagagga gaaattaact atgaaacatc accatcacca tcaccatggt gctgcaggta
      181 caagtttgta caaaaaagca ggcttcatgg tgagcaaggg cgaggagacc acaatgggcg
      241 taatcaagcc cgacatgaag atcaagctga agatggaggg caacgtgaat ggccacgcct
      301 tcgtgatcga gggcgagggc gagggcaagc cctacgacgg caccaacacc atcaacctgg
      361 aggtgaagga gggagccccc ctgcccttct cctacgacat tctgaccacc gcgttcgcct
      421 acggcaacag ggccttcacc aagtaccccg acgacatccc caactacttc aagcagtcct
      481 tccccgaggg ctactcttgg gagcgcacca tgaccttcga ggacaagggc atcgtgaagg
      541 tgaagtccga catctccatg gaggaggact ccttcatcta cgagatacac ctcaagggcg
      601 agaacttccc ccccaacggc cccgtgatgc agaagaagac caccggctgg gacgcctcca
      661 ccgagaggat gtacgtgcgc gacggcgtgc tgaagggcga cgtcaagcac aagctgctgc
      721 tggagggcgg cggccaccac cgcgttgact tcaagaccat ctacagggcc aagaaggcgg
      781 tgaagctgcc cgactatcac tttgtggacc accgcatcga gatcctgaac cacgacaagg
      841 actacaacaa ggtgaccgtt tacgagagcg ccgtggcccg caactccacc gacggcatgg
      901 acgagctgta caagaccggt gccggaggtg ggggcctagg aatggtgagc aagggcgagg
      961 agctgttcac cggggtggtg cccatcctgg tcgagctgga cggcgacgta aacggccaca
     1021 agttcagcgt gtccggcgag ggcgagggcg atgccaccta cggcaagctg accctgaagc
     1081 tgatctgcac caccggcaag ctgcccgtgc cctggcccac cctcgtgacc accctgggct
     1141 acggcctgca gtgcttcgcc cgctaccccg accacatgaa gcagcacgac ttcttcaagt
     1201 ccgccatgcc cgaaggctac gtccaggagc gcaccatctt cttcaaggac gacggcaact
     1261 acaagacccg cgccgaggtg aagttcgagg gcgacaccct ggtgaaccgc atcgagctga
     1321 agggcatcga cttcaaggag gacggcaaca tcctggggca caagctggag tacaactaca
     1381 acagccacaa cgtctatatc accgccgaca agcagaagaa cggcatcaag gccaacttca
     1441 agatccgcca caacatcgag gacggcggcg tgcagctcgc cgaccactac cagcagaaca
     1501 cccccatcgg cgacggcccc gtgctgctgc ccgacaacca ctacctgagc taccagtcca
     1561 agctgagcaa agaccccaac gagaagcgcg atcacatggt cctgctggag ttcgtgaccg
     1621 ccgccgggat cactctcggc atggacgagt aaaagcttga cccagctttc ttgtacaaag
     1681 tggtaagctt aattagctga gcttggactc ctgttgatag atccagtaat gacctcagaa
     1741 ctccatctgg atttgttcag aacgctcggt tgccgccggg cgttttttat tggtgagaat
     1801 ccaagtaaca acaccattta aatggagtgg ttacaaatgg agtggttaat taaggctagc
     1861 ttggcgagat tttcaggagc taaggaagct aaaatggaga aaaaaatcac tggatatacc
     1921 accgttgata tatcccaatg gcatcgtaaa gaacattttg aggcatttca gtcagttgct
     1981 caatgtacct ataaccagac cgttcagctg gatattacgg cctttttaaa gaccgtaaag
     2041 aaaaataagc acaagtttta tccggccttt attcacattc ttgcccgcct gatgaatgct
     2101 catccggaat ttcgtatggc aatgaaagac ggtgagctgg tgatatggga tagtgttcac
     2161 ccttgttaca ccgttttcca tgagcaaact gaaacgtttt catcgctctg gagtgaatac
     2221 cacgacgatt tccggcagtt tctacacata tattcgcaag atgtggcgtg ttacggtgaa
     2281 aacctggcct atttccctaa agggtttatt gagaatatgt ttttcgtctc agccaatccc
     2341 tgggtgagtt tcaccagttt tgatttaaac gtggccaata tggacaactt cttcgccccc
     2401 gttttcacca tgggcaaata ttatacgcaa ggcgacaagg tgctgatgcc gctggcgatt
     2461 caggttcatc atgccgtttg tgatggcttc catgtcggca gaatgcttaa tgaattacaa
     2521 cagtactgcg atgagtggca gggcggggcg taattttttt aaggcagtta ttggtgccct
     2581 taaacgcctg gggtaatgac tctctagctt gaggcatcaa ataaaacgaa aggctcagtc
     2641 gaaagactgg gcctttcgtt ttatctgttg tttgtcggtg aacgctctcc tgagtaggac
     2701 aaatccgccc tctagattac gtgcagtcga tgataagctg tcaaacatga gaattgtgcc
     2761 taatgagtga gctaacttac attaattgcg ttgcgctcac tgcccgcttt ccagtcggga
     2821 aacctgtcgt gccagctgca ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt
     2881 attgggcgcc agggtggttt ttcttttcac cagtgagacg ggcaacagct gattgccctt
     2941 caccgcctgg ccctgagaga gttgcagcaa gcggtccacg ctggtttgcc ccagcaggcg
     3001 aaaatcctgt ttgatggtgg ttaacggcgg gatataacat gagctgtctt cggtatcgtc
     3061 gtatcccact accgagatat ccgcaccaac gcgcagcccg gactcggtaa tggcgcgcat
     3121 tgcgcccagc gccatctgat cgttggcaac cagcatcgca gtgggaacga tgccctcatt
     3181 cagcatttgc atggtttgtt gaaaaccgga catggcactc cagtcgcctt cccgttccgc
     3241 tatcggctga atttgattgc gagtgagata tttatgccag ccagccagac gcagacgcgc
     3301 cgagacagaa cttaatgggc ccgctaacag cgcgatttgc tggtgaccca atgcgaccag
     3361 atgctccacg cccagtcgcg taccgtcttc atgggagaaa ataatactgt tgatgggtgt
     3421 ctggtcagag acatcaagaa ataacgccgg aacattagtg caggcagctt ccacagcaat
     3481 ggcatcctgg tcatccagcg gatagttaat gatcagccca ctgacgcgtt gcgcgagaag
     3541 attgtgcacc gccgctttac aggcttcgac gccgcttcgt tctaccatcg acaccaccac
     3601 gctggcaccc agttgatcgg cgcgagattt aatcgccgcg acaatttgcg acggcgcgtg
     3661 cagggccaga ctggaggtgg caacgccaat cagcaacgac tgtttgcccg ccagttgttg
     3721 tgccacgcgg ttgggaatgt aattcagctc cgccatcgcc gcttccactt tttcccgcgt
     3781 tttcgcagaa acgtggctgg cctggttcac cacgcgggaa acggtctgat aagagacacc
     3841 ggcatactct gcgacatcgt ataacgttac tggtttcaca ttcaccaccc tgaattgact
     3901 ctcttccggg cgctatcatg ccataccgcg aaaggttttg caccattcga tggtgtcgga
     3961 atttcgggca gcgttgggtc ctggccacgg gtgcgcatga tctagagctg cctcgcgcgt
     4021 ttcggtgatg acggtgaaaa cctctgacac atgcagctcc cggagacggt cacagcttgt
     4081 ctgtaagcgg atgccgggag cagacaagcc cgtcagggcg cgtcagcggg tgttggcggg
     4141 tgtcggggcg cagccatgac ccagtcacgt agcgatagcg gagtgtatac tggcttaact
     4201 atgcggcatc agagcagatt gtactgagag tgcaccacat gcggtgtgaa ataccgcaca
     4261 gatgcgtaag gagaaaatac cgcatcaggc gctcttccgc ttcctcgctc actgactcgc
     4321 tgcgctcggt cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt
     4381 tatccacaga atcaggggat aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg
     4441 ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca taggctccgc ccccctgacg
     4501 agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga ctataaagat
     4561 accaggcgtt tccccctgga agctccctcg tgcgctctcc tgttccgacc ctgccgctta
     4621 ccggatacct gtccgccttt ctcccttcgg gaagcgtggc gctttctcat agctcacgct
     4681 gtaggtatct cagttcggtg taggtcgttc gctccaagct gggctgtgtg cacgaacccc
     4741 ccgttcagcc cgaccgctgc gccttatccg gtaactatcg tcttgagtcc aacccggtaa
     4801 gacacgactt atcgccactg gcagcagcca ctggtaacag gattagcaga gcgaggtatg
     4861 taggcggtgc tacagagttc ttgaagtggt ggcctaacta cggctacact agaaggacag
     4921 tatttggtat ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt
     4981 gatccggcaa acaaaccacc gctggtagcg gtggtttttt tgtttgcaag cagcagatta
     5041 cgcgcagaaa aaaaggatct caagaagatc ctttgatctt ttctacgggg tctgacgctc
     5101 agtggaacga aaactcacgt taagggattt tggtcatgag attatcaaaa aggatcttca
     5161 cctagatcct tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa
     5221 cttggtctga cagttaccaa tgcttaatca gtgaggcacc tatctcagcg atctgtctat
     5281 ttcgttcatc catagttgcc tgactccccg tcgtgtagat aactacgata cgggagggct
     5341 taccatctgg ccccagtgct gcaatgatac cgcgagaccc acgctcaccg gctccagatt
     5401 tatcagcaat aaaccagcca gccggaaggg ccgagcgcag aagtggtcct gcaactttat
     5461 ccgcctccat ccagtctatt aattgttgcc gggaagctag agtaagtagt tcgccagtta
     5521 atagtttgcg caacgttgtt gccattgcta caggcatcgt ggtgtcacgc tcgtcgtttg
     5581 gtatggcttc attcagctcc ggttcccaac gatcaaggcg agttacatga tcccccatgt
     5641 tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt tgtcagaagt aagttggccg
     5701 cagtgttatc actcatggtt atggcagcac tgcataattc tcttactgtc atgccatccg
     5761 taagatgctt ttctgtgact ggtgagtact caaccaagtc attctgagaa tagtgtatgc
     5821 ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca catagcagaa
     5881 ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg aaaactctca aggatcttac
     5941 cgctgttgag atccagttcg atgtaaccca ctcgtgcacc caactgatct tcagcatctt
     6001 ttactttcac cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg
     6061 gaataagggc gacacggaaa tgttgaatac tcatactctt cctttttcaa tattattgaa
     6121 gcatttatca gggttattgt ctcatgagcg gatacatatt tgaatgtatt tagaaaaata
     6181 aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc acctgacgtc taagaaacca
     6241 ttattatcat gacattaacc tataaaaata ggcgtatcac gaggcccttt cgtcttcac
//
