LOCUS       sequence-446222-        5489 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses Hisx6-Tagged msYFP2 in bacteria.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5489)
  TITLE     Yellow and oxidation-resistant derivatives of a monomeric 
            superfolder GFP
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 5489)
  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: 
            "Unpublished"
COMMENT     Sequence Label: pQE81XN-msYFP2
FEATURES             Location/Qualifiers
     source          1..5489
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(10..24,25..30,31..47,48..54)
                     /label=T5 promoter
                     /note="bacteriophage T5 promoter for E. coli RNA 
                     polymerase, with embedded lac operator"
     protein_bind    30..46
                     /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    62..78
                     /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             97..108
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             127..144
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     regulatory      160..169
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             166..873
                     /codon_start=1
                     /product="photostable monomeric superfolder GFP (Valbuena 
                     et al., 2020)"
                     /label=msGFP2
                     /translation="MDSTESLFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTLK
                     FICTTGKLPVPWPTLVTSLGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTITFKDDG
                     TYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIKA
                     NFKIRHNVEDGGVQLADHYQQNTPIGDGPVLLPDNHYLSYQSKLSKDPNEKRDHMVLLE
                     FVTAAGITGGSGGS"
     primer_bind     complement(208..229)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     816..837
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     terminator      946..1040
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             1084..1743
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(1150..1169)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     terminator      1808..1894
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     CDS             complement(1987..3069)
                     /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     3031..3050
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(3070..3147)
                     /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(3231..3253)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    3339..3479
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     3394..3413
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(3494..3511)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3665..4253)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3745..3764)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4424..5215,5216..5284))
                     /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     5047..5066
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5285..5389)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5457..5475
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 ctcgagaaat cataaaaaat ttatttgctt tgtgagcgga taacaattat aatagattca
       61 attgtgagcg gataacaatt tcacacagaa ttcattaaag aggagaaatt aactatgaga
      121 ggatctcacc atcaccatca ccatacggat ccactagtcg ccaccatgga tagcactgag
      181 agcctgttca ccggggtggt gcccatcctg gtcgagctgg acggcgacgt aaacggccac
      241 aagttcagcg tgcgcggcga gggcgagggc gatgccacca acggcaagct gaccctgaag
      301 ttcatctgca ccaccggcaa gctgcccgtg ccctggccca ccctcgtgac ctccctgggc
      361 tacggcctgc agtgcttcgc ccgctacccc gaccacatga agcagcacga cttcttcaag
      421 tccgccatgc ccgaaggcta cgtccaggag cgcaccatca ccttcaagga cgacggcacc
      481 tacaagaccc gcgccgaggt gaagttcgag ggcgacaccc tggtgaaccg catcgagctg
      541 aagggcatcg acttcaagga ggacggcaac atcctggggc acaagctgga gtacaacttc
      601 aacagccaca acgtctatat caccgccgac aagcagaaga acggcatcaa ggccaacttc
      661 aagatccgcc acaacgtgga ggacggcggc gtgcagctcg ccgaccacta ccagcagaac
      721 acccccatcg gcgacggccc cgtgctgctg cccgacaacc actacctgag ctaccagtcc
      781 aagctgagca aagaccccaa cgagaagcgc gatcacatgg tcctgctgga gttcgtgacc
      841 gccgccggga tcactggagg cagtggcggt tcttaaagcg gccgcgactc tagatcactc
      901 ggtaccccgg gtcgacctgc agccaagctt aattagctga gcttggactc ctgttgatag
      961 atccagtaat gacctcagaa ctccatctgg atttgttcag aacgctcggt tgccgccggg
     1021 cgttttttat tggtgagaat ccaagctagc ttggcgagat tttcaggagc taaggaagct
     1081 aaaatggaga aaaaaatcac tggatatacc accgttgata tatcccaatg gcatcgtaaa
     1141 gaacattttg aggcatttca gtcagttgct caatgtacct ataaccagac cgttcagctg
     1201 gatattacgg cctttttaaa gaccgtaaag aaaaataagc acaagtttta tccggccttt
     1261 attcacattc ttgcccgcct gatgaatgct catccggaat ttcgtatggc aatgaaagac
     1321 ggtgagctgg tgatatggga tagtgttcac ccttgttaca ccgttttcca tgagcaaact
     1381 gaaacgtttt catcgctctg gagtgaatac cacgacgatt tccggcagtt tctacacata
     1441 tattcgcaag atgtggcgtg ttacggtgaa aacctggcct atttccctaa agggtttatt
     1501 gagaatatgt ttttcgtctc agccaatccc tgggtgagtt tcaccagttt tgatttaaac
     1561 gtggccaata tggacaactt cttcgccccc gttttcacta tgggcaaata ttatacgcaa
     1621 ggcgacaagg tgctgatgcc gctggcgatt caggttcatc atgccgtttg tgatggcttc
     1681 catgtcggca gaatgcttaa tgaattacaa cagtactgcg atgagtggca gggcggggcg
     1741 taattttttt aaggcagtta ttggtgccct taaacgcctg gggtaatgac tctctagctt
     1801 gaggcatcaa ataaaacgaa aggctcagtc gaaagactgg gcctttcgtt ttatctgttg
     1861 tttgtcggtg aacgctctcc tgagtaggac aaatccgccc tctagattac gtgcagtcga
     1921 tgataagctg tcaaacatga gaattgtgcc taatgagtga gctaacttac attaattgcg
     1981 ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt gccagctgca ttaatgaatc
     2041 ggccaacgcg cggggagagg cggtttgcgt attgggcgcc agggtggttt ttcttttcac
     2101 cagtgagacg ggcaacagct gattgccctt caccgcctgg ccctgagaga gttgcagcaa
     2161 gcggtccacg ctggtttgcc ccagcaggcg aaaatcctgt ttgatggtgg ttaacggcgg
     2221 gatataacat gagctgtctt cggtatcgtc gtatcccact accgagatat ccgcaccaac
     2281 gcgcagcccg gactcggtaa tggcgcgcat tgcgcccagc gccatctgat cgttggcaac
     2341 cagcatcgca gtgggaacga tgccctcatt cagcatttgc atggtttgtt gaaaaccgga
     2401 catggcactc cagtcgcctt cccgttccgc tatcggctga atttgattgc gagtgagata
     2461 tttatgccag ccagccagac gcagacgcgc cgagacagaa cttaatgggc ccgctaacag
     2521 cgcgatttgc tggtgaccca atgcgaccag atgctccacg cccagtcgcg taccgtcttc
     2581 atgggagaaa ataatactgt tgatgggtgt ctggtcagag acatcaagaa ataacgccgg
     2641 aacattagtg caggcagctt ccacagcaat ggcatcctgg tcatccagcg gatagttaat
     2701 gatcagccca ctgacgcgtt gcgcgagaag attgtgcacc gccgctttac aggcttcgac
     2761 gccgcttcgt tctaccatcg acaccaccac gctggcaccc agttgatcgg cgcgagattt
     2821 aatcgccgcg acaatttgcg acggcgcgtg cagggccaga ctggaggtgg caacgccaat
     2881 cagcaacgac tgtttgcccg ccagttgttg tgccacgcgg ttgggaatgt aattcagctc
     2941 cgccatcgcc gcttccactt tttcccgcgt tttcgcagaa acgtggctgg cctggttcac
     3001 cacgcgggaa acggtctgat aagagacacc ggcatactct gcgacatcgt ataacgttac
     3061 tggtttcaca ttcaccaccc tgaattgact ctcttccggg cgctatcatg ccataccgcg
     3121 aaaggttttg caccattcga tggtgtcgga atttcgggca gcgttgggtc ctggccacgg
     3181 gtgcgcatga tctagagctg cctcgcgcgt ttcggtgatg acggtgaaaa cctctgacac
     3241 atgcagctcc cggagacggt cacagcttgt ctgtaagcgg atgccgggag cagacaagcc
     3301 cgtcagggcg cgtcagcggg tgttggcggg tgtcggggcg cagccatgac ccagtcacgt
     3361 agcgatagcg gagtgtatac tggcttaact atgcggcatc agagcagatt gtactgagag
     3421 tgcaccatat gcggtgtgaa ataccgcaca gatgcgtaag gagaaaatac cgcatcaggc
     3481 gctcttccgc ttcctcgctc actgactcgc tgcgctcggt cgttcggctg cggcgagcgg
     3541 tatcagctca ctcaaaggcg gtaatacggt tatccacaga atcaggggat aacgcaggaa
     3601 agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg
     3661 cgtttttcca taggctccgc ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga
     3721 ggtggcgaaa cccgacagga ctataaagat accaggcgtt tccccctgga agctccctcg
     3781 tgcgctctcc tgttccgacc ctgccgctta ccggatacct gtccgccttt ctcccttcgg
     3841 gaagcgtggc gctttctcat agctcacgct gtaggtatct cagttcggtg taggtcgttc
     3901 gctccaagct gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc gccttatccg
     3961 gtaactatcg tcttgagtcc aacccggtaa gacacgactt atcgccactg gcagcagcca
     4021 ctggtaacag gattagcaga gcgaggtatg taggcggtgc tacagagttc ttgaagtggt
     4081 ggcctaacta cggctacact agaaggacag tatttggtat ctgcgctctg ctgaagccag
     4141 ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa acaaaccacc gctggtagcg
     4201 gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct caagaagatc
     4261 ctttgatctt ttctacgggg tctgacgctc agtggaacga aaactcacgt taagggattt
     4321 tggtcatgag attatcaaaa aggatcttca cctagatcct tttaaattaa aaatgaagtt
     4381 ttaaatcaat ctaaagtata tatgagtaaa cttggtctga cagttaccaa tgcttaatca
     4441 gtgaggcacc tatctcagcg atctgtctat ttcgttcatc catagttgcc tgactccccg
     4501 tcgtgtagat aactacgata cgggagggct taccatctgg ccccagtgct gcaatgatac
     4561 cgcgagaccc acgctcaccg gctccagatt tatcagcaat aaaccagcca gccggaaggg
     4621 ccgagcgcag aagtggtcct gcaactttat ccgcctccat ccagtctatt aattgttgcc
     4681 gggaagctag agtaagtagt tcgccagtta atagtttgcg caacgttgtt gccattgcta
     4741 caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc ggttcccaac
     4801 gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc
     4861 ctccgatcgt tgtcagaagt aagttggccg cagtgttatc actcatggtt atggcagcac
     4921 tgcataattc tcttactgtc atgccatccg taagatgctt ttctgtgact ggtgagtact
     4981 caaccaagtc attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa
     5041 tacgggataa taccgcgcca catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt
     5101 cttcggggcg aaaactctca aggatcttac cgctgttgag atccagttcg atgtaaccca
     5161 ctcgtgcacc caactgatct tcagcatctt ttactttcac cagcgtttct gggtgagcaa
     5221 aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa tgttgaatac
     5281 tcatactctt cctttttcaa tattattgaa gcatttatca gggttattgt ctcatgagcg
     5341 gatacatatt tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc acatttcccc
     5401 gaaaagtgcc acctgacgtc taagaaacca ttattatcat gacattaacc tataaaaata
     5461 ggcgtatcac gaggcccttt cgtcttcac
//
