LOCUS       sequence-233933-        5807 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses mKelly1 tagged N-terminally with an MHHHHHHGSG tag.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5807)
  AUTHORS   Wannier TM, Gillespie SK, Hutchins N, McIsaac RS, Wu SY, Shen Y, 
            Campbell RE, Brown KS, Mayo SL
  TITLE     Monomerization of far-red fluorescent proteins.
  JOURNAL   Proc Natl Acad Sci U S A. 2018 Nov 27;115(48):E11294-E11301. doi: 
            10.1073/pnas.1807449115. Epub 2018 Nov 13.
  PUBMED    30425172
REFERENCE   2  (bases 1 to 5807)
  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: 
            "10.1073/pnas.1807449115"; journalName: "Proc Natl Acad Sci U S A"; 
            date: "2018-11-27- 27"; volume: "115"; issue: "48"; pages: 
            "E11294-E11301"
COMMENT     Sequence Label: pET-mKelly1
FEATURES             Location/Qualifiers
     source          1..5807
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     550..569
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(589..666)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     872..891
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     979..998
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        979..997
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    998..1022
                     /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             1037..1059
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             1069..1086
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             1783..1806
                     /codon_start=1
                     /product="peptide that binds Strep-Tactin(R), an engineered
                     form of streptavidin"
                     /label=Strep-Tag II
                     /translation="WSHPQFEK"
     primer_bind     complement(1833..1851)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1847..1894
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      1931..2386
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2018..2037)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2228..2249
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(join(2501..3292,3293..3361))
                     /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     3124..3143
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3362..3454)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      3535..4123
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4024..4043
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4277..4294
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    4309..4448
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(4374..4393)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4534..4556
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(4550..4741)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(join(5313..5807,1..588))
                     /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
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEYSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
ORIGIN
        1 cagacgcgcc gagacagaac ttaatgggcc cgctaacagc gcgatttgct ggtgacccaa
       61 tgcgaccaga tgctccacgc ccagtcgcgt accgtcttca tgggagaaaa taatactgtt
      121 gatgggtgtc tggtcagaga catcaagaaa taacgccgga acattagtgc aggcagcttc
      181 cacagcaatg gcatcctggt catccagcgg atagttaatg atcagcccac tgacgcgttg
      241 cgcgagaaga ttgtgcaccg ccgctttaca ggcttcgacg ccgcttcgtt ctaccatcga
      301 caccaccacg ctggcaccca gttgatcggc gcgagattta atcgccgcga caatttgcga
      361 cggcgcgtgc agggccagac tggaggtggc aacgccaatc agcaacgact gtttgcccgc
      421 cagttgttgt gccacgcggt tgggaatgta attcagctcc gccatcgccg cttccacttt
      481 ttcccgcgtt ttcgcagaaa cgtggctggc ctggttcacc acgcgggaaa cggtctgata
      541 agagacaccg gcatactctg cgacatcgta taacgttact ggtttcacat tcaccaccct
      601 gaattgactc tcttccgggc gctatcatgc cataccgcga aaggttttgc gccattcgat
      661 ggtgtccggg atctcgacgc tctcccttat gcgactcctg cattaggaag cagcccagta
      721 gtaggttgag gccgttgagc accgccaccg caaggaatgg tgcatgcaag gagatggcgc
      781 ccaacagtcc cccggccacg gggcctgcca ccatacccac gccgaaacaa gcgctcatga
      841 gcccgaagtg gcgagcccga tcttccccat aggtgatgtc ggcgatatag gcgccagcaa
      901 ccgcacctgt ggcgccggtg atgccggcca cgatgcgtcc ggcgtagagg atcgagatcg
      961 atctcgatcc cgcgaaatta atacgactca ctatagggga attgtgagcg gataacaatt
     1021 cccctctaga aataattttg tttaacttta agaaggagat ataccatgca ccatcatcac
     1081 caccacggtt ctggtgaact gattaaagaa aacatgcgta tgaaactgta catggaaggt
     1141 accgttggca accatcactt caaatgcacc gcggaaggtg agggtaaacc gtacgaaggc
     1201 acccagaccc agcgtattaa agttgttgaa ggtggtccgc tgccgttcgc gttcgacatc
     1261 ctggcgacct gcttcatgta tggttctaaa acttttatca accaccctca gggtatcccg
     1321 gacttcttca aacagtcttt cccagaaggt ttcacctggg aacgtgttac cacctacgaa
     1381 gacggcggtg ttctgaccgt tacccaggac acctctctgc aggacggtta cctgatctac
     1441 aacgttaaac tgcgtggtgt taacttcccg tctaacggcc ctgttatgca gaaaaagact
     1501 ctgggttggg aaccgaccac cgagacactg gtcccggttg acggtggcct ggtgggtaga
     1561 tgtgatatgg cgctgcgtct ggttggtggc ggtcacctga agtgcaatct gaaaaccacg
     1621 tatcgttcca aaaaaccagc gaaaaacctg aagatgccgg gtaagtacca ggttgaccgt
     1681 aagctggagc gtatcaaaga ggcggacaac gaaacctacg ttgaacagca cgaagttgcg
     1741 gttgcgagat aatgagcttt cttgtacaaa gtggtttccg cttggagcca cccgcagttc
     1801 gaaaagtaag tgattaacct aggctgctgc caccgctgag caataactag cataacccct
     1861 tggggcctct aaacgggtct tgaggggttt tttgctgaaa ggaggaacta tatccggatt
     1921 ggcgaatggg acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc
     1981 agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc ctttcgcttt cttcccttcc
     2041 tttctcgcca cgttcgccgg ctttccccgt caagctctaa atcgggggct ccctttaggg
     2101 ttccgattta gtgctttacg gcacctcgac cccaaaaaac ttgattaggg tgatggttca
     2161 cgtagtgggc catcgccctg atagacggtt tttcgccctt tgacgttgga gtccacgttc
     2221 tttaatagtg gactcttgtt ccaaactgga acaacactca accctatctc ggtctattct
     2281 tttgatttat aagggatttt gccgatttcg gcctattggt taaaaaatga gctgatttaa
     2341 caaaaattta acgcgaattt taacaaaata ttaacgttta caatttctgg cggcacgatg
     2401 gcatgagatt atcaaaaagg atcttcacct agatcctttt aaattaaaaa tgaagtttta
     2461 aatcaatcta aagtatatat gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg
     2521 aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga ctccccgtcg
     2581 tgtagataac tacgatacgg gagggcttac catctggccc cagtgctgca atgataccgc
     2641 gagacccacg ctcaccggct ccagatttat cagcaataaa ccagccagcc ggaagggccg
     2701 agcgcagaag tggtcctgca actttatccg cctccatcca gtctattaat tgttgccggg
     2761 aagctagagt aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc attgctacag
     2821 gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt cagctccggt tcccaacgat
     2881 caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc
     2941 cgatcgttgt cagaagtaag ttggccgcag tgttatcact catggttatg gcagcactgc
     3001 ataattctct tactgtcatg ccatccgtaa gatgcttttc tgtgactggt gagtactcaa
     3061 ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac
     3121 gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga aaacgttctt
     3181 cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg taacccactc
     3241 gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg tgagcaaaaa
     3301 caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt tgaatactca
     3361 tactcttcct ttttcaatca tgattgaagc atttatcagg gttattgtct catgagcgga
     3421 tacatatttg aatgtattta gaaaaataaa caaataggtc atgaccaaaa tcccttaacg
     3481 tgagttttcg ttccactgag cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga
     3541 tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt
     3601 ggtttgtttg ccggatcaag agctaccaac tctttttccg aaggtaactg gcttcagcag
     3661 agcgcagata ccaaatactg tccttctagt gtagccgtag ttaggccacc acttcaagaa
     3721 ctctgtagca ccgcctacat acctcgctct gctaatcctg ttaccagtgg ctgctgccag
     3781 tggcgataag tcgtgtctta ccgggttgga ctcaagacga tagttaccgg ataaggcgca
     3841 gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa cgacctacac
     3901 cgaactgaga tacctacagc gtgagctatg agaaagcgcc acgcttcccg aagggagaaa
     3961 ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga gggagcttcc
     4021 agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct gacttgagcg
     4081 tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaaaacgcca gcaacgcggc
     4141 ctttttacgg ttcctggcct tttgctggcc ttttgctcac atgttctttc ctgcgttatc
     4201 ccctgattct gtggataacc gtattaccgc ctttgagtga gctgataccg ctcgccgcag
     4261 ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg gaagagcgcc tgatgcggta
     4321 ttttctcctt acgcatctgt gcggtatttc acaccgcaat ggtgcactct cagtacaatc
     4381 tgctctgatg ccgcatagtt aagccagtat acactccgct atcgctacgt gactgggtca
     4441 tggctgcgcc ccgacacccg ccaacacccg ctgacgcgcc ctgacgggct tgtctgctcc
     4501 cggcatccgc ttacagacaa gctgtgaccg tctccgggag ctgcatgtgt cagaggtttt
     4561 caccgtcatc accgaaacgc gcgaggcagc tgcggtaaag ctcatcagcg tggtcgtgaa
     4621 gcgattcaca gatgtctgcc tgttcatccg cgtccagctc gttgagtttc tccagaagcg
     4681 ttaatgtctg gcttctgata aagcgggcca tgttaagggc ggttttttcc tgtttggtca
     4741 ctgatgcctc cgtgtaaggg ggatttctgt tcatgggggt aatgataccg atgaaacgag
     4801 agaggatgct cacgatacgg gttactgatg atgaacatgc ccggttactg gaacgttgtg
     4861 agggtaaaca actggcggta tggatgcggc gggaccagag aaaaatcact cagggtcaat
     4921 gccagcgctt cgttaataca gatgtaggtg ttccacaggg tagccagcag catcctgcga
     4981 tgcagatccg gaacataatg gtgcagggcg ctgacttccg cgtttccaga ctttacgaaa
     5041 cacggaaacc gaagaccatt catgttgttg ctcaggtcgc agacgttttg cagcagcagt
     5101 cgcttcacgt tcgctcgcgt atcggtgatt cattctgcta accagtaagg caaccccgcc
     5161 agcctagccg ggtcctcaac gacaggagca cgatcatgct agtcatgccc cgcgcccacc
     5221 ggaaggagct gactgggttg aaggctctca agggcatcgg tcgagatccc ggtgcctaat
     5281 gagtgagcta acttacatta attgcgttgc gctcactgcc cgctttccag tcgggaaacc
     5341 tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg
     5401 ggcgccaggg tggtttttct tttcaccagt gagacgggca acagctgatt gcccttcacc
     5461 gcctggccct gagagagttg cagcaagcgg tccacgctgg tttgccccag caggcgaaaa
     5521 tcctgtttga tggtggttaa cggcgggata taacatgagc tgtattcggt atcgtcgtat
     5581 cccactaccg agatgtccgc accaacgcgc agcccggact cggtaatggc gcgcattgcg
     5641 cccagcgcca tctgatcgtt ggcaaccagc atcgcagtgg gaacgatgcc ctcattcagc
     5701 atttgcatgg tttgttgaaa accggacatg gcactccagt cgccttcccg ttccgctatc
     5761 ggctgaattt gattgcgagt gagatattta tgccagccag ccagacg
//
