LOCUS       sequence-343755-        7819 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of HaloTag7 at the nuclear lamina of the mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7819)
  AUTHORS   Frei MS, Tarnawski M, Roberti MJ, Koch B, Hiblot J, Johnsson K
  TITLE     Engineered HaloTag variants for fluorescence lifetime multiplexing.
  JOURNAL   Nat Methods. 2021 Dec 16. pii: 10.1038/s41592-021-01341-x. doi: 
            10.1038/s41592-021-01341-x.
  PUBMED    34916672
REFERENCE   2  (bases 1 to 7819)
  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: "Nat 
            Methods. 2021 Dec 16. pii: 10.1038/s41592-021-01341-x. doi: 
            10.1038/s41592-021-01341-x."
COMMENT     Sequence Label: pcDNA5/FRT/TO_HaloTag7_LamB1
FEATURES             Location/Qualifiers
     source          1..7819
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(join(624..1415,1416..1484))
                     /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     1247..1266
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1485..1589)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(1664..1683)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        1855..2234
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        2235..2438
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     2389..2409
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     protein_bind    2440..2458
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    2461..2479
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     2489..2513
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             2672..3556
                     /codon_start=1
                     /product="modified bacterial dehalogenase that forms 
                     covalent bonds with chloroalkane derivatives"
                     /label=HaloTag(R)
                     /translation="EIGTGFPFDPHYVEVLGERMHYVDVGPRDGTPVLFLHGNPTSSYV
                     WRNIIPHVAPTHRCIAPDLIGMGKSDKPDLGYFFDDHVRFMDAFIEALGLEEVVLVIHD
                     WGSALGFHWAKRNPERVKGIAFMEFIRPIPTWDEWPEFARETFQAFRTTDVGRKLIIDQ
                     NVFIEGTLPMGVVRPLTEVEMDHYREPFLNPVDREPLWRFPNELPIAGEPANIVALVEE
                     YMDWLHQSPVPKLLFWGTPGVLIPPAEAARLAKSLPNCKAVDIGPGLNLLQEDNPDLIG
                     SEIARWLSTLEISG"
     primer_bind     complement(5391..5408)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    5397..5621
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(5754..5773)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     protein_bind    5905..5952
                     /label=FRT
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="FLP-mediated recombination occurs in the 8-bp core 
                     sequence TCTAGAAA (Turan and Bode, 2011)."
     CDS             5960..6982
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     CDS             5960..6982
                     /codon_start=1
                     /gene="aph(4)-Ia"
                     /product="aminoglycoside phosphotransferase from E. coli"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     polyA_signal    7112..7233
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(7149..7168)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     7203..7222
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(7282..7298)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(7282..7298)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(7295..7317)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    7306..7322
                     /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)."
     promoter        complement(join(7330..7336,7337..7354,7355..7360))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    7375..7396
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(7513..7530)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(join(7684..7819,1..453))
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(7764..7783)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc gctttctcat
       61 agctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct gggctgtgtg
      121 cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg tcttgagtcc
      181 aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag gattagcaga
      241 gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta cggctacact
      301 agaagaacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt
      361 ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt tgtttgcaag
      421 cagcagatta cgcgcagaaa aaaaggatct caagaagatc ctttgatctt ttctacgggg
      481 tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgag attatcaaaa
      541 aggatcttca cctagatcct tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata
      601 tatgagtaaa cttggtctga cagttaccaa tgcttaatca gtgaggcacc tatctcagcg
      661 atctgtctat ttcgttcatc catagttgcc tgactccccg tcgtgtagat aactacgata
      721 cgggagggct taccatctgg ccccagtgct gcaatgatac cgcgagaccc acgctcaccg
      781 gctccagatt tatcagcaat aaaccagcca gccggaaggg ccgagcgcag aagtggtcct
      841 gcaactttat ccgcctccat ccagtctatt aattgttgcc gggaagctag agtaagtagt
      901 tcgccagtta atagtttgcg caacgttgtt gccattgcta caggcatcgt ggtgtcacgc
      961 tcgtcgtttg gtatggcttc attcagctcc ggttcccaac gatcaaggcg agttacatga
     1021 tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt tgtcagaagt
     1081 aagttggccg cagtgttatc actcatggtt atggcagcac tgcataattc tcttactgtc
     1141 atgccatccg taagatgctt ttctgtgact ggtgagtact caaccaagtc attctgagaa
     1201 tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca
     1261 catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg aaaactctca
     1321 aggatcttac cgctgttgag atccagttcg atgtaaccca ctcgtgcacc caactgatct
     1381 tcagcatctt ttactttcac cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc
     1441 gcaaaaaagg gaataagggc gacacggaaa tgttgaatac tcatactctt cctttttcaa
     1501 tattattgaa gcatttatca gggttattgt ctcatgagcg gatacatatt tgaatgtatt
     1561 tagaaaaata aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc acctgacgtc
     1621 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
     1681 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
     1741 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
     1801 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
     1861 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
     1921 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
     1981 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
     2041 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
     2101 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
     2161 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
     2221 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
     2281 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
     2341 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
     2401 gtaggcgtgt acggtgggag gtctatataa gcagagctct ccctatcagt gatagagatc
     2461 tccctatcag tgatagagat cgtcgacgag ctcgtttagt gaaccgtcag atcgcctgga
     2521 gacgccatcc acgctgtttt gacctccata gaagacaccg ggaccgatcc agcctccgga
     2581 ctctagcgtt taaacttaag cttggtaccg agctcggatc cactagtcca gtgtggtgga
     2641 attctgcaga tatccagcaa ccatgggatc cgaaatcggt actggctttc cattcgaccc
     2701 ccattatgtg gaagtcctgg gcgagcgcat gcactacgtc gatgttggtc cgcgcgatgg
     2761 cacccctgtg ctgttcctgc acggtaaccc gacctcctcc tacgtgtggc gcaacatcat
     2821 cccgcatgtt gcaccgaccc atcgctgcat tgctccagac ctgatcggta tgggcaaatc
     2881 cgacaaacca gacctgggtt atttcttcga cgaccacgtc cgcttcatgg atgccttcat
     2941 cgaagccctg ggtctggaag aggtcgtcct ggtcattcac gactggggct ccgctctggg
     3001 tttccactgg gccaagcgca atccagagcg cgtcaaaggt attgcattta tggagttcat
     3061 ccgccctatc ccgacctggg acgaatggcc agaatttgcc cgcgagacct tccaggcctt
     3121 ccgcaccacc gacgtcggcc gcaagctgat catcgatcag aacgttttta tcgagggtac
     3181 gctgccgatg ggtgtcgtcc gcccgctgac tgaagtcgag atggaccatt accgcgagcc
     3241 gttcctgaat cctgttgacc gcgagccact gtggcgcttc ccaaacgagc tgccaatcgc
     3301 cggtgagcca gcgaacatcg tcgcgctggt cgaagaatac atggactggc tgcaccagtc
     3361 ccctgtcccg aagctgctgt tctggggcac cccaggcgtt ctgatcccac cggccgaagc
     3421 cgctcgcctg gccaaaagcc tgcctaactg caaggctgtg gacatcggcc cgggtctgaa
     3481 tctgctgcaa gaagacaacc cggacctgat cggcagcgag atcgcgcgct ggctgtctac
     3541 tctggagatt tccggttccg gactcagatc tcgagctcaa gcttcgatgg cgactgcgac
     3601 ccccgtgccg ccgcggatgg gcagccgcgc tggcggcccc accacgccgc tgagccccac
     3661 gcgcctgtcg cggctccagg agaaggagga gctgcgcgag ctcaatgacc ggctggcggt
     3721 gtacatcgac aaggtgcgca gcctggagac ggagaacagc gcgctgcagc tgcaggtgac
     3781 ggagcgcgag gaggtgcgcg gccgtgagct caccggcctc aaggcgctct acgagaccga
     3841 gctggccgac gcgcgacgcg cgctcgacga cacggcccgc gagcgcgcca agctgcagat
     3901 cgagctgggc aagtgcaagg cggaacacga ccagctgctc ctcaactatg ctaagaagga
     3961 atctgatctt aatggcgccc agatcaagct tcgagaatat gaagcagcac tcaattcgaa
     4021 agatgcagct cttgctactg cacttggtga caaaaaaagt ttagagggag atttggagga
     4081 tctgaaggat cagattgccc agttggaagc ctccttagct gcagccaaaa aacagttagc
     4141 agatgaaact ttacttaaag tagatttgga gaatcgttgt cagagcctta ctgaggactt
     4201 ggagtttcgc aaaagcatgt atgaagagga gattaacgag accagaagga agcatgaaac
     4261 gcgcttggta gaggtggatt ctgggcgtca aattgagtat gagtacaagc tggcgcaagc
     4321 ccttcatgag atgagagagc aacatgatgc ccaagtgagg ctgtataagg aggagctgga
     4381 gcagacttac catgccaaac ttgagaatgc cagactgtca tcagagatga atacttctac
     4441 tgtcaacagt gccagggaag aactgatgga aagccgcatg agaattgaga gcctttcatc
     4501 ccagctttct aatctacaga aagagtctag agcatgtttg gaaaggattc aagaattaga
     4561 ggacttgctt gctaaagaaa aagacaactc tcgtcgcatg ctgacagaca aagagagaga
     4621 gatggcggaa ataagggatc aaatgcagca acagctgaat gactatgaac agcttcttga
     4681 tgtaaagtta gccctggaca tggaaatcag tgcttacagg aaactcttag aaggcgaaga
     4741 agagaggttg aagctgtctc caagcccttc ttcccgtgtg acagtatccc gagcatcctc
     4801 aagtcgtagt gtacgtacaa ctagaggaaa gcggaagagg gttgatgtgg aagaatcaga
     4861 ggcgagtagt agtgttagca tctctcattc cgcctcagcc actggaaatg tttgcatcga
     4921 agaaattgat gttgatggga aatttatccg cttgaagaac acttctgaac aggatcaacc
     4981 aatgggaggc tgggagatga tcagaaaaat tggagacaca tcagtcagtt ataaatatac
     5041 ctcaagatat gtgctgaagg caggccagac tgttacaatt tgggctgcaa acgctggtgt
     5101 cacagccagc cccccaactg acctcatctg gaagaaccag aactcgtggg gcactggcga
     5161 agatgtgaag gttatattga aaaattctca gggagaggag gttgctcaaa gaagtacagt
     5221 ctttaaaaca accatacctg aagaagagga ggaggaggaa gaagcagctg gagtggttgt
     5281 tgaggaagaa cttttccacc agcagggaac cccaagagca tccaatagaa gctgtgcaat
     5341 tatgtagtaa ccgctcgagt ctagagggcc cgtttaaacc cgctgatcag cctcgactgt
     5401 gccttctagt tgccagccat ctgttgtttg cccctccccc gtgccttcct tgaccctgga
     5461 aggtgccact cccactgtcc tttcctaata aaatgaggaa attgcatcgc attgtctgag
     5521 taggtgtcat tctattctgg ggggtggggt ggggcaggac agcaaggggg aggattggga
     5581 agacaatagc aggcatgctg gggatgcggt gggctctatg gcttctgagg cggaaagaac
     5641 cagctggggc tctagggggt atccccacgc gccctgtagc ggcgcattaa gcgcggcggg
     5701 tgtggtggtt acgcgcagcg tgaccgctac acttgccagc gccctagcgc ccgctccttt
     5761 cgctttcttc ccttcctttc tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg
     5821 ggggctccct ttagggttcc gatttagtgc tttacggcac ctcgacccca aaaaacttga
     5881 ttagggtgat ggttcacgta cctagaagtt cctattccga agttcctatt ctctagaaag
     5941 tataggaact tccttggcca aaaagcctga actcaccgcg acgtctgtcg agaagtttct
     6001 gatcgaaaag ttcgacagcg tctccgacct gatgcagctc tcggagggcg aagaatctcg
     6061 tgctttcagc ttcgatgtag gagggcgtgg atatgtcctg cgggtaaata gctgcgccga
     6121 tggtttctac aaagatcgtt atgtttatcg gcactttgca tcggccgcgc tcccgattcc
     6181 ggaagtgctt gacattgggg aattcagcga gagcctgacc tattgcatct cccgccgtgc
     6241 acagggtgtc acgttgcaag acctgcctga aaccgaactg cccgctgttc tgcagccggt
     6301 cgcggaggcc atggatgcga tcgctgcggc cgatcttagc cagacgagcg ggttcggccc
     6361 attcggaccg caaggaatcg gtcaatacac tacatggcgt gatttcatat gcgcgattgc
     6421 tgatccccat gtgtatcact ggcaaactgt gatggacgac accgtcagtg cgtccgtcgc
     6481 gcaggctctc gatgagctga tgctttgggc cgaggactgc cccgaagtcc ggcacctcgt
     6541 gcacgcggat ttcggctcca acaatgtcct gacggacaat ggccgcataa cagcggtcat
     6601 tgactggagc gaggcgatgt tcggggattc ccaatacgag gtcgccaaca tcttcttctg
     6661 gaggccgtgg ttggcttgta tggagcagca gacgcgctac ttcgagcgga ggcatccgga
     6721 gcttgcagga tcgccgcggc tccgggcgta tatgctccgc attggtcttg accaactcta
     6781 tcagagcttg gttgacggca atttcgatga tgcagcttgg gcgcagggtc gatgcgacgc
     6841 aatcgtccga tccggagccg ggactgtcgg gcgtacacaa atcgcccgca gaagcgcggc
     6901 cgtctggacc gatggctgtg tagaagtact cgccgatagt ggaaaccgac gccccagcac
     6961 tcgtccgagg gcaaaggaat agcacgtact acgagatttc gattccaccg ccgccttcta
     7021 tgaaaggttg ggcttcggaa tcgttttccg ggacgccggc tggatgatcc tccagcgcgg
     7081 ggatctcatg ctggagttct tcgcccaccc caacttgttt attgcagctt ataatggtta
     7141 caaataaagc aatagcatca caaatttcac aaataaagca tttttttcac tgcattctag
     7201 ttgtggtttg tccaaactca tcaatgtatc ttatcatgtc tgtataccgt cgacctctag
     7261 ctagagcttg gcgtaatcat ggtcatagct gtttcctgtg tgaaattgtt atccgctcac
     7321 aattccacac aacatacgag ccggaagcat aaagtgtaaa gcctggggtg cctaatgagt
     7381 gagctaactc acattaattg cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc
     7441 gtgccagctg cattaatgaa tcggccaacg cgcggggaga ggcggtttgc gtattgggcg
     7501 ctcttccgct tcctcgctca ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt
     7561 atcagctcac tcaaaggcgg taatacggtt atccacagaa tcaggggata acgcaggaaa
     7621 gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc
     7681 gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct caagtcagag
     7741 gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa gctccctcgt
     7801 gcgctctcct gttccgacc
//
