LOCUS       sequence-418214-        7475 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of ProDOL probe - LynAnchor-eGFP-SNAP-tag-HaloTag.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7475)
  AUTHORS   Tashev SA, Euchner J, Yserentant K, Hanselmann S, Hild F, 
            Chmielewicz W, Hummert J, Schworer F, Tsopoulidis N, Germer S, 
            Sassmannshausen Z, Fackler OT, Klingmuller U, Herten DP
  TITLE     ProDOL: a general method to determine the degree of labeling for 
            staining optimization and molecular counting.
  JOURNAL   Nat Methods. 2024 Sep;21(9):1708-1715. doi: 
            10.1038/s41592-024-02376-6. Epub 2024 Aug 8.
  PUBMED    39117875
REFERENCE   2  (bases 1 to 7475)
  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: 
            "10.1038/s41592-024-02376-6"; journalName: "Nat Methods"; date: 
            "2024-09"; volume: "21"; issue: "9"; pages: "1708-1715"
COMMENT     Sequence Label: pBABE-ProDOL
FEATURES             Location/Qualifiers
     source          1..7475
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(join(496..1287,1288..1356))
                     /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     1119..1138
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1357..1461)
                     /gene="bla"
                     /label=AmpR promoter
     LTR             1492..1961
                     /note="long terminal repeat from Moloney murine leukemia 
                     virus"
     misc_feature    2026..2225
                     /label=MMLV Psi
                     /note="packaging signal of Moloney murine leukemia virus 
                     (MMLV)"
     primer_bind     2772..2794
                     /label=pLXSN 5'
                     /note="Murine stem cell virus, forward primer. Also called 
                     MSCV"
     CDS             join(2990..2992,2993..2995,2996..3706)
                     /codon_start=1
                     /product="enhanced GFP with monomerizing A206K mutation 
                     (Zacharias et al., 2002)"
                     /label=mEGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(3035..3056)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(3296..3315)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3643..3664
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             3713..4600
                     /codon_start=1
                     /product="modified bacterial dehalogenase that forms 
                     covalent bonds with chloroalkane derivatives"
                     /label=HaloTag(R)
                     /translation="AEIGTGFPFDPHYVEVLGERMHYVDVGPRDGTPVLFLHGNPTSSY
                     VWRNIIPHVAPTHRCIAPDLIGMGKSDKPDLGYFFDDHVRFMDAFIEALGLEEVVLVIH
                     DWGSALGFHWAKRNPERVKGIAFMEFIRPIPTWDEWPEFARETFQAFRTTDVGRKLIID
                     QNVFIEGTLPMGVVRPLTEVEMDHYREPFLNPVDREPLWRFPNELPIAGEPANIVALVE
                     EYMDWLHQSPVPKLLFWGTPGVLIPPAEAARLAKSLPNCKAVDIGPGLNLLQEDNPDLI
                     GSEIARWLSTLEISG"
     CDS             4685..5230
                     /codon_start=1
                     /product="human O6-alkylguanine-DNA-alkyltransferase 
                     variant that forms covalent bonds with benzylguanine 
                     derivatives"
                     /label=SNAP-tag(R)
                     /note="fast-reacting variant, optimized for mammalian 
                     expression"
                     /translation="MDKDCEMKRTTLDSPLGKLELSGCEQGLHRIIFLGKGTSAADAVE
                     VPAPAAVLGGPEPLMQATAWLNAYFHQPEAIEEFPVPALHHPVFQQESFTRQVLWKLLK
                     VVKFGEVISYSHLAALAGNPAATAAVKTALSGNPVPILIPCHRVVQGDLDVGGYEGGLA
                     VKEWLLAHEGHRLGKPGLG"
     CDS             5237..5254
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(5289..5309)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        5294..5623
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      5474..5609
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             5633..6232
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(5633..5652)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     6129..6149
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     LTR             6453..6922
                     /note="long terminal repeat from Moloney murine leukemia 
                     virus"
     primer_bind     complement(7052..7069)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(join(7223..7475,1..336))
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(7303..7322)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag
       61 tccaacccgg taagacacga cttatcgcca ctggcagcag ccactggtaa caggattagc
      121 agagcgaggt atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa ctacggctac
      181 actagaagaa cagtatttgg tatctgcgct ctgctgaagc cagttacctt cggaaaaaga
      241 gttggtagct cttgatccgg caaacaaacc accgctggta gcggtggttt ttttgtttgc
      301 aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag atcctttgat cttttctacg
      361 gggtctgacg ctcagtggaa cgaaaactca cgttaaggga ttttggtcat gagattatca
      421 aaaaggatct tcacctagat ccttttgcgg ccgcaaatca atctaaagta tatatgagta
      481 aacttggtct gacagttacc aatgcttaat cagtgaggca cctatctcag cgatctgtct
      541 atttcgttca tccatagttg cctgactccc cgtcgtgtag ataactacga tacgggaggg
      601 cttaccatct ggccccagtg ctgcaatgat accgcgagac ccacgctcac cggctccaga
      661 tttatcagca ataaaccagc cagccggaag ggccgagcgc agaagtggtc ctgcaacttt
      721 atccgcctcc atccagtcta ttaattgttg ccgggaagct agagtaagta gttcgccagt
      781 taatagtttg cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt
      841 tggtatggct tcattcagct ccggttccca acgatcaagg cgagttacat gatcccccat
      901 gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc gttgtcagaa gtaagttggc
      961 cgcagtgtta tcactcatgg ttatggcagc actgcataat tctcttactg tcatgccatc
     1021 cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag tcattctgag aatagtgtat
     1081 gcggcgaccg agttgctctt gcccggcgtc aatacgggat aataccgcgc cacatagcag
     1141 aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct caaggatctt
     1201 accgctgttg agatccagtt cgatgtaacc cactcgtgca cccaactgat cttcagcatc
     1261 ttttactttc accagcgttt ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa
     1321 gggaataagg gcgacacgga aatgttgaat actcatactc ttcctttttc aatattattg
     1381 aagcatttat cagggttatt gtctcatgag cggatacata tttgaatgta tttagaaaaa
     1441 taaacaaata ggggttccgc gcacatttcc ccgaaaagtg ccacctgcag cctgaatatg
     1501 ggccaaacag gatatctgtg gtaagcagtt cctgccccgg ctcagggcca agaacagatg
     1561 gaacagctga atatgggcca aacaggatat ctgtggtaag cagttcctgc cccggctcag
     1621 ggccaagaac agatggtccc cagatgcggt ccagccctca gcagtttcta gagaaccatc
     1681 agatgtttcc agggtgcccc aaggacctga aatgaccctg tgccttattt gaactaacca
     1741 atcagttcgc ttctcgcttc tgttcgcgcg cttctgctcc ccgagctcaa taaaagagcc
     1801 cacaacccct cactcggggc gccagtcctc cgattgactg agtcgcccgg gtacccgtgt
     1861 atccaataaa ccctcttgca gttgcatccg acttgtggtc tcgctgttcc ttgggagggt
     1921 ctcctctgag tgattgacta cccgtcagcg ggggtctttc atttgggggc tcgtccggga
     1981 tcgggagacc cctgcccagg gaccaccgac ccaccaccgg gaggcaagct ggccagcaac
     2041 ttatctgtgt ctgtccgatt gtctagtgtc tatgactgat tttatgcgcc tgcgtcggta
     2101 ctagttagct aactagctct gtatctggcg gacccgtggt ggaactgacg agttctgaac
     2161 acccggccgc aaccctggga gacgtcccag ggactttggg ggccgttttt gtggcccgac
     2221 ctgaggaagg gagtcgatgt ggaatccgac cccgtcagga tatgtggttc tggtaggaga
     2281 cgagaaccta aaacagttcc cgcctccgtc tgaatttttg ctttcggttt ggaaccgaag
     2341 ccgcgcgtct tgtctgctgc agcgctgcag catcgttctg tgttgtctct gtctgactgt
     2401 gtttctgtat ttgtctgaaa attagggcca gactgttacc actcccttaa gtttgacctt
     2461 aggtcactgg aaagatgtcg agcggatcgc tcacaaccag tcggtagatg tcaagaagag
     2521 acgttgggtt accttctgct ctgcagaatg gccaaccttt aacgtcggat ggccgcgaga
     2581 cggcaccttt aaccgagacc tcatcaccca ggttaagatc aaggtctttt cacctggccc
     2641 gcatggacac ccagaccagg tcccctacat cgtgacctgg gaagccttgg cttttgaccc
     2701 ccctccctgg gtcaagccct ttgtacaccc taagcctccg cctcctcttc ctccatccgc
     2761 cccgtctctc ccccttgaac ctcctcgttc gaccccccct cgagctgcag agcttgctga
     2821 cccagctgtg gactagggga aaggggaaag aggaaaggga aggagaggtg ggggggacga
     2881 aacaggggcg ctggagccct gagcttcctg aagctagggc tgcatcgaat tcatgggatg
     2941 tatcaagagt aagcgtaagg ataatctcaa tgacgacgag ctcgagacca tggtgagcaa
     3001 gggcgaggag ctgttcaccg gggtggtgcc catcctggtc gagctggacg gcgacgtaaa
     3061 cggccacaag ttcagcgtgt ccggcgaggg cgagggcgat gccacctacg gcaagctgac
     3121 cctgaagttc atctgcacca ccggcaagct gcccgtgccc tggcccaccc tcgtgaccac
     3181 cctgacctac ggcgtgcagt gcttcagccg ctaccccgac cacatgaagc agcacgactt
     3241 cttcaagtcc gccatgcccg aaggctacgt ccaggagcgc accatcttct tcaaggacga
     3301 cggcaactac aagacccgcg ccgaggtgaa gttcgagggc gacaccctgg tgaaccgcat
     3361 cgagctgaag ggcatcgact tcaaggagga cggcaacatc ctggggcaca agctggagta
     3421 caactacaac agccacaacg tctatatcat ggccgacaag cagaagaacg gcatcaaggt
     3481 gaacttcaag atccgccaca acatcgagga cggcagcgtg cagctcgccg accactacca
     3541 gcagaacacc cccatcggcg acggccccgt gctgctgccc gacaaccact acctgagcac
     3601 ccagtccaaa ctgagcaaag accccaacga gaagcgcgat cacatggtcc tgctggagtt
     3661 cgtgaccgcc gccgggatca ctctcggcat ggacgagctg tacaagggat ccgcagaaat
     3721 cggtactggc tttccattcg acccccatta tgtggaagtc ctgggcgagc gcatgcacta
     3781 cgtcgatgtt ggtccgcgcg atggcacccc tgtgctgttc ctgcacggta acccgacctc
     3841 ctcctacgtg tggcgcaaca tcatcccgca tgttgcaccg acccatcgct gcattgctcc
     3901 agacctgatc ggtatgggca aatccgacaa accagacctg ggttatttct tcgacgacca
     3961 cgtccgcttc atggatgcct tcatcgaagc cctgggtctg gaagaggtcg tcctggtcat
     4021 tcacgactgg ggctccgctc tgggtttcca ctgggccaag cgcaatccag agcgcgtcaa
     4081 aggtattgca tttatggagt tcatccgccc tatcccgacc tgggacgaat ggccagaatt
     4141 tgcccgcgag accttccagg ccttccgcac caccgacgtc ggccgcaagc tgatcatcga
     4201 tcagaacgtt tttatcgagg gtacgctgcc gatgggtgtc gtccgcccgc tgactgaagt
     4261 cgagatggac cattaccgcg agccgttcct gaatcctgtt gaccgcgagc cactgtggcg
     4321 cttcccaaac gagctgccaa tcgccggtga gccagcgaac atcgtcgcgc tggtcgaaga
     4381 atacatggac tggctgcacc agtcccctgt cccgaagctg ctgttctggg gcaccccagg
     4441 cgttctgatc ccaccggccg aagccgctcg cctggccaaa agcctgccta actgcaaggc
     4501 tgtggacatc ggcccgggtc tgaatctgct gcaagaagac aacccggacc tgatcggcag
     4561 cgagatcgcg cgctggctgt cgacgctgga gatttccggc accggtttgg cggaggcggc
     4621 ggcgaaggag gcggcggcga aggaggcggc ggcgaaggag gcggcggcga aggcggcggc
     4681 gcatatggac aaagactgcg aaatgaagcg caccaccctg gatagccctc tgggcaagct
     4741 ggaactgtct gggtgcgaac agggcctgca ccgtatcatc ttcctgggca aaggaacatc
     4801 tgccgccgac gccgtggaag tgcctgcccc agccgccgtg ctgggcggac cagagccact
     4861 gatgcaggcc accgcctggc tcaacgccta ctttcaccag cctgaggcca tcgaggagtt
     4921 ccctgtgcca gccctgcacc acccagtgtt ccagcaggag agctttaccc gccaggtgct
     4981 gtggaaactg ctgaaagtgg tgaagttcgg agaggtcatc agctacagcc acctggccgc
     5041 cctggccggc aatcccgccg ccaccgccgc cgtgaaaacc gccctgagcg gaaatcccgt
     5101 gcccattctg atcccctgcc accgggtggt gcagggcgac ctggacgtgg ggggctacga
     5161 gggcgggctc gccgtgaaag agtggctgct ggcccacgag ggccacagac tgggcaagcc
     5221 tgggctgggt caattgcacc atcaccatca ccactaatta attaagccag cacagtggtc
     5281 gaccctgtgg aatgtgtgtc agttagggtg tggaaagtcc ccaggctccc cagcaggcag
     5341 aagtatgcaa agcatgcatc tcaattagtc agcaaccagg tgtggaaagt ccccaggctc
     5401 cccagcaggc agaagtatgc aaagcatgca tctcaattag tcagcaacca tagtcccgcc
     5461 cctaactccg cccatcccgc ccctaactcc gcccagttcc gcccattctc cgccccatgg
     5521 ctgactaatt ttttttattt atgcagaggc cgaggccgcc tcggcctctg agctattcca
     5581 gaagtagtga ggaggctttt ttggaggcct aggcttttgc aaaaagctta ccatgaccga
     5641 gtacaagccc acggtgcgcc tcgccacccg cgacgacgtc cccagggccg tacgcaccct
     5701 cgccgccgcg ttcgccgact accccgccac gcgccacacc gtcgatccgg accgccacat
     5761 cgagcgggtc accgagctgc aagaactctt cctcacgcgc gtcgggctcg acatcggcaa
     5821 ggtgtgggtc gcggacgacg gcgccgcggt ggcggtctgg accacgccgg agagcgtcga
     5881 agcgggggcg gtgttcgccg agatcggccc gcgcatggcc gagttgagcg gttcccggct
     5941 ggccgcgcag caacagatgg aaggcctcct ggcgccgcac cggcccaagg agcccgcgtg
     6001 gttcctggcc accgtcggcg tctcgcccga ccaccagggc aagggtctgg gcagcgccgt
     6061 cgtgctcccc ggagtggagg cggccgagcg cgccggggtg cccgccttcc tggagacctc
     6121 cgcgccccgc aacctcccct tctacgagcg gctcggcttc accgtcaccg ccgacgtcga
     6181 ggtgcccgaa ggaccgcgca cctggtgcat gacccgcaag cccggtgcct gacgcccgcc
     6241 ccacgacccg cagcgcccga ccgaaaggag cgcacgaccc catgcatcga taaaataaaa
     6301 gattttattt agtctccaga aaaagggggg aatgaaagac cccacctgta ggtttggcaa
     6361 gctagcttaa gtaacgccat tttgcaaggc atggaaaaat acataactga gaatagagaa
     6421 gttcagatca aggtcaggaa cagatggaac agctgaatat gggccaaaca ggatatctgt
     6481 ggtaagcagt tcctgccccg gctcagggcc aagaacagat ggaacagctg aatatgggcc
     6541 aaacaggata tctgtggtaa gcagttcctg ccccggctca gggccaagaa cagatggtcc
     6601 ccagatgcgg tccagccctc agcagtttct agagaaccat cagatgtttc cagggtgccc
     6661 caaggacctg aaatgaccct gtgccttatt tgaactaacc aatcagttcg cttctcgctt
     6721 ctgttcgcgc gcttctgctc cccgagctca ataaaagagc ccacaacccc tcactcgggg
     6781 cgccagtcct ccgattgact gagtcgcccg ggtacccgtg tatccaataa accctcttgc
     6841 agttgcatcc gacttgtggt ctcgctgttc cttgggaggg tctcctctga gtgattgact
     6901 acccgtcagc gggggtcttt cacatgcagc atgtatcaaa attaatttgg ttttttttct
     6961 taagtattta cattaaatgg ccatagttgc attaatgaat cggccaacgc gcggggagag
     7021 gcggtttgcg tattgggcgc tcttccgctt cctcgctcac tgactcgctg cgctcggtcg
     7081 ttcggctgcg gcgagcggta tcagctcact caaaggcggt aatacggtta tccacagaat
     7141 caggggataa cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta
     7201 aaaaggccgc gttgctggcg tttttccata ggctccgccc ccctgacgag catcacaaaa
     7261 atcgacgctc aagtcagagg tggcgaaacc cgacaggact ataaagatac caggcgtttc
     7321 cccctggaag ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt
     7381 ccgcctttct cccttcggga agcgtggcgc tttctcatag ctcacgctgt aggtatctca
     7441 gttcggtgta ggtcgttcgc tccaagctgg gctgt
//
