LOCUS       sequence-311841-        8268 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Shield-1 inducible transient mammalian expression of Dam-LMNB1 for 
            DamID, containing the V133A mutant of Dam..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8268)
  AUTHORS   Altemose N, Maslan A, Rios-Martinez C, Lai A, White JA, Streets A
  TITLE     muDamID: A Microfluidic Approach for Joint Imaging and Sequencing of
            Protein-DNA Interactions in Single Cells.
  JOURNAL   Cell Syst. 2020 Oct 21;11(4):354-366.e9. doi: 
            10.1016/j.cels.2020.08.015. Epub 2020 Sep 23.
  PUBMED    33099405
REFERENCE   2  (bases 1 to 8268)
  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: 
            "10.1016/j.cels.2020.08.015"; journalName: "Cell Syst"; date: 
            "2020-10-21- 21"; volume: "11"; issue: "4"; pages: "354-366.e9"
COMMENT     Sequence Label: DD-DamV133A-LMNB1-IRES2-mCherry
FEATURES             Location/Qualifiers
     source          1..8268
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        61..364
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        365..568
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     519..539
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     CDS             join(597..599,600..920)
                     /codon_start=1
                     /product="destabilization domain that can be stabilized by 
                     Shield1 in the ProteoTuner(TM) system"
                     /label=DD
                     /note="L106P mutant of FKBP12"
                     /translation="MGVQVETISPGDGRTFPKRGQTCVVHYTGMLEDGKKVDSSRDRNK
                     PFKFMLGKQEVIRGWEEGVAQMSVGQRAKLTISPDYAYGATGHPGIIPPHATLVFDVEL
                     LKPE"
     CDS             1806..1847
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     protein_bind    1863..1887
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     protein_bind    complement(3685..3709)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     misc_feature    join(3741..4315,4316..4318,4319..4327)
                     /label=IRES2
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(3908..3925)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     4135..4154
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             4328..5038
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(4473..4491)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     4741..4760
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     polyA_signal    5056..5177
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5093..5112)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5147..5166
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(5184..5639)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5321..5342)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     5533..5552
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        5666..5770
                     /gene="bla"
                     /label=AmpR promoter
     promoter        5772..6129
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        5772..5963
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(5795..5815)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      5980..6115
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             6164..6958
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             6164..6958
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(6218..6237)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     6828..6847
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(7146..7165)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    7190..7237
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      7566..8154
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     8055..8074
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg
       61 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
      121 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      181 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      241 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      301 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      361 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
      421 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
      481 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
      541 acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatcc gctagcatgg
      601 gagtgcaggt ggaaaccatc tccccaggag acgggcgcac cttccccaag cgcggccaga
      661 cctgtgtggt gcactacacc gggatgcttg aagatggaaa gaaagtcgat tcctcccggg
      721 acagaaacaa gccctttaag tttatgctag gcaagcagga ggtgatccga ggctgggaag
      781 aaggggttgc ccagatgagt gtgggtcaga gagccaaact gactatatct ccagattatg
      841 cctatggtgc cactgggcac ccaggcatca tcccaccaca tgccactctc gtcttcgatg
      901 tggagcttct aaaaccggaa gctagcgcta ccggactcag atcttcacac acagctagcg
      961 ccaccatgaa gaaaaatcgc gcttttttga agtgggcagg gggcaagtat cccctgcttg
     1021 atgatattaa acggcatttg cccaagggcg aatgtctggt tgagcctttt gtaggtgccg
     1081 ggtcggtgtt tctcaacacc gacttttctc gttatatcct tgccgatatc aatagcgacc
     1141 tgatcagtct ctataacatt gtgaagatgc gtactgatga gtacgtacag gccgcacgcg
     1201 agctgtttgt tcccgaaaca aattgcgccg aggtttacta tcagttccgc gaagagttca
     1261 acaaaagcca ggatccgttc cgtcgggcgg tactgttttt atatttgaac cgctacggtt
     1321 acaacggcct gtgtcgttac aatctgcgcg gtgagtttaa cgcgccgttc ggccgctaca
     1381 aaaaacccta tttcccggaa gcagagttgt atcacttcgc tgaaaaagcg cagaatgcct
     1441 ttttctattg tgagtcttac gccgatagca tggcgcgcgc agatgatgca tccgtcgtct
     1501 attgcgatcc gccttatgca ccgctgtctg cgaccgccaa ctttacggcg tatcacacaa
     1561 acagttttac gcttgaacaa caagcgcatc tggcggagat cgccgaaggt ctggttgagc
     1621 gccatattcc agtgctgatc tccaatcacg atacgatgtt aacgcgtgag tggtatcagc
     1681 gcgcaaaatt gcatgtcgtc aaagttcgac gcagtataag cagcaacggc ggcacacgta
     1741 aaaaggtgga cgaactgctg gctttgtaca aaccaggagt cgtttcaccc gcgaaaaaaa
     1801 ccggtggtaa gcctatccct aaccctctcc tcggtctcga ttctacgatc gatttcgatc
     1861 aaacaagttt gtacaaaaaa gcaggctccg cggccgcccc cttcaccatg gcgactgcga
     1921 cccccgtgcc gccgcggatg ggcagccgcg ctggcggccc caccacgccg ctgagcccca
     1981 cgcgcctgtc gcggctccag gagaaggagg agctgcgcga gctcaatgac cggctggcgg
     2041 tgtacatcga caaggtgcgc agcctggaga cggagaacag cgcgctgcag ctgcaggtga
     2101 cggagcgcga ggaggtgcgc ggccgtgagc tcaccggcct caaggcgctc tacgagaccg
     2161 agctggccga cgcgcgacgc gcgctcgacg acacggcccg cgagcgcgcc aagctgcaga
     2221 tcgagctggg caagtgcaag gcggaacacg accagctgct cctcaactat gctaagaagg
     2281 aatctgatct taatggcgcc cagatcaagc ttcgagaata tgaagcagca ctgaattcga
     2341 aagatgcagc tcttgctact gcacttggtg acaaaaaaag tttagaggga gatttggagg
     2401 atctgaagga tcagattgcc cagttggaag cctccttagc tgcagccaaa aaacagttag
     2461 cagatgaaac tttacttaaa gtagatttgg agaatcgttg tcagagcctt actgaggact
     2521 tggagtttcg caaaagcatg tatgaagagg agattaacga gaccagaagg aagcatgaaa
     2581 cgcgcttggt agaggtggat tctgggcgtc aaattgagta tgagtacaag ctggcgcaag
     2641 cccttcatga gatgagagag caacatgatg cccaagtgag gctgtataag gaggagctgg
     2701 agcagactta ccatgccaaa cttgagaatg ccagactgtc atcagagatg aatacttcta
     2761 ctgtcaacag tgccagggaa gaactgatgg aaagccgcat gagaattgag agcctttcat
     2821 cccagctttc taatctacag aaagagtcta gagcatgttt ggaaaggatt caagaattag
     2881 aggacttgct tgctaaagaa aaagacaact ctcgtcgcat gctgacagac aaagagagag
     2941 agatggcgga aataagggat caaatgcagc aacagctgaa tgactatgaa cagcttcttg
     3001 atgtaaagtt agccctggac atggaaatca gtgcttacag gaaactctta caaggcgaag
     3061 aagagaggtt gaagctgtct ccaagccctt cttcccgtgt gacagtatcc cgagcatcct
     3121 caagtcgtag tgtacgtaca actagaggaa agcggaagag ggttgatgtg gaagaatcag
     3181 aggcgagtag tagtgttagc atctctcatt ccgcctcagc cactggaaat gtttgcatcg
     3241 aagaaattga tgttgatggg aaatttatcc gcttgaagaa cacttctgaa caggatcaac
     3301 caatgggagg ctgggagatg atcagaaaaa ttggagacac atcagtcagt tataaatata
     3361 cctcaagata tgtgctgaag gcaggccaga ctgttacaat ttgggctgca aacgctggtg
     3421 tcacagccag ccccccaact gacctcatct ggaagaacca gaactcgtgg ggcactggcg
     3481 aagatgtgaa ggttatattg aaaaattctc agggagagga ggttgctcaa agaagtacag
     3541 tctttaaaac aaccatacct gaagaagagg aggaggagga agaagcagct ggagtggttg
     3601 ttgaggaaga acttttccac cagcagggaa ccccaagagc atccaataga agctgtgcaa
     3661 ttatgtaaaa gggtgggcgc gccgacccag ctttcttgta caaagtggtt cggtcgacgg
     3721 taccgcgggc ccgggatccg cccctctccc tccccccccc ctaacgttac tggccgaagc
     3781 cgcttggaat aaggccggtg tgcgtttgtc tatatgttat tttccaccat attgccgtct
     3841 tttggcaatg tgagggcccg gaaacctggc cctgtcttct tgacgagcat tcctaggggt
     3901 ctttcccctc tcgccaaagg aatgcaaggt ctgttgaatg tcgtgaagga agcagttcct
     3961 ctggaagctt cttgaagaca aacaacgtct gtagcgaccc tttgcaggca gcggaacccc
     4021 ccacctggcg acaggtgcct ctgcggccaa aagccacgtg tataagatac acctgcaaag
     4081 gcggcacaac cccagtgcca cgttgtgagt tggatagttg tggaaagagt caaatggctc
     4141 tcctcaagcg tattcaacaa ggggctgaag gatgcccaga aggtacccca ttgtatggga
     4201 tctgatctgg ggcctcggta cacatgcttt acatgtgttt agtcgaggtt aaaaaaacgt
     4261 ctaggccccc cgaaccacgg ggacgtggtt ttcctttgaa aaacacgatg ataatatggc
     4321 cacaaccatg gtgagcaagg gcgaggagga taacatggcc atcatcaagg agttcatgcg
     4381 cttcaaggtg cacatggagg gctccgtgaa cggccacgag ttcgagatcg agggcgaggg
     4441 cgagggccgc ccctacgagg gcacccagac cgccaagctg aaggtgacca agggtggccc
     4501 cctgcccttc gcctgggaca tcctgtcccc tcagttcatg tacggctcca aggcctacgt
     4561 gaagcacccc gccgacatcc ccgactactt gaagctgtcc ttccccgagg gcttcaagtg
     4621 ggagcgcgtg atgaacttcg aggacggcgg cgtggtgacc gtgacccagg actcctccct
     4681 gcaggacggc gagttcatct acaaggtgaa gctgcgcggc accaacttcc cctccgacgg
     4741 ccccgtaatg cagaagaaga ccatgggctg ggaggcctcc tccgagcgga tgtaccccga
     4801 ggacggcgcc ctgaagggcg agatcaagca gaggctgaag ctgaaggacg gcggccacta
     4861 cgacgctgag gtcaagacca cctacaaggc caagaagccc gtgcagctgc ccggcgccta
     4921 caacgtcaac atcaagttgg acatcacctc ccacaacgag gactacacca tcgtggaaca
     4981 gtacgaacgc gccgagggcc gccactccac cggcggcatg gacgagctgt acaagtaatg
     5041 acaattgttg ttgttaactt gtttattgca gcttataatg gttacaaata aagcaatagc
     5101 atcacaaatt tcacaaataa agcatttttt tcactgcatt ctagttgtgg tttgtccaaa
     5161 ctcatcaatg tatcttaagg cgtaaattgt aagcgttaat attttgttaa aattcgcgtt
     5221 aaatttttgt taaatcagct cattttttaa ccaataggcc gaaatcggca aaatccctta
     5281 taaatcaaaa gaatagaccg agatagggtt gagtgttgtt ccagtttgga acaagagtcc
     5341 actattaaag aacgtggact ccaacgtcaa agggcgaaaa accgtctatc agggcgatgg
     5401 cccactacgt gaaccatcac cctaatcaag ttttttgggg tcgaggtgcc gtaaagcact
     5461 aaatcggaac cctaaaggga gcccccgatt tagagcttga cggggaaagc cggcgaacgt
     5521 ggcgagaaag gaagggaaga aagcgaaagg agcgggcgct agggcgctgg caagtgtagc
     5581 ggtcacgctg cgcgtaacca ccacacccgc cgcgcttaat gcgccgctac agggcgcgtc
     5641 aggtggcact tttcggggaa atgtgcgcgg aacccctatt tgtttatttt tctaaataca
     5701 ttcaaatatg tatccgctca tgagacaata accctgataa atgcttcaat aatattgaaa
     5761 aaggaagagt cctgaggcgg aaagaaccag ctgtggaatg tgtgtcagtt agggtgtgga
     5821 aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa ttagtcagca
     5881 accaggtgtg gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag catgcatctc
     5941 aattagtcag caaccatagt cccgccccta actccgccca tcccgcccct aactccgccc
     6001 agttccgccc attctccgcc ccatggctga ctaatttttt ttatttatgc agaggccgag
     6061 gccgcctcgg cctctgagct attccagaag tagtgaggag gcttttttgg aggcctaggc
     6121 ttttgcaaag atcgatcaag agacaggatg aggatcgttt cgcatgattg aacaagatgg
     6181 attgcacgca ggttctccgg ccgcttgggt ggagaggcta ttcggctatg actgggcaca
     6241 acagacaatc ggctgctctg atgccgccgt gttccggctg tcagcgcagg ggcgcccggt
     6301 tctttttgtc aagaccgacc tgtccggtgc cctgaatgaa ctgcaagacg aggcagcgcg
     6361 gctatcgtgg ctggccacga cgggcgttcc ttgcgcagct gtgctcgacg ttgtcactga
     6421 agcgggaagg gactggctgc tattgggcga agtgccgggg caggatctcc tgtcatctca
     6481 ccttgctcct gccgagaaag tatccatcat ggctgatgca atgcggcggc tgcatacgct
     6541 tgatccggct acctgcccat tcgaccacca agcgaaacat cgcatcgagc gagcacgtac
     6601 tcggatggaa gccggtcttg tcgatcagga tgatctggac gaagagcatc aggggctcgc
     6661 gccagccgaa ctgttcgcca ggctcaaggc gagcatgccc gacggcgagg atctcgtcgt
     6721 gacccatggc gatgcctgct tgccgaatat catggtggaa aatggccgct tttctggatt
     6781 catcgactgt ggccggctgg gtgtggcgga ccgctatcag gacatagcgt tggctacccg
     6841 tgatattgct gaagagcttg gcggcgaatg ggctgaccgc ttcctcgtgc tttacggtat
     6901 cgccgctccc gattcgcagc gcatcgcctt ctatcgcctt cttgacgagt tcttctgagc
     6961 gggactctgg ggttcgaaat gaccgaccaa gcgacgccca acctgccatc acgagatttc
     7021 gattccaccg ccgccttcta tgaaaggttg ggcttcggaa tcgttttccg ggacgccggc
     7081 tggatgatcc tccagcgcgg ggatctcatg ctggagttct tcgcccaccc tagggggagg
     7141 ctaactgaaa cacggaagga gacaataccg gaaggaaccc gcgctatgac ggcaataaaa
     7201 agacagaata aaacgcacgg tgttgggtcg tttgttcata aacgcggggt tcggtcccag
     7261 ggctggcact ctgtcgatac cccaccgaga ccccattggg gccaatacgc ccgcgtttct
     7321 tccttttccc caccccaccc cccaagttcg ggtgaaggcc cagggctcgc agccaacgtc
     7381 ggggcggcag gccctgccat agcctcaggt tactcatata tactttagat tgatttaaaa
     7441 cttcattttt aatttaaaag gatctaggtg aagatccttt ttgataatct catgaccaaa
     7501 atcccttaac gtgagttttc gttccactga gcgtcagacc ccgtagaaaa gatcaaagga
     7561 tcttcttgag atcctttttt tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg
     7621 ctaccagcgg tggtttgttt gccggatcaa gagctaccaa ctctttttcc gaaggtaact
     7681 ggcttcagca gagcgcagat accaaatact gttcttctag tgtagccgta gttaggccac
     7741 cacttcaaga actctgtagc accgcctaca tacctcgctc tgctaatcct gttaccagtg
     7801 gctgctgcca gtggcgataa gtcgtgtctt accgggttgg actcaagacg atagttaccg
     7861 gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca cacagcccag cttggagcga
     7921 acgacctaca ccgaactgag atacctacag cgtgagctat gagaaagcgc cacgcttccc
     7981 gaagggagaa aggcggacag gtatccggta agcggcaggg tcggaacagg agagcgcacg
     8041 agggagcttc cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc
     8101 tgacttgagc gtcgattttt gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc
     8161 agcaacgcgg cctttttacg gttcctggcc ttttgctggc cttttgctca catgttcttt
     8221 cctgcgttat cccctgattc tgtggataac cgtattaccg ccatgcat
//
