LOCUS       sequence-491089-        6419 bp    DNA     circular SYN 29-OCT-2025
DEFINITION  anchoring mCherry-1x HA to the chromatin/nucleus.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6419)
  AUTHORS   Vigano MA, Ell CM, Kustermann MMM, Aguilar G, Matsuda S, Zhao N, 
            Stasevich TJ, Affolter M, Pyrowolakis G
  TITLE     Protein manipulation using single copies of short peptide tags in 
            cultured cells and in Drosophila melanogaster.
  JOURNAL   Development. 2021 Mar 16;148(6):dev191700. doi: 10.1242/dev.191700.
  PUBMED    33593816
REFERENCE   2  (bases 1 to 6419)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Oct 29, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: 
            "Development."; date: "2021-03-16"; volume: "148(6):dev191700. doi";
            pages: " 10.1242/dev.191700"
COMMENT     Sequence Label: mCherry_HA_H2B
FEATURES             Location/Qualifiers
     source          1..6419
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     protein_bind    820..838
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    841..859
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     869..893
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     regulatory      990..999
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             996..1703
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFSEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(1141..1159)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     1409..1428
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     CDS             1719..1745
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             1752..2132
                     /codon_start=1
                     /gene="HIST1H2BJ"
                     /product="human histone H2B"
                     /label=H2B
                     /translation="MPEPAKSAPAPKKGSKKAVTKAQKKGGKKRKRSRKESYSIYVYKV
                     LKQVHPDTGISSKAMGIMNSFVNDIFERIAGEASRLAHYNKRSTITSREIQTAVRLLLP
                     GELAKHAVSEGTKAITKYTSAK"
     primer_bind     complement(2170..2187)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2176..2400
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2446..2874
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2533..2552)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2743..2764
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2883..2903)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2888..3217
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3068..3203
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3130..3149
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3280..3879
                     /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(3280..3299)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     3776..3796
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    4092..4213
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4129..4148)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4183..4202
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4262..4278)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4262..4278)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4275..4297)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4286..4302
                     /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(4310..4316,4317..4334,4335..4340))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4355..4376
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4493..4510)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4664..5252)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4744..4763)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5423..6214,6215..6283))
                     /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     6046..6065
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6284..6388)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ccctatcagt gatagagatc
      841 tccctatcag tgatagagat cgtcgacgag ctcgtttagt gaaccgtcag atcgcctgga
      901 gacgccatcc acgctgtttt gacctccata gaagacaccg ggaccgatcc agcctccgga
      961 ctctagcgtt taaacttaag cttggtaccg ccaccatggt gagcaagggc gaggaggata
     1021 acatggccat catcaaggag ttcatgcgct tcaaggtgca catggagggc tccgtgaacg
     1081 gccacgagtt cgagatcgag ggcgagggcg agggccgccc ctacgagggc acccagaccg
     1141 ccaagctgaa ggtgaccaag ggtggccccc tgcccttcgc ctgggacatc ctgtcccctc
     1201 agttcatgta cggctccaag gcctacgtga agcaccccgc cgacatcccc gactacttga
     1261 agctgtcctt ctccgagggc ttcaagtggg agcgcgtgat gaacttcgag gacggcggcg
     1321 tggtgaccgt gacccaggac tcctccctgc aggacggcga gttcatctac aaggtgaagc
     1381 tgcgcggcac caacttcccc tccgacggcc ccgtaatgca gaagaagacc atgggctggg
     1441 aggcctcctc cgagcggatg taccccgagg acggcgccct gaagggcgag atcaagcaga
     1501 ggctgaagct gaaggacggc ggccactacg acgctgaggt caagaccacc tacaaggcca
     1561 agaagcccgt gcagctgccc ggcgcctaca acgtcaacat caagttggac atcacctccc
     1621 acaacgagga ctacaccatc gtggaacagt acgaacgcgc cgagggccgc cactccaccg
     1681 gcggcatgga cgagctgtac aaggatcaaa gcggtccgta cccatacgat gttccagatt
     1741 acgctaccat catgccagag ccagcgaagt ctgctcccgc cccgaaaaag ggctccaaga
     1801 aggcggtgac taaggcgcag aagaaaggcg gcaagaagcg caagcgcagc cgcaaggaga
     1861 gctattccat ctatgtgtac aaggttctga agcaggtcca ccctgacacc ggcatttcgt
     1921 ccaaggccat gggcatcatg aattcgtttg tgaacgacat tttcgagcgc atcgcaggtg
     1981 aggcttcccg cctggcgcat tacaacaagc gctcgaccat cacctccagg gagatccaga
     2041 cggccgtgcg cctgctgctg cctggggagt tggccaagca cgccgtgtcc gagggtacta
     2101 aggccatcac caagtacacc agcgctaagt gactcgagtc tagagggccc gtttaaaccc
     2161 gctgatcagc ctcgactgtg ccttctagtt gccagccatc tgttgtttgc ccctcccccg
     2221 tgccttcctt gaccctggaa ggtgccactc ccactgtcct ttcctaataa aatgaggaaa
     2281 ttgcatcgca ttgtctgagt aggtgtcatt ctattctggg gggtggggtg gggcaggaca
     2341 gcaaggggga ggattgggaa gacaatagca ggcatgctgg ggatgcggtg ggctctatgg
     2401 cttctgaggc ggaaagaacc agctggggct ctagggggta tccccacgcg ccctgtagcg
     2461 gcgcattaag cgcggcgggt gtggtggtta cgcgcagcgt gaccgctaca cttgccagcg
     2521 ccctagcgcc cgctcctttc gctttcttcc cttcctttct cgccacgttc gccggctttc
     2581 cccgtcaagc tctaaatcgg gggctccctt tagggttccg atttagtgct ttacggcacc
     2641 tcgaccccaa aaaacttgat tagggtgatg gttcacgtag tgggccatcg ccctgataga
     2701 cggtttttcg ccctttgacg ttggagtcca cgttctttaa tagtggactc ttgttccaaa
     2761 ctggaacaac actcaaccct atctcggtct attcttttga tttataaggg attttgccga
     2821 tttcggccta ttggttaaaa aatgagctga tttaacaaaa atttaacgcg aattaattct
     2881 gtggaatgtg tgtcagttag ggtgtggaaa gtccccaggc tccccagcag gcagaagtat
     2941 gcaaagcatg catctcaatt agtcagcaac caggtgtgga aagtccccag gctccccagc
     3001 aggcagaagt atgcaaagca tgcatctcaa ttagtcagca accatagtcc cgcccctaac
     3061 tccgcccatc ccgcccctaa ctccgcccag ttccgcccat tctccgcccc atggctgact
     3121 aatttttttt atttatgcag aggccgaggc cgcctctgcc tctgagctat tccagaagta
     3181 gtgaggaggc ttttttggag gcctaggctt ttgcaaaaag ctcccgggag cttgtatatc
     3241 cattttcgga tctgatcaag agacaggatg aagcttacca tgaccgagta caagcccacg
     3301 gtgcgcctcg ccacccgcga cgacgtcccc agggccgtac gcaccctcgc cgccgcgttc
     3361 gccgactacc ccgccacgcg ccacaccgtc gacccggacc gccacatcga gcgggtcacc
     3421 gagctgcaag aactcttcct cacgcgcgtc gggctcgaca tcggcaaggt gtgggtcgcg
     3481 gacgacggcg ccgcggtggc ggtctggacc acgccggaga gcgtcgaagc gggggcggtg
     3541 ttcgccgaga tcggcccgcg catggccgag ttgagcggtt cccggctggc cgcgcagcaa
     3601 cagatggaag gcctcctggc gccgcaccgg cccaaggagc ccgcgtggtt cctggccacc
     3661 gtcggcgtct cgcccgacca ccagggcaag ggtctgggca gcgccgtcgt gctccccgga
     3721 gtggaggcgg ccgagcgcgc cggggtgccc gccttcctgg agacctccgc gccccgcaac
     3781 ctccccttct acgagcggct cggcttcacc gtcaccgccg acgtcgaggt gcccgaagga
     3841 ccgcgcacct ggtgcatgac ccgcaagccc ggtgcctgac gcccgcccca cgacccgcag
     3901 cgcccgaccg aaaggagcgc acgaccccat gcatcgaaat gaccgaccaa gcgacgccca
     3961 acctgccatc acgagatttc gattccaccg ccgccttcta tgaaaggttg ggcttcggaa
     4021 tcgttttccg ggacgccggc tggatgatcc tccagcgcgg ggatctcatg ctggagttct
     4081 tcgcccaccc caacttgttt attgcagctt ataatggtta caaataaagc aatagcatca
     4141 caaatttcac aaataaagca tttttttcac tgcattctag ttgtggtttg tccaaactca
     4201 tcaatgtatc ttatcatgtc tgtataccgt cgacctctag ctagagcttg gcgtaatcat
     4261 ggtcatagct gtttcctgtg tgaaattgtt atccgctcac aattccacac aacatacgag
     4321 ccggaagcat aaagtgtaaa gcctggggtg cctaatgagt gagctaactc acattaattg
     4381 cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa
     4441 tcggccaacg cgcggggaga ggcggtttgc gtattgggcg ctcttccgct tcctcgctca
     4501 ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg
     4561 taatacggtt atccacagaa tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc
     4621 agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc
     4681 cccctgacga gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac
     4741 tataaagata ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc
     4801 tgccgcttac cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata
     4861 gctcacgctg taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc
     4921 acgaaccccc cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca
     4981 acccggtaag acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag
     5041 cgaggtatgt aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta
     5101 gaagaacagt atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg
     5161 gtagctcttg atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc
     5221 agcagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt
     5281 ctgacgctca gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa
     5341 ggatcttcac ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat
     5401 atgagtaaac ttggtctgac agttaccaat gcttaatcag tgaggcacct atctcagcga
     5461 tctgtctatt tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatac
     5521 gggagggctt accatctggc cccagtgctg caatgatacc gcgagaccca cgctcaccgg
     5581 ctccagattt atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg
     5641 caactttatc cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt
     5701 cgccagttaa tagtttgcgc aacgttgttg ccattgctac aggcatcgtg gtgtcacgct
     5761 cgtcgtttgg tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat
     5821 cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta
     5881 agttggccgc agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca
     5941 tgccatccgt aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat
     6001 agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac
     6061 atagcagaac tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa
     6121 ggatcttacc gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt
     6181 cagcatcttt tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg
     6241 caaaaaaggg aataagggcg acacggaaat gttgaatact catactcttc ctttttcaat
     6301 attattgaag catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt
     6361 agaaaaataa acaaataggg gttccgcgca catttccccg aaaagtgcca cctgacgtc
//
