LOCUS       sequence-308972-        6049 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  expresses squirrel monkey retroCHMP3 with self-cleaving tag in 
            mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6049)
  AUTHORS   Rheinemann L, Downhour DM, Bredbenner K, Mercenne G, Davenport KA, 
            Schmitt PT, Necessary CR, McCullough J, Schmitt AP, Simon SM, 
            Sundquist WI, Elde NC
  TITLE     RetroCHMP3 blocks budding of enveloped viruses without blocking 
            cytokinesis.
  JOURNAL   Cell. 2021 Sep 27. pii: S0092-8674(21)01055-2. doi: 
            10.1016/j.cell.2021.09.008.
  PUBMED    34597582
REFERENCE   2  (bases 1 to 6049)
  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: "Cell. 2021 
            Sep 27. pii: S0092-8674(21)01055-2. doi: 
            10.1016/j.cell.2021.09.008."
COMMENT     Sequence Label: pEF1-alpha-squirrel-monkey-retroCHMP3-T2A-H2B-BFP
FEATURES             Location/Qualifiers
     source          1..6049
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(156..172)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(156..172)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(169..191)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    180..196
                     /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(204..210,211..228,229..234))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    249..270
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        329..524
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      375..510
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    530..664
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(567..586)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     621..640
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(732..749)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(903..1491)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(983..1002)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(1662..2453,2454..2522))
                     /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     2285..2304
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2523..2627)
                     /gene="bla"
                     /label=AmpR promoter
     promoter        2689..3871
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          2920..3862
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     3819..3839
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     CDS             4430..4456
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             4457..4510
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     CDS             4511..4888
                     /codon_start=1
                     /gene="HIST1H2BB"
                     /product="human histone H2B"
                     /label=H2B
                     /translation="MPEPSKSAPAPKKGSKKAITKAQKKDGKKRKRSRKESYSIYVYKV
                     LKQVHPDTGISSKAMGIMNSFVNDIFERIAGEASRLAHYNKRSTITSREIQTAVRLLLP
                     GELAKHAVSEGTKAVTKYTSSK"
     regulatory      4901..4910
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             4907..5605
                     /codon_start=1
                     /product="monomeric blue fluorescent protein"
                     /label=TagBFP
                     /note="mammalian codon-optimized"
                     /translation="MVSELIKENMHMKLYMEGTVDNHHFKCTSEGEGKPYEGTQTMRIK
                     VVEGGPLPFAFDILATSFLYGSKTFINHTQGIPDFFKQSFPEGFTWERVTTYEDGGVLT
                     ATQDTSLQDGCLIYNVKIRGVNFTSNGPVMQKKTLGWEAFTETLYPADGGLEGRNDMAL
                     KLVGGSHLIANIKTTYRSKKPAKNLKMPGVYYVDYRLERIKEANNETYVEQHEVAVARY
                     CDLPSKLGHKL"
     primer_bind     complement(5724..5743)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    5789..5844
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(5843..5862)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
ORIGIN
        1 gttttgtgtt atttttttct ttaacatccc taaaattttc cttacatgtt ttactagcca
       61 gatttttcct cctctcctga ctactcccag tcatagctgt ccctcttctc ttatggagat
      121 ccctcgacct gcagcccaag cttggcgtaa tcatggtcat agctgtttcc tgtgtgaaat
      181 tgttatccgc tcacaattcc acacaacata cgagccggaa gcataaagtg taaagcctgg
      241 ggtgcctaat gagtgagcta actcacatta attgcgttgc gctcactgcc cgctttccag
      301 tcgggaaacc tgtcgtgcca gcggatccgc atctcaatta gtcagcaacc atagtcccgc
      361 ccctaactcc gcccatcccg cccctaactc cgcccagttc cgcccattct ccgccccatg
      421 gctgactaat tttttttatt tatgcagagg ccgaggccgc ctcggcctct gagctattcc
      481 agaagtagtg aggaggcttt tttggaggcc taggcttttg caaaaagcta acttgtttat
      541 tgcagcttat aatggttaca aataaagcaa tagcatcaca aatttcacaa ataaagcatt
      601 tttttcactg cattctagtt gtggtttgtc caaactcatc aatgtatctt atcatgtctg
      661 gatccgctgc attaatgaat cggccaacgc gcggggagag gcggtttgcg tattgggcgc
      721 tcttccgctt cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta
      781 tcagctcact caaaggcggt aatacggtta tccacagaat caggggataa cgcaggaaag
      841 aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg
      901 tttttccata ggctccgccc ccctgacgag catcacaaaa atcgacgctc aagtcagagg
      961 tggcgaaacc cgacaggact ataaagatac caggcgtttc cccctggaag ctccctcgtg
     1021 cgctctcctg ttccgaccct gccgcttacc ggatacctgt ccgcctttct cccttcggga
     1081 agcgtggcgc tttctcatag ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc
     1141 tccaagctgg gctgtgtgca cgaacccccc gttcagcccg accgctgcgc cttatccggt
     1201 aactatcgtc ttgagtccaa cccggtaaga cacgacttat cgccactggc agcagccact
     1261 ggtaacagga ttagcagagc gaggtatgta ggcggtgcta cagagttctt gaagtggtgg
     1321 cctaactacg gctacactag aagaacagta tttggtatct gcgctctgct gaagccagtt
     1381 accttcggaa aaagagttgg tagctcttga tccggcaaac aaaccaccgc tggtagcggt
     1441 ggtttttttg tttgcaagca gcagattacg cgcagaaaaa aaggatctca agaagatcct
     1501 ttgatctttt ctacggggtc tgacgctcag tggaacgaaa actcacgtta agggattttg
     1561 gtcatgagat tatcaaaaag gatcttcacc tagatccttt taaattaaaa atgaagtttt
     1621 aaatcaatct aaagtatata tgagtaaact tggtctgaca gttaccaatg cttaatcagt
     1681 gaggcaccta tctcagcgat ctgtctattt cgttcatcca tagttgcctg actccccgtc
     1741 gtgtagataa ctacgatacg ggagggctta ccatctggcc ccagtgctgc aatgataccg
     1801 cgagacccac gctcaccggc tccagattta tcagcaataa accagccagc cggaagggcc
     1861 gagcgcagaa gtggtcctgc aactttatcc gcctccatcc agtctattaa ttgttgccgg
     1921 gaagctagag taagtagttc gccagttaat agtttgcgca acgttgttgc cattgctaca
     1981 ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat tcagctccgg ttcccaacga
     2041 tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag cggttagctc cttcggtcct
     2101 ccgatcgttg tcagaagtaa gttggccgca gtgttatcac tcatggttat ggcagcactg
     2161 cataattctc ttactgtcat gccatccgta agatgctttt ctgtgactgg tgagtactca
     2221 accaagtcat tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata
     2281 cgggataata ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg aaaacgttct
     2341 tcggggcgaa aactctcaag gatcttaccg ctgttgagat ccagttcgat gtaacccact
     2401 cgtgcaccca actgatcttc agcatctttt actttcacca gcgtttctgg gtgagcaaaa
     2461 acaggaaggc aaaatgccgc aaaaaaggga ataagggcga cacggaaatg ttgaatactc
     2521 atactcttcc tttttcaata ttattgaagc atttatcagg gttattgtct catgagcgga
     2581 tacatatttg aatgtattta gaaaaataaa caaatagggg ttccgcgcac atttccccga
     2641 aaagtgccac ctgggtcgac attgattatt gactagatca tcgcgtgagg ctccggtgcc
     2701 cgtcagtggg cagagcgcac atcgcccaca gtccccgaga agttgggggg aggggtcggc
     2761 aattgaaccg gtgcctagag aaggtggcgc ggggtaaact gggaaagtga tgtcgtgtac
     2821 tggctccgcc tttttcccga gggtggggga gaaccgtata taagtgcagt agtcgccgtg
     2881 aacgttcttt ttcgcaacgg gtttgccgcc agaacacagg taagtgccgt gtgtggttcc
     2941 cgcgggcctg gcctctttac gggttatggc ccttgcgtgc cttgaattac ttccacgccc
     3001 ctggctgcag tacgtgattc ttgatcccga gcttcgggtt ggaagtgggt gggagagttc
     3061 gaggccttgc gcttaaggag ccccttcgcc tcgtgcttga gttgaggcct ggcctgggcg
     3121 ctggggccgc cgcgtgcgaa tctggtggca ccttcgcgcc tgtctcgctg ctttcgataa
     3181 gtctctagcc atttaaaatt tttgatgacc tgctgcgacg ctttttttct ggcaagatag
     3241 tcttgtaaat gcgggccaag atctgcacac tggtatttcg gtttttgggg ccgcgggcgg
     3301 cgacggggcc cgtgcgtccc agcgcacatg ttcggcgagg cggggcctgc gagcgcggcc
     3361 accgagaatc ggacgggggt agtctcaagc tggccggcct gctctggtgc ctggcctcgc
     3421 gccgccgtgt atcgccccgc cctgggcggc aaggctggcc cggtcggcac cagttgcgtg
     3481 agcggaaaga tggccgcttc ccggccctgc tgcagggagc tcaaaatgga ggacgcggcg
     3541 ctcgggagag cgggcgggtg agtcacccac acaaaggaaa agggcctttc cgtcctcagc
     3601 cgtcgcttca tgtgactcca cggagtaccg ggcgccgtcc aggcacctcg attagttctc
     3661 gagcttttgg agtacgtcgt ctttaggttg gggggagggg ttttatgcga tggagtttcc
     3721 ccacactgag tgggtggaga ctgaagttag gccagcttgg cacttgatgt aattctcctt
     3781 ggaatttgcc ctttttgagt ttggatcttg gttcattctc aagcctcaga cagtggttca
     3841 aagttttttt cttccatttc aggtgtcgtg aggaattctg cagtcgacgg taccgcgggc
     3901 ccgggatcca ccggtcgcca ccgggaacgg aaagccggaa ggagcaagac gagttcagtt
     3961 cgccatggga ctcttggtta aaacccagga gaagccacca aaagaactgg tcaatgaatg
     4021 gtcattgaag ataagaaagg aaatgagagt tgttgacagg caaatacggg atatccaaag
     4081 agaagaagaa aaagtgaaac gatctgtgaa agaagcagtc aagaagggcc agaaggatgt
     4141 ctgcacagtt ctggccaggg agatgatccg gtcagggacg gctgtgagca agctgtatga
     4201 atccaaagcg cacctggact cggtgctcac ggggatgagg aaccagttgg cggtcttgcg
     4261 aggggctggt tccctgcaga agagtgcaga agtgacgagg gctgtgcaaa gtcttgtgaa
     4321 ggttccagaa attcaggcca ccatgaggga gctgtccaaa gaaatgacga aggctgggat
     4381 catagaggag atgttagagg acactttcga aagcatggac gatcgggaat acccatacga
     4441 tgttccagat tacgctgagg gcagaggaag tctgctaaca tgcggtgacg tcgaggagaa
     4501 tcctggccca atgcctgaac cctctaagtc tgctccagcc cctaaaaagg gttctaagaa
     4561 ggctatcact aaggcgcaga agaaggatgg taagaagcgt aagcgcagcc gcaaggagag
     4621 ctattctatc tatgtgtaca aggttctgaa gcaggtccac cccgacaccg gcatctcatc
     4681 caaggccatg gggatcatga actccttcgt caacgacatc ttcgagcgca tcgcgggcga
     4741 ggcttctcgc ctggctcact acaataagcg ctcgaccatc acctccaggg agattcagac
     4801 ggctgtgcgc ctgctgctgc ctggggagct ggctaagcat gctgtgtccg agggcactaa
     4861 ggcagttacc aagtacacta gctctaagga tccaccggtc gccaccatgg tgagcgagct
     4921 gattaaggag aacatgcaca tgaagctgta catggagggc accgtggaca accatcactt
     4981 caagtgcaca tccgagggcg aaggcaagcc ctacgagggc acccagacca tgagaatcaa
     5041 ggtggtcgag ggcggccctc tccccttcgc cttcgacatc ctggctacta gcttcctcta
     5101 cggcagcaag accttcatca accacaccca gggcatcccc gacttcttca agcagtcctt
     5161 ccctgagggc ttcacatggg agagagtcac cacatacgaa gacgggggcg tgctgaccgc
     5221 tacccaggac accagcctcc aggacggctg cctcatctac aacgtcaaga tcagaggggt
     5281 gaacttcaca tccaacggcc ctgtgatgca gaagaaaaca ctcggctggg aggccttcac
     5341 cgagacgctg taccccgctg acggcggcct ggaaggcaga aacgacatgg ccctgaagct
     5401 cgtgggcggg agccatctga tcgcaaacat caagaccaca tatagatcca agaaacccgc
     5461 taagaacctc aagatgcctg gcgtctacta tgtggactac agactggaaa gaatcaagga
     5521 ggccaacaac gagacctacg tcgagcagca cgaggtggca gtggccagat actgcgacct
     5581 ccctagcaaa ctggggcaca agcttggcat ggacgagctg tacaagtaat gaaagcttgc
     5641 ggccgcactc ctcaggtgca ggctgcctat cagaaggtgg tggctggtgt ggccaatgcc
     5701 ctggctcaca aataccactg agatcttttt ccctctgcca aaaattatgg ggacatcatg
     5761 aagccccttg agcatctgac ttctggctaa taaaggaaat ttattttcat tgcaatagtg
     5821 tgttggaatt ttttgtgtct ctcactcgga aggacatatg ggagggcaaa tcatttaaaa
     5881 catcagaatg agtatttggt ttagagtttg gcaacatatg cccatatgct ggctgccatg
     5941 aacaaaggtt ggctataaag aggtcatcag tatatgaaac agccccctgc tgtccattcc
     6001 ttattccata gaaaagcctt gacttgaggt tagatttttt ttatatttt
//
