LOCUS       sequence-381948-        6942 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  3xHA endogenous tagging of pandoravirus/mollivirus genes.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6942)
  AUTHORS   Bisio H, Legendre M, Giry C, Philippe N, Alempic JM, Jeudy S, 
            Abergel C
  TITLE     Evolution of giant pandoravirus revealed by CRISPR/Cas9.
  JOURNAL   Nat Commun. 2023 Jan 26;14(1):428. doi: 10.1038/s41467-023-36145-4.
  PUBMED    36702819
REFERENCE   2  (bases 1 to 6942)
  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 
            Commun."; date: "2023-01-26"; volume: "14(1):428. doi"; pages: " 
            10.1038/s41467-023-36145-4"
COMMENT     Sequence Label: vHB178
FEATURES             Location/Qualifiers
     source          1..6942
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             744..1421
                     /codon_start=1
                     /product="monomeric derivative of DsRed (Campbell et al., 
                     2002)"
                     /label=mRFP1
                     /translation="MASSEDVIKEFMRFKVRMEGSVNGHEFEIEGEGEGRPYEGTQTAK
                     LKVTKGGPLPFAWDILSPQFQYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFEDGGV
                     VTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASTERMYPEDGALKGEIKM
                     RLKLKDGGHYDAEVKTTYMAKKPVQLPGAYKTDIKLDITSHNEDYTIVEQYERAEGRHS
                     TGA"
     CDS             744..1421
                     /codon_start=1
                     /product="monomeric derivative of DsRed (Campbell et al., 
                     2002)"
                     /label=mRFP1
                     /note="mammalian codon-optimized"
                     /translation="MASSEDVIKEFMRFKVRMEGSVNGHEFEIEGEGEGRPYEGTQTAK
                     LKVTKGGPLPFAWDILSPQFQYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFEDGGV
                     VTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASTERMYPEDGALKGEIKM
                     RLKLKDGGHYDAEVKTTYMAKKPVQLPGAYKTDIKLDITSHNEDYTIVEQYERAEGRHS
                     TGA"
     primer_bind     complement(874..892)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     1142..1161
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     1336..1359
                     /label=DsRed1-C
                     /note="DsRed1, forward primer"
     rep_origin      1621..2076
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1708..1727)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1918..1939
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        2193..2553
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2193..2384
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2216..2236)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     CDS             2662..2751
                     /codon_start=1
                     /product="three tandem HA epitope tags"
                     /label=3xHA
                     /translation="YPYDVPDYAGYPYDVPDYAGSYPYDVPDYA"
     CDS             4010..4582
                     /codon_start=1
                     /gene="Streptomyces noursei nat1"
                     /product="nourseothricin acetyltransferase"
                     /label=NrsR
                     /note="confers resistance to nourseothricin"
                     /translation="MGTTLDDTAYRYRTSVPGDAEAIEALDGSFTTDTVFRVTATGDGF
                     TLREVPVDPPLTKVFPDDESDDESDDGEDGDPDSRTFVAYGDDGDLAGFVVVSYSGWNR
                     RLTVEDIEVAPEHRGHGVGRALMGLATEFARERGAGHLWLEVTNVNAPAIHAYRRMGFT
                     LCGLDTALYDGTASDGEQALYMSMPCP"
     polyA_signal    4797..4845
                     /note="synthetic polyadenylation signal"
     primer_bind     complement(4886..4905)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5005..5027
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5065..5083)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5151..5255
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5256..5324,5325..6116)
                     /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     complement(5474..5493)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6287..6875
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6776..6795
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 agatctatat gcaaccctgt gacgagagac agggagtgct gttagagaag agcgcacatt
       61 cagcgctcaa gtggtcttgc gtttggtgtc agcacgaggt gcgccagacc caggtcgcac
      121 gcaggcccct cagcgcggtc gccgtgtgtt ggtattgtgt gcctgagtga tgcggaatgc
      181 gttctgcttg taggcgtacg agaccgacgc caggcggcct gtcgtgctgg gctcgtccgc
      241 ggcaggctct gtagggctgg cgcggagtgg gcgatcgtcc tcccgccgcg aggaaacagc
      301 aacggacagg cttcattgcc acctcatcgc cgctttcgcg ctcgaggcca gggcgtggcg
      361 agcaggtaag aatctgagaa gttgaattgt gaattaacac aaagaagacg aacgaacccg
      421 ccgtggtatg gaagaaggcc acctcaacca atctcaagat cacctcctaa tcgtccaacc
      481 cggcctacaa tcgcgcacat cagcactcga tctcgcactt cggtccgttt ggccctcgag
      541 ccaaagccaa gcagttttcc aggcaccgtt gaagtgcgaa tagatgaaat ttccggtcta
      601 ccccaatgac atggcgtttt gtcacaacac atggcgctgt gctggtggtc tcggtggggc
      661 ctatttattt ggagttgttt gcttgctctc tttcatcttt gtcttcctac acagcagcaa
      721 acaccactct ctcactaaga catatggcct cctccgagga cgtcatcaag gagttcatgc
      781 gcttcaaggt gcgcatggag ggctccgtga acggccacga gttcgagatc gagggcgagg
      841 gcgagggccg cccctacgag ggcacccaga ccgccaagct gaaggtgacc aagggcggcc
      901 ccctgccctt cgcctgggac atcctgtccc ctcagttcca gtacggctcc aaggcctacg
      961 tgaagcaccc cgccgacatc cccgactact tgaagctgtc cttccccgag ggcttcaagt
     1021 gggagcgcgt gatgaacttc gaggacggcg gcgtggtgac cgtgacccag gactcctccc
     1081 tgcaggacgg cgagttcatc tacaaggtga agctgcgcgg caccaacttc ccctccgacg
     1141 gccccgtaat gcagaagaag accatgggct gggaggcctc caccgagcgg atgtaccccg
     1201 aggacggcgc cctgaagggc gagatcaaga tgaggctgaa gctgaaggac ggcggccact
     1261 acgacgccga ggtcaagacc acctacatgg ccaagaagcc cgtgcagctg cccggcgcct
     1321 acaagaccga catcaagctg gacatcacct cccacaacga ggactacacc atcgtggaac
     1381 agtacgagcg cgccgagggc cgccactcca ccggcgccta atctagagtc gacccaacaa
     1441 caacaattgc attcatttta tgtttcaggt tcagggggag atgtgggagg ttttttaaag
     1501 caagtaaaac ctctacaaat gtggtaaaat ccgataagga tcgatccggg ctggcgtaat
     1561 agcgaagagg cccgcaccga tcgcccttcc caacagttgc gcagcctgaa tggcgaatgg
     1621 acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg
     1681 ctacacttgc cagcgcccta gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca
     1741 cgttcgccgg ctttccccgt caagctctaa atcgggggct ccctttaggg ttccgattta
     1801 gtgctttacg gcacctcgac cccaaaaaac ttgattaggg tgatggttca cgtagtgggc
     1861 catcgccctg atagacggtt tttcgccctt tgacgttgga gtccacgttc tttaatagtg
     1921 gactcttgtt ccaaactgga acaacactca accctatctc ggtctattct tttgatttat
     1981 aagggatttt gccgatttcg gcctattggt taaaaaatga gctgatttaa caaaaattta
     2041 acgcgaattt taacaaaata ttaacgctta caatttcctg atgcggtatt ttctccttac
     2101 gcatctgtgc ggtatttcac accgcatacg cggatctgcg cagcaccatg gcctgaaata
     2161 acctctgaaa gaggaacttg gttaggtacc ttctgaggcg gaaagaacca gctgtggaat
     2221 gtgtgtcagt tagggtgtgg aaagtcccca ggctccccag caggcagaag tatgcaaagc
     2281 atgcatctca attagtcagc aaccaggtgt ggaaagtccc caggctcccc agcaggcaga
     2341 agtatgcaaa gcatgcatct caattagtca gcaaccatag tcccgcccct aactccgccc
     2401 atcccgcccc taactccgcc cagttccgcc cattctccgc cccatggctg actaattttt
     2461 tttatttatg cagaggccga ggccgcctcg gcctctgagc tattccagaa gtagtgagga
     2521 ggcttttttg gaggatcaaa cgacgccttg caacaagctg agaaaaaacc aggatcggca
     2581 aggaaggatt ttcaacggaa acgttggcat caccgggtat aaaaaggggc caattttttt
     2641 gttgatttgt tgcgcgaatt ctacccatac gatgttcctg actatgcggg ctatccgtat
     2701 gacgtcccgg actatgcagg ttcctatcca tatgacgttc cagattacgc ttaactcttg
     2761 ttcttatttt ctcaatagga aaagaagacg ggattattgc tttacctata attatagcgc
     2821 ccgaactaag cgcccggaaa aaggcttagt tgacgaggat ggaggttatc gaattttcgg
     2881 cggatgcctc ccggctgagt gtgcagatca cagccgtaag gatttcttca aaccaagggg
     2941 gtgactcctt gaacaaagag aaatcacatg atcttccaaa aaacatgtag gaggggacga
     3001 ttgaaagtcc ccttgaaatt tgactttctt cgtctccttt tacaatctta ggaggaagaa
     3061 aaataattct ctttgcctcg tctttttctg tctgtctttg tgccaaaaac tcgtgttccg
     3121 ttttttcttg tgcgcggtcg ctctcgggcc gatccgcaga ggcagcgcga gtcgcgcaat
     3181 aatcaaaaaa aaaagaagga gaacaagaaa aaaaagcggc aacgtagcgc acaaactgga
     3241 ttttttatga ttgagcacaa gccgcatgag actttttgat cgcaaggcag cattggacgg
     3301 gacgatgtcg ccgccaccag ggcgacggga gtcgcccaaa aatcggcaaa cgaggacggc
     3361 ccgacgggag cgagcctcaa gcttcggctc tgcagttgat ctcttgcgtc tgcggttttt
     3421 ctcctagcga ccttgttggg ccgagggaaa aaaatagtcg ggccgctggt ggctctctgg
     3481 gtcagcgcgg agaggaaata gacaagaaaa taaattaaaa aaagaccaaa caaatcgtgc
     3541 cgatggtgcg agaaaaaggg aggccacaac gccttggcct cttctctttc gttttccctc
     3601 tgcgcggtgc ggcgctgggc ggggcgcgtg ggtccaaaaa aaaaaggacg acacacacac
     3661 acaaaaaata cgaacaaaaa atctacgacg aatagaaggc cccggttcga ggccggcccg
     3721 aaaggaggca ccccaatggc gcccaacggc ggggcacgcc gaaacgacct gccgtcgccg
     3781 gccaatgccc taccgtaaac aaaaaaagaa cacaagcgtc ttcaaaaaaa gacgattggt
     3841 tgcatgcttt gataaataga tcggacctcg catcgtcaaa ccgtatcgac aacagcaacc
     3901 aacacactct cgacaacaac aacaacaaca ataaccgcaa ccgcaaccat catcagcttg
     3961 attcttctga cacaacagtc tcgaacttaa ggctagagcc accgatatca tgggtaccac
     4021 tcttgacgac acggcttacc ggtaccgcac cagtgtcccg ggggacgccg aggccatcga
     4081 ggcactggat gggtccttca ccaccgacac cgtcttccgc gtcaccgcca ccggggacgg
     4141 cttcaccctg cgggaggtgc cggtggaccc gcccctgacc aaggtgttcc ccgacgacga
     4201 atcggacgac gaatcggacg acggggagga cggcgacccg gactctcgga cgttcgtcgc
     4261 gtacggggac gacggcgacc tggcgggctt cgtggtcgtc tcgtactccg gctggaaccg
     4321 ccggctgacc gtcgaggaca tcgaggtcgc cccggagcac cgggggcacg gggtcgggcg
     4381 cgcgttgatg gggctcgcga cggagttcgc ccgcgagcgg ggtgccgggc acctctggct
     4441 ggaggtcacc aacgtcaacg caccggcgat ccacgcgtac cggcggatgg ggttcaccct
     4501 ctgcggcctg gacaccgccc tgtacgacgg caccgcctcg gacggcgagc aggcgctcta
     4561 catgagcatg ccctgcccct aataagcggc gccgtgccgc ctgtctgttt tctctatctc
     4621 ctttttgcag ttcatgcgtg tggacgcgca acttttttca atgcaaggac cacaacaggc
     4681 aataaaaaac ataaaaacac aaaatgtctc taaggcacag cggccgcgat atcgcgggac
     4741 tctggggttc gaaatgaccg accaagcgac gcccaacctg ccatcacgat ggccgcaata
     4801 aaatatcttt attttcatta catctgtgtg ttggtttttt gtgtgaatcg atagcgataa
     4861 ggatccgcgt atggtgcact ctcagtacaa tctgctctga tgccgcatag ttaagccagc
     4921 cccgacaccc gccaacaccc gctgacgcgc cctgacgggc ttgtctgctc ccggcatccg
     4981 cttacagaca agctgtgacc gtctccggga gctgcatgtg tcagaggttt tcaccgtcat
     5041 caccgaaacg cgcgagacga aagggcctcg tgatacgcct atttttatag gttaatgtca
     5101 tgataataat ggtttcttag acgtcaggtg gcacttttcg gggaaatgtg cgcggaaccc
     5161 ctatttgttt atttttctaa atacattcaa atatgtatcc gctcatgaga caataaccct
     5221 gataaatgct tcaataatat tgaaaaagga agagtatgag tattcaacat ttccgtgtcg
     5281 cccttattcc cttttttgcg gcattttgcc ttcctgtttt tgctcaccca gaaacgctgg
     5341 tgaaagtaaa agatgctgaa gatcagttgg gtgcacgagt gggttacatc gaactggatc
     5401 tcaacagcgg taagatcctt gagagttttc gccccgaaga acgttttcca atgatgagca
     5461 cttttaaagt tctgctatgt ggcgcggtat tatcccgtat tgacgccggg caagagcaac
     5521 tcggtcgccg catacactat tctcagaatg acttggttga gtactcacca gtcacagaaa
     5581 agcatcttac ggatggcatg acagtaagag aattatgcag tgctgccata accatgagtg
     5641 ataacactgc ggccaactta cttctgacaa cgatcggagg accgaaggag ctaaccgctt
     5701 ttttgcacaa catgggggat catgtaactc gccttgatcg ttgggaaccg gagctgaatg
     5761 aagccatacc aaacgacgag cgtgacacca cgatgcctgt agcaatggca acaacgttgc
     5821 gcaaactatt aactggcgaa ctacttactc tagcttcccg gcaacaatta atagactgga
     5881 tggaggcgga taaagttgca ggaccacttc tgcgctcggc ccttccggct ggctggttta
     5941 ttgctgataa atctggagcc ggtgagcgtg ggtctcgcgg tatcattgca gcactggggc
     6001 cagatggtaa gccctcccgt atcgtagtta tctacacgac ggggagtcag gcaactatgg
     6061 atgaacgaaa tagacagatc gctgagatag gtgcctcact gattaagcat tggtaactgt
     6121 cagaccaagt ttactcatat atactttaga ttgatttaaa acttcatttt taatttaaaa
     6181 ggatctaggt gaagatcctt tttgataatc tcatgaccaa aatcccttaa cgtgagtttt
     6241 cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttcttga gatccttttt
     6301 ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt
     6361 tgccggatca agagctacca actctttttc cgaaggtaac tggcttcagc agagcgcaga
     6421 taccaaatac tgttcttcta gtgtagccgt agttaggcca ccacttcaag aactctgtag
     6481 caccgcctac atacctcgct ctgctaatcc tgttaccagt ggctgctgcc agtggcgata
     6541 agtcgtgtct taccgggttg gactcaagac gatagttacc ggataaggcg cagcggtcgg
     6601 gctgaacggg gggttcgtgc acacagccca gcttggagcg aacgacctac accgaactga
     6661 gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca
     6721 ggtatccggt aagcggcagg gtcggaacag gagagcgcac gagggagctt ccagggggaa
     6781 acgcctggta tctttatagt cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt
     6841 tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg gcctttttac
     6901 ggttcctggc cttttgctgg ccttttgctc acatggctcg ac
//
