LOCUS       sequence-420143-        8907 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  piggyBac vector to express FLAG-tNGFR with puromycin resistance.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8907)
  AUTHORS   Najia MA, Borrajo J, Le A, Tsai F, Huang JY, Griffith LG, Daley GQ, 
            Blainey PC
  TITLE     Live-cell transcriptomics with engineered virus-like particles
  JOURNAL   bioRxiv 2024.10.01.616098
REFERENCE   2  (bases 1 to 8907)
  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: "bioRxiv 
            2024.10.01.616098"
COMMENT     Sequence Label: pPB_EF1a_FLAG-tNGFR_PuroR
FEATURES             Location/Qualifiers
     source          1..8907
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   127..189
                     /label=piggyBac right (3') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     promoter        1356..2534
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          1587..2525
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     2482..2502
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     regulatory      2541..2550
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2652..2675
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             3420..3476
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     CDS             3477..4076
                     /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(3477..3496)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     3973..3993
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     misc_feature    4092..4680
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(4145..4165)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(4563..4574)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    4730..4851
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4767..4786)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4821..4840
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     repeat_region   5614..5648
                     /label=piggyBac left (5') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     primer_bind     complement(6139..6155)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6139..6155)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(6152..6174)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    6163..6179
                     /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(6187..6193,6194..6211,6212..6217))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6232..6253
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(6370..6387)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6541..7129)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6621..6640)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(7300..8091,8092..8160))
                     /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     7923..7942
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(8161..8265)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(8291..8746)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(8428..8449)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     8640..8659
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     8873..8895
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     8887..8904
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     8888..8904
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
ORIGIN
        1 cgcgcctcgt tcattcacgt ttttgaaccc gtggaggacg ggcagactcg cggtgcaaat
       61 gtgttttaca gcgtgatgga gcagatgaag atgctcgaca cgctgcagaa cacgcagcta
      121 gattaaccct agaaagataa tcatattgtg acgtacgtta aagataatca tgtgtaaaat
      181 tgacgcatgt gttttatcgg tctgtatatc gaggtttatt tattaatttg aatagatatt
      241 aagttttatt atatttacac ttacatacta ataataaatt caacaaacaa tttatttatg
      301 tttatttatt tattaaaaaa aacaaaaact caaaatttct tctataaagt aacaaaactt
      361 ttatgaggga cagccccccc ccaaagcccc cagggatgta attacgtccc tcccccgcta
      421 gggggcagca gcgagccgcc cggggctccg ctccggtccg gcgctccccc cgcatccccg
      481 agccggcagc gtgcggggac agcccgggca cggggaaggt ggcacgggat cgctttcctc
      541 tgaacgcttc tcgctgctct ttgagcctgc agacacctgg ggggatacgg ggaaaaggcc
      601 tccaccataa agatggttaa ttaacccacc caagatctgg atcgataagc ttgatatcga
      661 attcgggagg tggtccctgc agttacgcca atgataaccc ccgccagaaa aatcttagta
      721 gccttccctt tttgttttcc gtgccccaac tcggcggatt gactcggccc cttccggaaa
      781 cacccgaatc aacttctagt caaattattg ttcacgccgc aatgacccac ccctggcccg
      841 cgtctgtgga actgacccct ggtgtacagg agagttcgct gctgaaagtg gtcccaaagg
      901 ggtactagtt tttaagctcc caactccccc tcccccagcg tctggaggat tccacaccct
      961 cgcaccgcag gggcgaggaa gtgggcggag tccggttttg gcgccagccg ctgaggctgc
     1021 caagcagaaa agccaccgct gaggagactc cggtcactgt cctcgccccg cctccccctt
     1081 ccctcccctt ggggaccacc gggcgccacg ccgcgaacgg taagtgccgc ggtcgtcggc
     1141 gcctccgccc tccccctagg gccccaattc ccagcgggcg cggcgcgcgg ccccgcgcgc
     1201 agctcccggc tccctccccc ttcggatgtg gcttgagctg taggcgcgga gtcctgcagc
     1261 cccgataaaa taaaagattt tatttagtct ccagaaaaag gggggaatga aagaccccac
     1321 ctgtaggttt ggcaatcttg aaaggagtgg gaattggctc cggtgcccgt cagtgggcag
     1381 agcgcacatc gcccacagtc cccgagaagt tggggggagg ggtcggcaat tgatccggtg
     1441 cctagagaag gtggcgcggg gtaaactggg aaagtgatgt cgtgtactgg ctccgccttt
     1501 ttcccgaggg tgggggagaa ccgtatataa gtgcagtagt cgccgtgaac gttctttttc
     1561 gcaacgggtt tgccgccaga acacaggtaa gtgccgtgtg tggttcccgc gggcctggcc
     1621 tctttacggg ttatggccct tgcgtgcctt gaattacttc cacctggctg cagtacgtga
     1681 ttcttgatcc cgagcttcgg gttggaagtg ggtgggagag ttcgaggcct tgcgcttaag
     1741 gagccccttc gcctcgtgct tgagttgagg cctggcctgg gcgctggggc cgccgcgtgc
     1801 gaatctggtg gcaccttcgc gcctgtctcg ctgctttcga taagtctcta gccatttaaa
     1861 atttttgatg acctgctgcg acgctttttt tctggcaaga tagtcttgta aatgcgggcc
     1921 aagatctgca cactggtatt tcggtttttg gggccgcggg cggcgacggg gcccgtgcgt
     1981 cccagcgcac atgttcggcg aggcggggcc tgcgagcgcg gccaccgaga atcggacggg
     2041 ggtagtctca agctggccgg cctgctctgg tgcctggcct cgcgccgccg tgtatcgccc
     2101 cgccctgggc ggcaaggctg gcccggtcgg caccagttgc gtgagcggaa agatggccgc
     2161 ttcccggccc tgctgcaggg agctcaaaat ggaggacgcg gcgctcggga gagcgggcgg
     2221 gtgagtcacc cacacaaagg aaaagggcct ttccgtcctc agccgtcgct tcatgtgact
     2281 ccacggagta ccgggcgccg tccaggcacc tcgattagtt ctcgagcttt tggagtacgt
     2341 cgtctttagg ttggggggag gggttttatg cgatggagtt tccccacact gagtgggtgg
     2401 agactgaagt taggccagct tggcacttga tgtaattctc cttggaattt gccctttttg
     2461 agtttggatc ttggttcatt ctcaagcctc agacagtggt tcaaagtttt tttcttccat
     2521 ttcaggtgtc gtgacgtacg gccaccatgg gggcaggtgc caccggccgc gccatggacg
     2581 ggccgcgcct gctgctgttg ctgcttctgg gggtgtccct tggaggtgcc aaggaggcat
     2641 gcgggggagg ggactacaaa gacgatgacg acaagggagg aggccccaca ggcctgtaca
     2701 cacacagcgg tgagtgctgc aaagcctgca acctgggcga gggtgtggcc cagccttgtg
     2761 gagccaacca gaccgtgtgt gagccctgcc tggacagcgt gacgttctcc gacgtggtga
     2821 gcgcgaccga gccgtgcaag ccgtgcaccg agtgcgtggg gctccagagc atgtcggcgc
     2881 cgtgcgtgga ggccgacgac gccgtgtgcc gctgcgccta cggctactac caggatgaga
     2941 cgactgggcg ctgcgaggcg tgccgcgtgt gcgaggcggg ctcgggcctc gtgttctcct
     3001 gccaggacaa gcagaacacc gtgtgcgagg agtgccccga cggcacgtat tccgacgagg
     3061 ccaaccacgt ggacccgtgc ctgccctgca ccgtgtgcga ggacaccgag cgccagctcc
     3121 gcgagtgcac acgctgggcc gacgccgagt gcgaggagat ccctggccgt tggattacac
     3181 ggtccacacc cccagagggc tcggacagca cagcccccag cacccaggag cctgaggcac
     3241 ctccagaaca agacctcata gccagcacgg tggcaggtgt ggtgaccaca gtgatgggca
     3301 gctcccagcc cgtggtgacc cgaggcacca ccgacaacct catccctgtc tattgctcca
     3361 tcctggctgc tgtggttgtg ggccttgtgg cctacatagc cttcaagagg ggatccggcg
     3421 caacaaactt ctctctgctc aaacaagccg gcgacgtcga agagaatcct gggccgatga
     3481 ccgagtacaa gcccacggtg cgcctcgcca cccgcgacga cgtccccagg gccgtacgca
     3541 ccctcgccgc cgcgttcgcc gactaccccg ccacgcgcca caccgtcgat ccggaccgcc
     3601 acatcgagcg ggtcaccgag ctgcaagaac tcttcctcac gcgcgtcggg ctcgacatcg
     3661 gcaaggtgtg ggtcgcggac gacggcgccg cggtggcggt ctggaccacg ccggagagcg
     3721 tcgaagcggg ggcggtgttc gccgagatcg gcccgcgcat ggccgagttg agcggttccc
     3781 ggctggccgc gcagcaacag atggaaggcc tcctggcgcc gcaccggccc aaggagcccg
     3841 cgtggttcct ggccaccgtc ggcgtctcgc ccgaccacca gggcaagggt ctgggcagcg
     3901 ccgtcgtgct ccccggagtg gaggcggccg agcgcgccgg ggtgcccgcc ttcctggaga
     3961 cctccgcgcc ccgcaacctc cccttctacg agcggctcgg cttcaccgtc accgccgacg
     4021 tcgaggtgcc cgaaggaccg cgcacctggt gcatgacccg caagcccggt gcctaaacgc
     4081 gttaagtcga caatcaacct ctggattaca aaatttgtga aagattgact ggtattctta
     4141 actatgttgc tccttttacg ctatgtggat acgctgcttt aatgcctttg tatcatgcta
     4201 ttgcttcccg tatggctttc attttctcct ccttgtataa atcctggttg ctgtctcttt
     4261 atgaggagtt gtggcccgtt gtcaggcaac gtggcgtggt gtgcactgtg tttgctgacg
     4321 caacccccac tggttggggc attgccacca cctgtcagct cctttccggg actttcgctt
     4381 tccccctccc tattgccacg gcggaactca tcgccgcctg ccttgcccgc tgctggacag
     4441 gggctcggct gttgggcact gacaattccg tggtgttgtc ggggaaatca tcgtcctttc
     4501 cttggctgct cgcctgtgtt gccacctgga ttctgcgcgg gacgtccttc tgctacgtcc
     4561 cttcggccct caatccagcg gaccttcctt cccgcggcct gctgccggct ctgcggcctc
     4621 ttccgcgtct tcgccttcgc cctcagacga gtcggatctc cctttgggcc gcctccccgc
     4681 ctcgagtcta gagggcccgt ttaaacccgc tgatcagcct aactaactaa acttgtttat
     4741 tgcagcttat aatggttaca aataaagcaa tagcatcaca aatttcacaa ataaagcatt
     4801 tttttcactg cattctagtt gtggtttgtc caaactcatc aatgtatctt atcatgtctg
     4861 gaattgactc aaatgatgtc aattagtcta tcagaagctc atctggtctc ccttccgggg
     4921 gacaagacat ccctgtttaa tatttaaaca gcagtgttcc caaactgggt tcttatatcc
     4981 cttgctctgg tcaaccaggt tgcagggttt cctgtcctca caggaacgaa gtccctaaag
     5041 aaacagtggc agccaggttt agccccggaa ttgactggat tcctttttta gggcccattg
     5101 gtatggcttt ttccccgtat ccccccaggt gtctgcaggc tcaaagagca gcgagaagcg
     5161 ttcagaggaa agcgatcccg tgccaccttc cccgtgcccg ggctgtcccc gcacgctgcc
     5221 ggctcgggga tgcgggggga gcgccggacc ggagcggagc cccgggcggc tcgctgctgc
     5281 cccctagcgg gggagggacg taattacatc cctgggggct ttgggggggg gctgtccctg
     5341 atatctataa caagaaaata tatatataat aagttatcac gtaagtagaa catgaaataa
     5401 caatataatt atcgtatgag ttaaatctta aaagtcacgt aaaagataat catgcgtcat
     5461 tttgactcac gcggtcgtta tagttcaaaa tcagtgacac ttaccgcatt gacaagcacg
     5521 cctcacggga gctccaagcg gcgactgaga tgtcctaaat gcacagcgac ggattcgcgc
     5581 tatttagaaa gagagagcaa tatttcaaga atgcatgcgt caattttacg cagactatct
     5641 ttctagggtt aatctagctg catcaggatc atatcgtcgg gtcttttttc cggctcagtc
     5701 atcgcccaag ctggcgctat ctgggcatcg gggaggaaga agcccgtgcc ttttcccgcg
     5761 aggttgaagc ggcatggaaa gagtttgccg aggatgactg ctgctgcatt gacgttgagc
     5821 gaaaacgcac gtttaccatg atgattcggg aaggtgtggc catgcacgcc tttaacggtg
     5881 aactgttcgt tcaggccacc tgggatacca gttcgtcgcg gcttttccgg acacagttcc
     5941 ggatggtcag cccgaagcgc atcagcaacc cgaacaatac cggcgacagc cggaactgcc
     6001 gtgccggtgt gcagattaat gacagcggtg cggcgctggg atattacgtc agcgaggacg
     6061 ggtatcctgg ctggatgccg cagaaatgga catggatacc ccgtgagtta cccggcgggc
     6121 gcgcttggcg taatcatggt catagctgtt tcctgtgtga aattgttatc cgctcacaat
     6181 tccacacaac atacgagccg gaagcataaa gtgtaaagcc tggggtgcct aatgagtgag
     6241 ctaactcaca ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg
     6301 ccagctgcat taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc
     6361 ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc
     6421 agctcactca aaggcggtaa tacggttatc cacagaatca ggggataacg caggaaagaa
     6481 catgtgagca aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt
     6541 tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg
     6601 gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg
     6661 ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag
     6721 cgtggcgctt tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc
     6781 caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa
     6841 ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg
     6901 taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc
     6961 taactacggc tacactagaa gaacagtatt tggtatctgc gctctgctga agccagttac
     7021 cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg
     7081 tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt
     7141 gatcttttct acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt
     7201 catgagatta tcaaaaagga tcttcaccta gatcctttta aattaaaaat gaagttttaa
     7261 atcaatctaa agtatatatg agtaaacttg gtctgacagt taccaatgct taatcagtga
     7321 ggcacctatc tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt
     7381 gtagataact acgatacggg agggcttacc atctggcccc agtgctgcaa tgataccgcg
     7441 agacccacgc tcaccggctc cagatttatc agcaataaac cagccagccg gaagggccga
     7501 gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga
     7561 agctagagta agtagttcgc cagttaatag tttgcgcaac gttgttgcca ttgctacagg
     7621 catcgtggtg tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc
     7681 aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc
     7741 gatcgttgtc agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca
     7801 taattctctt actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac
     7861 caagtcattc tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg
     7921 ggataatacc gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc
     7981 ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg
     8041 tgcacccaac tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac
     8101 aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat
     8161 actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata
     8221 catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa
     8281 agtgccacct aaattgtaag cgttaatatt ttgttaaaat tcgcgttaaa tttttgttaa
     8341 atcagctcat tttttaacca ataggccgaa atcggcaaaa tcccttataa atcaaaagaa
     8401 tagaccgaga tagggttgag tgttgttcca gtttggaaca agagtccact attaaagaac
     8461 gtggactcca acgtcaaagg gcgaaaaacc gtctatcagg gcgatggccc actacgtgaa
     8521 ccatcaccct aatcaagttt tttggggtcg aggtgccgta aagcactaaa tcggaaccct
     8581 aaagggagcc cccgatttag agcttgacgg ggaaagccgg cgaacgtggc gagaaaggaa
     8641 gggaagaaag cgaaaggagc gggcgctagg gcgctggcaa gtgtagcggt cacgctgcgc
     8701 gtaaccacca cacccgccgc gcttaatgcg ccgctacagg gcgcgtccca ttcgccattc
     8761 aggctgcgca actgttggga agggcgatcg gtgcgggcct cttcgctatt acgccagctg
     8821 gcgaaagggg gatgtgctgc aaggcgatta agttgggtaa cgccagggtt ttcccagtca
     8881 cgacgttgta aaacgacggc cagtgag
//
