LOCUS       sequence-467877-        5955 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Evolved human-codon optimized PseTnsAB in mammalian cells (for 
            evoCAST).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5955)
  AUTHORS   Witte IP, Lampe GD, Eitzinger S, Miller SM, Berrios KN, McElroy AN, 
            King RT, Stringham OG, Gelsinger DR, Vo PLH, Chen AT, Tolar J, 
            Osborn MJ, Sternberg SH, Liu DR
  TITLE     Programmable gene insertion in human cells with a laboratory-evolved
            CRISPR-associated transposase.
  JOURNAL   Science. 2025 May 15;388(6748):eadt5199. doi: 
            10.1126/science.adt5199. Epub 2025 May 15.
  PUBMED    40373119
REFERENCE   2  (bases 1 to 5955)
  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: "Science."; 
            date: "2025-05-15"; volume: "388(6748):eadt5199. doi"; pages: " 
            10.1126/science.adt5199. Epub 2025 May 15"
COMMENT     Sequence Label: pCMV-evoCAST-evoTnsAB
FEATURES             Location/Qualifiers
     source          1..5955
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(join(171..962,963..1031))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLMDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAILNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     794..813
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1032..1136)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(1215..1234)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        1406..1785
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        1786..1989
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     1940..1960
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     2031..2050
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        2031..2049
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             2738..2758
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             4624..4641
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(4664..4681)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    4670..4894
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(4965..4981)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4965..4981)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4978..5000)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4989..5005
                     /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(5013..5019,5020..5037,5038..5043))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5058..5079
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5196..5213)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5367..5955)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5447..5466)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 gaagatcctt tgatcttttc tacggggtct gacactcagt ggaacgaaaa ctcacgttaa
       61 gggattttgg tcatgagatt atcaaaaagg atcttcacct agatcctttt aaattaaaaa
      121 tgaagtttta aatcaatcta aagtatatat gagtaaactt ggtctgacag ttaccaatgc
      181 ttaatcagtg aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga
      241 ctccccgtcg tgtagataac tacgatacgg gagggcttac catctggccc cagtgctgca
      301 atgataccgc gagacccacg ctcaccggct ccagatttat cagcaataaa ccagccagcc
      361 ggaagggccg agcgcagaag tggtcctgca actttatccg cctccatcca gtctattaat
      421 tgttgccggg aagctagagt aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc
      481 attgctacag gcatcgtggt gtcacgctcg tcgtttagta tggcttcatt cagctccggt
      541 tcccaacgat caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc
      601 ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag tgttatcact catggttatg
      661 gcagcactgc ataattctct tactgtcatg ccatccataa gatgcttttc tgtgactggt
      721 gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg
      781 gcgtcaatac gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga
      841 aaacgttctt cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg
      901 taacccactc gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg
      961 tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt
     1021 tgaatactca tactcttcct ttttcaatat tattgaagca tttatcaggg ttattgtctc
     1081 atgagcggat acatatttga atgtatttag aaaaataaac aaataggggt tccgcgcaca
     1141 tttccccgaa aagtgccacc tgacgtcgac ggatcgggag atcgatctcc cgatccccta
     1201 gggtcgactc tcagtacaat ctgctctgat gccgcatagt taagccagta tctgctccct
     1261 gcttgtgtgt tggaggtcgc tgagtagtgc gcgagcaaaa tttaagctac aacaaggcaa
     1321 ggcttgaccg acaattgcat gaagaatctg cttagggtta ggcgttttgc gctgcttcgc
     1381 gatgtacggg ccagatatac gcgttgacat tgattattga ctagttatta atagtaatca
     1441 attacggggt cattagttca tagcccatat attgagttcc gcgttacata acttacggta
     1501 aatggcccgc ctggctgacc gcccaacgac ccccgcccat tgacgtcaat aatgacgtat
     1561 gttcccatag taacgccaat agggactttc cattgacgtc aatgggtgga gtatttacgg
     1621 taaactgccc acttggcagt acatcaagtg tatcatatgc caagtacgcc ccctattgac
     1681 gtcaatgacg gtaaatggcc cgcctggcat tatgcccagt acatgacctt atgggacttt
     1741 cctacttggc agtacatcta cgtattagtc atcgctatta ccatggtgat gcggttttgg
     1801 cagtacatca atgggcgtgg atagcggttt gactcacggg gatttccaag tctccacccc
     1861 attgacgtca atgggagttt gttttggcac caaaatcaac gggactttcc aaaatgtcgt
     1921 aacaactccg ccccattgac gcaaatgggc ggtaggcgtg tacggtggga ggtctatata
     1981 agcagagctg gtttagtgaa ccgtcagatc cgctagagat ccgcggccgc taatacgact
     2041 cactataggg agagccgcca ccatgtacat cagaaacctg agaaagcctt ccccaaacaa
     2101 gaacgtgttc aagttcgcct ccaccaaggt gtccagcgtg gtgatgtgcg agagcagcct
     2161 ggagttcgac gcctgctttc accacgagta caacgacctt atcgagtctt tcggcagcca
     2221 acctgagggc ttcaaatacg agttcatggg caagtccctg ccttatacac ctgatgccct
     2281 gatcagctac accgataaga cacagaaata ccacgagtac aagacctact ctaagatcgc
     2341 cagccctctg ttccgggctg aatttgccgc caagcgggcc gcttctctga aactgggaat
     2401 cgacctggtc ctggtgaccg acagacagat cagagtgaac cctatcctta acaatctgaa
     2461 gctgctgcat agatacagcg gcgtgtacgg catcagcggc gtccagaagg aactgctgag
     2521 ctttatccac aaaagcggcg ttatcaagct gaatgacatc agcagccagt tggggatccc
     2581 catcggcgaa acccggagct tactgcttgg cctcatgcac aagggcctgg tgaaggccga
     2641 cctgggctgt gatgacctga ccaacaaccc caccctgtgg gccacacctg gcagcggctc
     2701 cggaaaacgg acagccgacg gaagcgagtt cgagtcacca aagaagaagc ggaaagtcgg
     2761 atccggctct ggaggcatga ccgatttctt taacgagttc gacgagagcc tggtgcccct
     2821 gaaacctcag accccaacac agtacgtgaa gctggacgat gccaacctga tccagcggga
     2881 tctggacacc ttttccgata catccaagaa tcaggccctg cagagatata agctgatctc
     2941 caccatcgac aagaaactgt ccaggggctg gacacagcgc aacctggacc ccatcctgga
     3001 cgagctgttc aagggcggcg acgtggtgag gcctaattgg cgcacagtgg cccggtggag
     3061 aaagaagtac atcgagtcta acggcgatat cgccagcctg gccgacaaga atcacaagat
     3121 gggcaacagg accaatcgca tcaagggcga cgataagttc tttgataagg ccctggagcg
     3181 gttcctggac gccaagagac ctacaatcgc caccgcctat cagtactata aggatctgat
     3241 cgtgatcgag aacgagtcca tcgtggaggg caagatccca atcatctctt acaacgcctt
     3301 taataagagg atcaaggcca tcccacctta tgcagtagca gtggctcgcc acggcaagtt
     3361 caaggccgac cagtggtttg catactgcgc agcacacgtg ccaccaacaa gaatcctgga
     3421 gcgggtagag atcgatcaca cacccctgga cctgatccta ctggacgatg agctgctgat
     3481 cccaatcggc aggccctatc tgacactgct gatcgacgtg ttcagcggct gtgtgctggg
     3541 ctttcacctg agctacaaga gcccttccta cgtgagcgct gcaaaggcaa ttacccacgc
     3601 catcaagcca aagtccctgg atgccctgaa catcgagctg cagaatgact ggccctgctt
     3661 cggcaagttt gagaacctgg tagtggacaa tggcgccgag ttctggtcta agaacttgga
     3721 gcacgcctgt cagagcgccg gcatcaacat ccagtacaat cctgtgagga agccctggct
     3781 gaagcccttt atcgagcggt tcttcggcgt gatgaacgag aacttcctaa ccgagctgcc
     3841 tggcaagacc ttttctaata tcctggagaa ggaggagtat aagcccgaga aggacgccat
     3901 catgcggttc agcacatttg tggaggagtt ccacagatgg atcgtcgacg tgtaccacca
     3961 gaactctaat agccgggaga ccagaatccc tatcaagagg tggaagcagg gcttcgatgc
     4021 ctatcctcca ctgaccatga acgaggagga ggagacaagg ttttccatgc tgatgcgcat
     4081 ctccgactct aggaccctga cacgcaatgg ctttaagtac caggagctga tgtatgattc
     4141 taccgccctg gccgactacc gcaagcgcta tccacagaca aaggagaccg tgaagaagtt
     4201 gatcaaggtg gaccccgacg atatcagcaa aatctacgtg tacctggagg agctggagtc
     4261 ctacctggag gtgccttgca cagatccaac cggctacaca gacggcctga gcatctatga
     4321 gcacaagacc atcaagaaga tcaaccggga agagatcaga gagagcaagg actccctggg
     4381 cctggccaag gcaaggatgg ccatccacga gagagtgaag cgggagcagg aggtattcat
     4441 cgagtccaag accaaggcca agatcacagc tgtgaagaag caggcccaga tcgccgatgt
     4501 gagcaatacc ggcacatcca ccatcaaggt gtctgaggag agcgccgcac cagtgctgaa
     4561 gcacatctcc aacgacaatt ctgacgattg ggacgatgac ctggaggcct ttgagtaacc
     4621 ggtcatcatc accatcacca ttgagtttaa acccgctgat cagcctcgac tgtgccttct
     4681 agttgccagc catctgttgt ttgcccctcc cccgtgcctt ccttgaccct ggaaggtgcc
     4741 actcccactg tcctttccta ataaaatgag gaaattgcat cgcattgtct gagtaggtgt
     4801 cattctattc tggggggtgg ggtggggcag gacagcaagg gggaggattg ggaagacaat
     4861 agcaggcatg ctggggatgc ggtgggctct atggcttctg aggcggaaag aaccagctgg
     4921 ggctcgatac cgtcgacctc tagctagagc ttggcgtaat catggtcata gctgtttcct
     4981 gtgtgaaatt gttatccgct cacaattcca cacaacatac gagccggaag cataaagtgt
     5041 aaagcctagg gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg ctcactgccc
     5101 gctttccagt cgggaaacct gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg
     5161 agaggcggtt tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg
     5221 gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg cggtaatacg gttatccaca
     5281 gaatcagggg ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac
     5341 cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac
     5401 aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag ataccaggcg
     5461 tttccccctg gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac
     5521 ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc atagctcacg ctgtaggtat
     5581 ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag
     5641 cccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccggt aagacacgac
     5701 ttatcgccac tggcagcagc cactggtaac aggattagca gagcgaggta tgtaggcggt
     5761 gctacagagt tcttgaagtg gtggcctaac tacggctaca ctagaagaac agtatttggt
     5821 atctgcgctc tgctgaagcc agttaccttc ggaaaaagag ttggtagctc ttgatccggc
     5881 aaacaaacca ccgctggtag cggtggtttt tttgtttgca agcagcagat tacgcgcaga
     5941 aaaaaaggat ctcaa
//
