LOCUS       sequence-378566-        6990 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Transient expression of RvCAHS12 (fly codon 
            optimized)-GS_linker-AcGFP1 in Drosophila cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6990)
  AUTHORS   Tanaka A, Nakano T, Watanabe K, Masuda K, Honda G, Kamata S, Yasui 
            R, Kozuka-Hata H, Watanabe C, Chinen T, Kitagawa D, Sawai S, Oyama 
            M, Yanagisawa M, Kunieda T
  TITLE     Stress-dependent cell stiffening by tardigrade tolerance proteins 
            that reversibly form a filamentous network and gel.
  JOURNAL   PLoS Biol. 2022 Sep 6;20(9):e3001780. doi: 
            10.1371/journal.pbio.3001780. eCollection 2022 Sep.
  PUBMED    36067153
REFERENCE   2  (bases 1 to 6990)
  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: "PLoS 
            Biol."; date: "2022-09-6"; volume: "20(9):e3001780. doi"; pages: " 
            10.1371/journal.pbio.3001780. eCollection 2022 Sep"
COMMENT     Sequence Label: pAc5.1/V5-HisA-RvCAHS12-GS-AcGFP1
FEATURES             Location/Qualifiers
     source          1..6990
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        56..2565
                     /label=Ac5 promoter
                     /note="Drosophila melanogaster actin 5C promoter"
     CDS             3547..4266
                     /codon_start=1
                     /product="Aequorea coerulescens GFP"
                     /label=AcGFP1
                     /note="mammalian codon-optimized"
                     /translation="MVSKGAELFTGIVPILIELNGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLSYGVQCFSRYPDHMKQHDFFKSAMPEGYIQERTIFFEDD
                     GNYKSRAEVKFEGDTLVNRIELTGTDFKEDGNILGNKMEYNYNAHNVYIMTDKAKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMIYF
                     GFVTAAAITHGMDELYK"
     CDS             4296..4337
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     CDS             4347..4364
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(4387..4404)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    4437..4571
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4461..4480)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     4515..4534
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     rep_origin      complement(5172..5760)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5252..5271)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5931..6722,6723..6791))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     6554..6573
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6792..6896)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6964..6982
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 cttcaagaat taattctata ttctaaaaac acaaatgata cttctaaaaa aaaatcatga
       61 atggcatcaa ctctgaatca aatctttgca gatgcaccta cttctcattt ccactgtcac
      121 atcatttttc cagatctcgc tgcctgttat gtggcccaca aaccaagaca cgttttatgg
      181 ccattaaagc tggctgatcg tcgccaaaca ccaaatacat aatgaatatg tacacattcg
      241 agaaagaagc gatcaaagaa gcgtcttcgg gcggagtagg agaatgcgga ggagaaggag
      301 aacgagctga tctagtatct ctccacaatc caatgccaac tgaccaactg gccatattcg
      361 gagcaatttg aagccaattt ccatcgcctg gcgatcgctc cattcttggc tatatgtttt
      421 tcaccgttac ccggggccat tttcaaagac tcgtcggcaa gataagattg tgtcactcgc
      481 tgtctctctt catttgtcga agaatgctga ggaatttcgc gatgacgtcg gcgagtattt
      541 tgaagaatga gaataatttg tatttatacg aaaatcagtt agtggaattt tctacaaaaa
      601 catgttatct atagataatt ttgttgcaaa atatgttgac tatgacaaag attgtatgta
      661 tataccttta atgtattctc attttcttat gtatttataa tggcaatgat gatactgatg
      721 atattttaag atgatgccag accaaaaggc ttgaatttct gcgtcttttg ccgaacgcag
      781 tgcatgtgca attgttgttt tttggaatat tcaattttcg gactgtccgc tttgatttca
      841 gtttcttggc ttattcaaaa agcaaagtaa agccaaaaaa gcgagatggc aataccaaat
      901 gcggcaaaac ggtagtggaa ggaaaggggt gcggggcagc ggaaggaagg gtggggcggg
      961 gcgtggcggg gtctgtggct gggcgcgacg tcaccgacgt tggagccact cctttgacca
     1021 tgtgtgcgtg tgtgtattat tcgtgtctcg ccactcgccg gttgtttttt tctttttatg
     1081 ctgcgctctc tctagcgcca tctcgcttac gcatgctcaa cgcaccgcat gttgccgttt
     1141 ccttttatgc gtcattttgg ctcgaaatag gcaattattt aaacaaagat tagtcaacga
     1201 aaacgctaaa ataaataagt ctacaatatg gttacttatt gccatgtgtg tgcagccaac
     1261 gatagcaaca aaagcaacaa cacaggtggc tttccctctt tcactttttg tttgcaagcc
     1321 gcgtgcgagc aagacggcac gaccggcaaa cgcaattacg ctgacaaaga gcagacgaag
     1381 ttttggcgaa aaacatcaag gcgcctgata cgaatgcatt tgcaataaca attgcgatat
     1441 ttaatattgt ttatgaagct gtttgacttc aaaacacaca aaaaaaaaaa taaaacaaat
     1501 tatttgaaag agaattagga atcggacgct tatcgttagg gtaacaacaa gaaatgctta
     1561 ctgagtcaca gcctctggaa aactgccgca agccagagag agagagaaaa agagggagag
     1621 cagcttagac cgcatgtgct tgtgtgtgag gcgtctctct cttcgtctct gttgcgcaaa
     1681 cgcatagact gcactgaaaa aatcgattac ctatttttta tgaatgaata tttgcactat
     1741 tactattcaa aactattaag atagcaatca cattcaatag ccaaatacta taccacctga
     1801 gcgatgcaac gaaatgatca atttgagcaa aaatgctgca tatttaggac ggcatcatta
     1861 tagaaatgct tcttgctgtg tacttttctc tcgtctggca gctgtttcgc cgttattgtt
     1921 aaaaccggct taagttaggt gtgttttcta cgactagtga atgccctact agaagatgtg
     1981 tgttgcacaa aatgtccctg gaataaccaa tttgaagtgc agatagcagt aaacgtaagc
     2041 taatatgaat attatttaac tgtaatgttt taatatcgct ggacattact aataaaccca
     2101 ctataaacac atgtacatat gtatgttttg gcatacaatg agtagttggg gaaaaaatgt
     2161 gtaaaagcac cgtgaccatc acagcataaa gataaccagc tgaagtatcg aatatgagta
     2221 acccccaaat tgaatcacat gccgcaactg ataggaccca tggaagtaca ctcttcatgg
     2281 cgatatacaa gacacacaca agcacgaaca cccagttgcg gaggaaattc tccgtaaatg
     2341 aaaacccaat cggcgaacaa ttcataccca tatatggtaa aagttttgaa cgcgacttga
     2401 gagcggagag cattgcggct gataaggttt tagcgctaag cgggctttat aaaacgggct
     2461 gcgggaccag ttttcatatc actaccgttt gagttcttgt gctgtgtgga tactcctccc
     2521 gacacaaagc cgctccatca gccagcagtc gtctaatcca gagaccccgg atcggggtac
     2581 ctactagtcc agtgtggtgg aattgccacc atgagtcaca cccacgagca aaagttcgag
     2641 cgcgtggaag aacgcaaggt ggacgataag aaaggcctgc aagaagtgcg cgtcggcgtt
     2701 gacacaggac atggcgatcc cgccttgaac ttcaccccaa ccgatgccac actcgtgcgc
     2761 acaggtggtg ttggaggaac aacagccagc agccacagct cccatatgag cagtagtaca
     2821 ggcggagccg tgacaggcgc tagccagtat agtagcacca tgcaccaaga gggcggccac
     2881 atgacaacag aggccagcaa gaacaccagc tacacccaca ccgaagtgcg tgccccagtg
     2941 atcgatacag ccccgccaat tattagcacc ggcgccagtg gaatggccga gcagatagtt
     3001 ggacagggct ttacagccag cgccgctcgc attacaggat ccagcgccga tgtgaacatc
     3061 gtggaaacag ccgaggctcg cgagaagcgc atgcgtgatg aagagaagta cgcccgcgag
     3121 caagaggcca tcaatcgcca cgccgataag gacctggaaa agaaaaccga ggcctaccgc
     3181 aaagaggccg agcacgaggc cgaaaagatc cgcaaggctc tggaaaagca gcacgagcgc
     3241 gatatcgagt tccgcaaaga agtcgtgggc agcaccatcg agaagcagaa ggccgagatt
     3301 gagctggaag ccaagcgtgc caaggccgct ctggaacatg aacgacagct ggccaacgat
     3361 gccctggaac gcagcaagat gcacaccgat gtgcaagtga ccatggatac cgccgctgga
     3421 catacagtgt ccggcggaac aacaatgagc agcagcgagc agcatagcag ctcccacagt
     3481 agtagtcgca ccggcggact gggtggaggc ggttcaggcg gaggtggctc tggcggtggc
     3541 ggatcgatgg tgagcaaggg cgccgagctg ttcaccggca tcgtgcccat cctgatcgag
     3601 ctgaatggcg atgtgaatgg ccacaagttc agcgtgagcg gcgagggcga gggcgatgcc
     3661 acctacggca agctgaccct gaagttcatc tgcaccaccg gcaagctgcc tgtgccctgg
     3721 cccaccctgg tgaccaccct gagctacggc gtgcagtgct tctcacgcta ccccgatcac
     3781 atgaagcagc acgacttctt caagagcgcc atgcctgagg gctacatcca ggagcgcacc
     3841 atcttcttcg aggatgacgg caactacaag tcgcgcgccg aggtgaagtt cgagggcgat
     3901 accctggtga atcgcatcga gctgaccggc accgatttca aggaggatgg caacatcctg
     3961 ggcaataaga tggagtacaa ctacaacgcc cacaatgtgt acatcatgac cgacaaggcc
     4021 aagaatggca tcaaggtgaa cttcaagatc cgccacaaca tcgaggatgg cagcgtgcag
     4081 ctggccgacc actaccagca gaataccccc atcggcgatg gccctgtgct gctgcccgat
     4141 aaccactacc tgtccaccca gagcgccctg tccaaggacc ccaacgagaa gcgcgatcac
     4201 atgatctact tcggcttcgt gaccgccgcc gccatcaccc acggcatgga tgagctgtac
     4261 aagtgaggcc gctcgagtct agagggccct tcgaaggtaa gcctatccct aaccctctcc
     4321 tcggtctcga ttctacgcgt accggtcatc atcaccatca ccattgagtt taaacccgct
     4381 gatcagcctc gactgtgcct tctaaggcct gagctcgctg atcagcctcg atcgaggatc
     4441 cagacatgat aagatacatt gatgagtttg gacaaaccac aactagaatg cagtgaaaaa
     4501 aatgctttat ttgtgaaatt tgtgatgcta ttgctttatt tgtaaccatt ataagctgca
     4561 ataaacaagt taacaacaac aattgcattc attttatgtt tcaggttcag ggggaggtgt
     4621 gggaggtttt ttaaagcaag taaaacctct acaaatgtgg tatggctgat tatgatcagt
     4681 cgaccgatgc ccttgagagc cttcaaccca gtcagctcct tccggtgggc gcggggcatg
     4741 actatcgtcg ccgcacttat gactgtcttc tttatcatgc aactcgtagg acaggtgccg
     4801 gcagcgctct gggtcatttt cggcgaggac cgctttcgct ggagcgcgac gatgatcggc
     4861 ctgtcgcttg cggtattcgg aatcttgcac gccctcgctc aagccttcgt cactggtccc
     4921 gccaccaaac gtttcggcga gaagcaggcc attatcgccg gcatggcggc cgacgcgctg
     4981 ggctacgtct tgctggcgtt cgcgacgcga ggctggatgg ccttccccat tatgattctt
     5041 ctcgcttccg gcggcatcgg gatgcccgcg ttgcaggcca tgctgtccag gcaggtagat
     5101 gacgaccatc agggacagct tcaaggccag caaaaggcca ggaaccgtaa aaaggccgcg
     5161 ttgctggcgt ttttccatag gctccgcccc cctgacgagc atcacaaaaa tcgacgctca
     5221 agtcagaggt ggcgaaaccc gacaggacta taaagatacc aggcgtttcc ccctggaagc
     5281 tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg gatacctgtc cgcctttctc
     5341 ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta ggtatctcag ttcggtgtag
     5401 gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc
     5461 ttatccggta actatcgtct tgagtccaac ccggtaagac acgacttatc gccactggca
     5521 gcagccactg gtaacaggat tagcagagcg aggtatgtag gcggtgctac agagttcttg
     5581 aagtggtggc ctaactacgg ctacactaga aggacagtat ttggtatctg cgctctgctg
     5641 aagccagtta ccttcggaaa aagagttggt agctcttgat ccggcaaaca aaccaccgct
     5701 ggtagcggtg gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa
     5761 gaagatcctt tgatcttttc tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa
     5821 gggattttgg tcatgagatt atcaaaaagg atcttcacct agatcctttt aaattaaaaa
     5881 tgaagtttta aatcaatcta aagtatatat gagtaaactt ggtctgacag ttaccaatgc
     5941 ttaatcagtg aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga
     6001 ctccccgtcg tgtagataac tacgatacgg gagggcttac catctggccc cagtgctgca
     6061 atgataccgc gagacccacg ctcaccggct ccagatttat cagcaataaa ccagccagcc
     6121 ggaagggccg agcgcagaag tggtcctgca actttatccg cctccatcca gtctattaat
     6181 tgttgccggg aagctagagt aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc
     6241 attgctgcag gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt cagctccggt
     6301 tcccaacgat caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc
     6361 ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag tgttatcact catggttatg
     6421 gcagcactgc ataattctct tactgtcatg ccatccgtaa gatgcttttc tgtgactggt
     6481 gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg
     6541 gcgtcaacac gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga
     6601 aaacgttctt cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg
     6661 taacccactc gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg
     6721 tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt
     6781 tgaatactca tactcttcct ttttcaatat tattgaagca tttatcaggg ttattgtctc
     6841 atgagcggat acatatttga atgtatttag aaaaataaac aaataggggt tccgcgcaca
     6901 tttccccgaa aagtgccacc tgacgtctaa gaaaccatta ttatcatgac attaacctat
     6961 aaaaataggc gtatcacgag gccctttcgt
//
