LOCUS       sequence-378558-        5169 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of RvCAHS12-min-GS_linker-AcGFP1 in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5169)
  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 5169)
  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: pAcGFP1-N1-RvCAHS12-min-GS
FEATURES             Location/Qualifiers
     source          1..5169
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        61..364
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        365..568
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     519..539
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     CDS             1116..1835
                     /codon_start=1
                     /product="Aequorea coerulescens GFP"
                     /label=AcGFP1
                     /note="mammalian codon-optimized"
                     /translation="MVSKGAELFTGIVPILIELNGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLSYGVQCFSRYPDHMKQHDFFKSAMPEGYIQERTIFFEDD
                     GNYKSRAEVKFEGDTLVNRIELTGTDFKEDGNILGNKMEYNYNAHNVYIMTDKAKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMIYF
                     GFVTAAAITHGMDELYK"
     polyA_signal    1957..2078
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1994..2013)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2048..2067
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2085..2540)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2222..2243)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2434..2453
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2567..2671
                     /gene="bla"
                     /label=AmpR promoter
     promoter        2673..3030
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2673..2864
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2696..2716)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      2881..3016
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3065..3859
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             3065..3859
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(3119..3138)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3729..3748
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4047..4066)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4091..4138
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4467..5055
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4956..4975
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg
       61 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
      121 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      181 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      241 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      301 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      361 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
      421 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
      481 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
      541 acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatcc gccaccatgt
      601 ctggcatggc cgaacagatc gtcggtcaag gtttcacggc ctctgcagct cgtatcactg
      661 gctcgtcggc tgatgtcaac atcgttgaga ctgctgaggc tcgcgagaag cgaatgaggg
      721 atgaggagaa atatgctcgt gaacaggagg ccatcaaccg tcatgctgac aaggacctgg
      781 aaaagaagac ggaagcctac cgcaaggagg ccgagcacga ggctgagaag atccgcaagg
      841 cgttggagaa gcaacacgag cgagacatcg agttccgcaa ggaagttgtg ggaagtacca
      901 tcgagaagca gaaggcggaa atcgagctgg aagccaagcg agcaaaggcc gctttggagc
      961 acgaacgtca gctggccaac gatgcgttgg agcgttctaa gatgcatacc gatgtccaag
     1021 tcacgatgga cactgcagcc ggacataccg tcagcggtgg aaccacgatg ggtggaggcg
     1081 gttcaggcgg aggtggctct ggcggtggcg gatcgatggt gagcaagggc gccgagctgt
     1141 tcaccggcat cgtgcccatc ctgatcgagc tgaatggcga tgtgaatggc cacaagttca
     1201 gcgtgagcgg cgagggcgag ggcgatgcca cctacggcaa gctgaccctg aagttcatct
     1261 gcaccaccgg caagctgcct gtgccctggc ccaccctggt gaccaccctg agctacggcg
     1321 tgcagtgctt ctcacgctac cccgatcaca tgaagcagca cgacttcttc aagagcgcca
     1381 tgcctgaggg ctacatccag gagcgcacca tcttcttcga ggatgacggc aactacaagt
     1441 cgcgcgccga ggtgaagttc gagggcgata ccctggtgaa tcgcatcgag ctgaccggca
     1501 ccgatttcaa ggaggatggc aacatcctgg gcaataagat ggagtacaac tacaacgccc
     1561 acaatgtgta catcatgacc gacaaggcca agaatggcat caaggtgaac ttcaagatcc
     1621 gccacaacat cgaggatggc agcgtgcagc tggccgacca ctaccagcag aataccccca
     1681 tcggcgatgg ccctgtgctg ctgcccgata accactacct gtccacccag agcgccctgt
     1741 ccaaggaccc caacgagaag cgcgatcaca tgatctactt cggcttcgtg accgccgccg
     1801 ccatcaccca cggcatggat gagctgtaca agtgagcggc cgcgactcta gatcataatc
     1861 agccatacca catttgtaga ggttttactt gctttaaaaa acctcccaca cctccccctg
     1921 aacctgaaac ataaaatgaa tgcaattgtt gttgttaact tgtttattgc agcttataat
     1981 ggttacaaat aaagcaatag catcacaaat ttcacaaata aagcattttt ttcactgcat
     2041 tctagttgtg gtttgtccaa actcatcaat gtatcttaag gcgtaaattg taagcgttaa
     2101 tattttgtta aaattcgcgt taaatttttg ttaaatcagc tcatttttta accaataggc
     2161 cgaaatcggc aaaatccctt ataaatcaaa agaatagacc gagatagggt tgagtgttgt
     2221 tccagtttgg aacaagagtc cactattaaa gaacgtggac tccaacgtca aagggcgaaa
     2281 aaccgtctat cagggcgatg gcccactacg tgaaccatca ccctaatcaa gttttttggg
     2341 gtcgaggtgc cgtaaagcac taaatcggaa ccctaaaggg agcccccgat ttagagcttg
     2401 acggggaaag ccggcgaacg tggcgagaaa ggaagggaag aaagcgaaag gagcgggcgc
     2461 tagggcgctg gcaagtgtag cggtcacgct gcgcgtaacc accacacccg ccgcgcttaa
     2521 tgcgccgcta cagggcgcgt caggtggcac ttttcgggga aatgtgcgcg gaacccctat
     2581 ttgtttattt ttctaaatac attcaaatat gtatccgctc atgagacaat aaccctgata
     2641 aatgcttcaa taatattgaa aaaggaagag tcctgaggcg gaaagaacca gctgtggaat
     2701 gtgtgtcagt tagggtgtgg aaagtcccca ggctccccag caggcagaag tatgcaaagc
     2761 atgcatctca attagtcagc aaccaggtgt ggaaagtccc caggctcccc agcaggcaga
     2821 agtatgcaaa gcatgcatct caattagtca gcaaccatag tcccgcccct aactccgccc
     2881 atcccgcccc taactccgcc cagttccgcc cattctccgc cccatggctg actaattttt
     2941 tttatttatg cagaggccga ggccgcctcg gcctctgagc tattccagaa gtagtgagga
     3001 ggcttttttg gaggcctagg cttttgcaaa gatcgatcaa gagacaggat gaggatcgtt
     3061 tcgcatgatt gaacaagatg gattgcacgc aggttctccg gccgcttggg tggagaggct
     3121 attcggctat gactgggcac aacagacaat cggctgctct gatgccgccg tgttccggct
     3181 gtcagcgcag gggcgcccgg ttctttttgt caagaccgac ctgtccggtg ccctgaatga
     3241 actgcaagac gaggcagcgc ggctatcgtg gctggccacg acgggcgttc cttgcgcagc
     3301 tgtgctcgac gttgtcactg aagcgggaag ggactggctg ctattgggcg aagtgccggg
     3361 gcaggatctc ctgtcatctc accttgctcc tgccgagaaa gtatccatca tggctgatgc
     3421 aatgcggcgg ctgcatacgc ttgatccggc tacctgccca ttcgaccacc aagcgaaaca
     3481 tcgcatcgag cgagcacgta ctcggatgga agccggtctt gtcgatcagg atgatctgga
     3541 cgaagagcat caggggctcg cgccagccga actgttcgcc aggctcaagg cgagcatgcc
     3601 cgacggcgag gatctcgtcg tgacccatgg cgatgcctgc ttgccgaata tcatggtgga
     3661 aaatggccgc ttttctggat tcatcgactg tggccggctg ggtgtggcgg accgctatca
     3721 ggacatagcg ttggctaccc gtgatattgc tgaagagctt ggcggcgaat gggctgaccg
     3781 cttcctcgtg ctttacggta tcgccgctcc cgattcgcag cgcatcgcct tctatcgcct
     3841 tcttgacgag ttcttctgag cgggactctg gggttcaaaa tgaccgacca agcgacgccc
     3901 aacctgccat cacgagattt cgattccacc gccgccttct atgaaaggtt gggcttcgga
     3961 atcgttttcc gggacgccgg ctggatgatc ctccagcgcg gggatctcat gctggagttc
     4021 ttcgcccacc ctagggggag gctaactgaa acacggaagg agacaatacc ggaaggaacc
     4081 cgcgctatga cggcaataaa aagacagaat aaaacgcacg gtgttgggtc gtttgttcat
     4141 aaacgcgggg ttcggtccca gggctggcac tctgtcgata ccccaccgag accccattgg
     4201 ggccaatacg cccgcgtttc ttccttttcc ccaccccacc ccccaagttc gggtgaaggc
     4261 ccagggctcg cagccaacgt cggggcggca ggccctgcca tagcctcagg ttactcatat
     4321 atactttaga ttgatttaaa acttcatttt taatttaaaa ggatctaggt gaagatcctt
     4381 tttgataatc tcatgaccaa aatcccttaa cgtgagtttt cgttccactg agcgtcagac
     4441 cccgtagaaa agatcaaagg atcttcttga gatccttttt ttctgcgcgt aatctgctgc
     4501 ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt tgccggatca agagctacca
     4561 actctttttc cgaaggtaac tggcttcagc agagcgcaga taccaaatac tgttcttcta
     4621 gtgtagccgt agttaggcca ccacttcaag aactctgtag caccgcctac atacctcgct
     4681 ctgctaatcc tgttaccagt ggctgctgcc agtggcgata agtcgtgtct taccgggttg
     4741 gactcaagac gatagttacc ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc
     4801 acacagccca gcttggagcg aacgacctac accgaactga gatacctaca gcgtgagcta
     4861 tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca ggtatccggt aagcggcagg
     4921 gtcggaacag gagagcgcac gagggagctt ccagggggaa acgcctggta tctttatagt
     4981 cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg
     5041 cggagcctat ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc cttttgctgg
     5101 ccttttgctc acatgttctt tcctgcgtta tcccctgatt ctgtggataa ccgtattacc
     5161 gccatgcat
//
