LOCUS       sequence-179000-        5424 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  To express GFP-tagged MAHS in mammalian cells transiently.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5424)
  AUTHORS   Tanaka S, Tanaka J, Miwa Y, Horikawa DD, Katayama T, Arakawa K, 
            Toyoda A, Kubo T, Kunieda T
  TITLE     Novel mitochondria-targeted heat-soluble proteins identified in the 
            anhydrobiotic Tardigrade improve osmotic tolerance of human cells.
  JOURNAL   PLoS One. 2015 Feb 12;10(2):e0118272. eCollection 2015.
  PUBMED    25675104
REFERENCE   2  (bases 1 to 5424)
  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 One.";
            date: "2015-02-12"; volume: "10(2)"; pages: "e0118272. eCollection 
            2015"
COMMENT     Sequence Label: pAcGFP1-N1-MAHS
FEATURES             Location/Qualifiers
     source          1..5424
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        47..404
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        47..238
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(70..90)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      255..390
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             439..1233
                     /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             439..1233
                     /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(493..512)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     1103..1122
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(1421..1440)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    1465..1512
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      1841..2429
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2330..2349
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     enhancer        2604..2907
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        2908..3111
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     3062..3082
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     CDS             3914..4633
                     /codon_start=1
                     /product="Aequorea coerulescens GFP"
                     /label=AcGFP1
                     /note="mammalian codon-optimized"
                     /translation="MVSKGAELFTGIVPILIELNGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLSYGVQCFSRYPDHMKQHDFFKSAMPEGYIQERTIFFEDD
                     GNYKSRAEVKFEGDTLVNRIELTGTDFKEDGNILGNKMEYNYNAHNVYIMTDKAKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMIYF
                     GFVTAAAITHGMDELYK"
     polyA_signal    4755..4876
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4792..4811)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4846..4865
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(4883..5338)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5020..5041)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     5232..5251
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        join(5365..5424,1..45)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 caataaccct gataaatgct tcaataatat tgaaaaagga agagtcctga ggcggaaaga
       61 accagctgtg gaatgtgtgt cagttagggt gtggaaagtc cccaggctcc ccagcaggca
      121 gaagtatgca aagcatgcat ctcaattagt cagcaaccag gtgtggaaag tccccaggct
      181 ccccagcagg cagaagtatg caaagcatgc atctcaatta gtcagcaacc atagtcccgc
      241 ccctaactcc gcccatcccg cccctaactc cgcccagttc cgcccattct ccgccccatg
      301 gctgactaat tttttttatt tatgcagagg ccgaggccgc ctcggcctct gagctattcc
      361 agaagtagtg aggaggcttt tttggaggcc taggcttttg caaagatcga tcaagagaca
      421 ggatgaggat cgtttcgcat gattgaacaa gatggattgc acgcaggttc tccggccgct
      481 tgggtggaga ggctattcgg ctatgactgg gcacaacaga caatcggctg ctctgatgcc
      541 gccgtgttcc ggctgtcagc gcaggggcgc ccggttcttt ttgtcaagac cgacctgtcc
      601 ggtgccctga atgaactgca agacgaggca gcgcggctat cgtggctggc cacgacgggc
      661 gttccttgcg cagctgtgct cgacgttgtc actgaagcgg gaagggactg gctgctattg
      721 ggcgaagtgc cggggcagga tctcctgtca tctcaccttg ctcctgccga gaaagtatcc
      781 atcatggctg atgcaatgcg gcggctgcat acgcttgatc cggctacctg cccattcgac
      841 caccaagcga aacatcgcat cgagcgagca cgtactcgga tggaagccgg tcttgtcgat
      901 caggatgatc tggacgaaga gcatcagggg ctcgcgccag ccgaactgtt cgccaggctc
      961 aaggcgagca tgcccgacgg cgaggatctc gtcgtgaccc atggcgatgc ctgcttgccg
     1021 aatatcatgg tggaaaatgg ccgcttttct ggattcatcg actgtggccg gctgggtgtg
     1081 gcggaccgct atcaggacat agcgttggct acccgtgata ttgctgaaga gcttggcggc
     1141 gaatgggctg accgcttcct cgtgctttac ggtatcgccg ctcccgattc gcagcgcatc
     1201 gccttctatc gccttcttga cgagttcttc tgagcgggac tctggggttc gaaatgaccg
     1261 accaagcgac gcccaacctg ccatcacgag atttcgattc caccgccgcc ttctatgaaa
     1321 ggttgggctt cggaatcgtt ttccgggacg ccggctggat gatcctccag cgcggggatc
     1381 tcatgctgga gttcttcgcc caccctaggg ggaggctaac tgaaacacgg aaggagacaa
     1441 taccggaagg aacccgcgct atgacggcaa taaaaagaca gaataaaacg cacggtgttg
     1501 ggtcgtttgt tcataaacgc ggggttcggt cccagggctg gcactctgtc gataccccac
     1561 cgagacccca ttggggccaa tacgcccgcg tttcttcctt ttccccaccc caccccccaa
     1621 gttcgggtga aggcccaggg ctcgcagcca acgtcggggc ggcaggccct gccatagcct
     1681 caggttactc atatatactt tagattgatt taaaacttca tttttaattt aaaaggatct
     1741 aggtgaagat cctttttgat aatctcatga ccaaaatccc ttaacgtgag ttttcgttcc
     1801 actgagcgtc agaccccgta gaaaagatca aaggatcttc ttgagatcct ttttttctgc
     1861 gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt tgtttgccgg
     1921 atcaagagct accaactctt tttccgaagg taactggctt cagcagagcg cagataccaa
     1981 atactgttct tctagtgtag ccgtagttag gccaccactt caagaactct gtagcaccgc
     2041 ctacatacct cgctctgcta atcctgttac cagtggctgc tgccagtggc gataagtcgt
     2101 gtcttaccgg gttggactca agacgatagt taccggataa ggcgcagcgg tcgggctgaa
     2161 cggggggttc gtgcacacag cccagcttgg agcgaacgac ctacaccgaa ctgagatacc
     2221 tacagcgtga gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc
     2281 cggtaagcgg cagggtcgga acaggagagc gcacgaggga gcttccaggg ggaaacgcct
     2341 ggtatcttta tagtcctgtc gggtttcgcc acctctgact tgagcgtcga tttttgtgat
     2401 gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa cgcggccttt ttacggttcc
     2461 tggccttttg ctggcctttt gctcacatgt tctttcctgc gttatcccct gattctgtgg
     2521 ataaccgtat taccgccatg cattagttat taatagtaat caattacggg gtcattagtt
     2581 catagcccat atatggagtt ccgcgttaca taacttacgg taaatggccc gcctggctga
     2641 ccgcccaacg acccccgccc attgacgtca ataatgacgt atgttcccat agtaacgcca
     2701 atagggactt tccattgacg tcaatgggtg gagtatttac ggtaaactgc ccacttggca
     2761 gtacatcaag tgtatcatat gccaagtacg ccccctattg acgtcaatga cggtaaatgg
     2821 cccgcctggc attatgccca gtacatgacc ttatgggact ttcctacttg gcagtacatc
     2881 tacgtattag tcatcgctat taccatggtg atgcggtttt ggcagtacat caatgggcgt
     2941 ggatagcggt ttgactcacg gggatttcca agtctccacc ccattgacgt caatgggagt
     3001 ttgttttggc accaaaatca acgggacttt ccaaaatgtc gtaacaactc cgccccattg
     3061 acgcaaatgg gcggtaggcg tgtacggtgg gaggtctata taagcagagc tggtttagtg
     3121 aaccgtcaga tccgctagcg ctaccggact cagatctcga gctcaagctt cgaattctgc
     3181 agtcgacggt accatgtcca gatacctgct gcgcgatgtc caggctgtat tacgcggagt
     3241 tcgcaaagtg gccgagagta gcttaaagct ggagacggag aaagtcagtc tgcggcttgg
     3301 tgactttcgg tcacagcctt cccttcgcag tgtgcctgct tccctcacaa gtcgatcaca
     3361 ggcatttagc ctacaggaga tagctgctcg tgccggagtt gttctgcgag gagtgcaaca
     3421 acagttccga aacgtcactg gagtgaatgc cgctcctgtt gtagcctttg ataatggatc
     3481 agttctatac agtgaaagaa tccactcgca gagttcgcag aagcaggccc cgactacagt
     3541 accaacagga tccgtcagca attcccctca accggaagga aaggcaaacg aagctgctga
     3601 acgcgcaaaa cagtttatga atcctccagt tgcgccaatg gatcctgtcg acaagaatga
     3661 atttgtcgcc atgccggaga tgggtcgtag taatggaaat ggagaaaaca aacaagctgc
     3721 tgatttcatg aaaaaccaag gtgacaccga tatggattcc cagtacgcgc ctgattcatc
     3781 gaagaacacg aaatcggttc ccacgaagga aatcgttgct gaagatggtt cgatgagcat
     3841 tgaggatatc aagaaagcta cgcaggttac tcctggagtt gcagttaaaa acgagggtgt
     3901 tgatccaccg gtcatggtga gcaagggcgc cgagctgttc accggcatcg tgcccatcct
     3961 gatcgagctg aatggcgatg tgaatggcca caagttcagc gtgagcggcg agggcgaggg
     4021 cgatgccacc tacggcaagc tgaccctgaa gttcatctgc accaccggca agctgcctgt
     4081 gccctggccc accctggtga ccaccctgag ctacggcgtg cagtgcttct cacgctaccc
     4141 cgatcacatg aagcagcacg acttcttcaa gagcgccatg cctgagggct acatccagga
     4201 gcgcaccatc ttcttcgagg atgacggcaa ctacaagtcg cgcgccgagg tgaagttcga
     4261 gggcgatacc ctggtgaatc gcatcgagct gaccggcacc gatttcaagg aggatggcaa
     4321 catcctgggc aataagatgg agtacaacta caacgcccac aatgtgtaca tcatgaccga
     4381 caaggccaag aatggcatca aggtgaactt caagatccgc cacaacatcg aggatggcag
     4441 cgtgcagctg gccgaccact accagcagaa tacccccatc ggcgatggcc ctgtgctgct
     4501 gcccgataac cactacctgt ccacccagag cgccctgtcc aaggacccca acgagaagcg
     4561 cgatcacatg atctacttcg gcttcgtgac cgccgccgcc atcacccacg gcatggatga
     4621 gctgtacaag tgagcggccg cgactctaga tcataatcag ccataccaca tttgtagagg
     4681 ttttacttgc tttaaaaaac ctcccacacc tccccctgaa cctgaaacat aaaatgaatg
     4741 caattgttgt tgttaacttg tttattgcag cttataatgg ttacaaataa agcaatagca
     4801 tcacaaattt cacaaataaa gcattttttt cactgcattc tagttgtggt ttgtccaaac
     4861 tcatcaatgt atcttaaggc gtaaattgta agcgttaata ttttgttaaa attcgcgtta
     4921 aatttttgtt aaatcagctc attttttaac caataggccg aaatcggcaa aatcccttat
     4981 aaatcaaaag aatagaccga gatagggttg agtgttgttc cagtttggaa caagagtcca
     5041 ctattaaaga acgtggactc caacgtcaaa gggcgaaaaa ccgtctatca gggcgatggc
     5101 ccactacgtg aaccatcacc ctaatcaagt tttttggggt cgaggtgccg taaagcacta
     5161 aatcggaacc ctaaagggag cccccgattt agagcttgac ggggaaagcc ggcgaacgtg
     5221 gcgagaaagg aagggaagaa agcgaaagga gcgggcgcta gggcgctggc aagtgtagcg
     5281 gtcacgctgc gcgtaaccac cacacccgcc gcgcttaatg cgccgctaca gggcgcgtca
     5341 ggtggcactt ttcggggaaa tgtgcgcgga acccctattt gtttattttt ctaaatacat
     5401 tcaaatatgt atccgctcat gaga
//
