LOCUS       sequence-365643-        4584 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  For the mammalian expression of the tardigrade protein MAHS_RAMVA. 
            This is one of the bottom performing targets in our apoptosis assay 
            (it induced apoptosis)..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4584)
  AUTHORS   Veling MT, Nguyen DT, Thadani NN, Oster ME, Rollins NJ, Brock KP, 
            Bethel NP, Lim S, Baker D, Way JC, Marks DS, Chang RL, Silver PA
  TITLE     Natural and Designed Proteins Inspired by Extremotolerant Organisms 
            Can Form Condensates and Attenuate Apoptosis in Human Cells.
  JOURNAL   ACS Synth Biol. 2022 Mar 18;11(3):1292-1302. doi: 
            10.1021/acssynbio.1c00572. Epub 2022 Feb 18.
  PUBMED    35176859
REFERENCE   2  (bases 1 to 4584)
  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"; doi: 
            "10.1021/acssynbio.1c00572"; journalName: "ACS Synth Biol"; date: 
            "2022-03-18- 18"; volume: "11"; issue: "3"; pages: "1292-1302"
COMMENT     Sequence Label: MAHS_RAMVA_Tanaka pSecTag (pBS0253)
FEATURES             Location/Qualifiers
     source          1..4584
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(13..30)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(184..772)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(264..283)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(943..1734,1735..1803))
                     /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     1566..1585
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1804..1908)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(1983..2002)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        2174..2553
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        2554..2757
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     2708..2728
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     2802..2821
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        2802..2820
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      2837..2846
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2846..2869
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     promoter        3626..3673
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             3692..4066
                     /codon_start=1
                     /gene="Sh ble from Streptoalloteichus hindustanus"
                     /product="antibiotic-binding protein"
                     /label=BleoR
                     /note="confers resistance to bleomycin, phleomycin, and 
                     Zeocin(TM)"
                     /translation="MAKLTSAVPVLTARDVAGAVEFWTDRLGFSRDFVEDDFAGVVRDD
                     VTLFISAVQDQVVPDNTLAWVWVRGLDELYAEWSEVVSTNFRDASGPAMTEIGEQPWGR
                     EFALRDPAGNCVHFVAEEQD"
     polyA_signal    4196..4317
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4233..4252)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4287..4306
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4366..4382)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4366..4382)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4379..4401)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4390..4406
                     /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(4414..4420,4421..4438,4439..4444))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
ORIGIN
        1 ctcttccgct tcctcgctca ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt
       61 atcagctcac tcaaaggcgg taatacggtt atccacagaa tcaggggata acgcaggaaa
      121 gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc
      181 gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct caagtcagag
      241 gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa gctccctcgt
      301 gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc tcccttcggg
      361 aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt aggtcgttcg
      421 ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg ccttatccgg
      481 taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg cagcagccac
      541 tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg
      601 gcctaactac ggctacacta gaagaacagt atttggtatc tgcgctctgc tgaagccagt
      661 taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg
      721 tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc aagaagatcc
      781 tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt aagggatttt
      841 ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa aatgaagttt
      901 taaatcaatc taaagtatat atgagtaaac ttggtctgac agttaccaat gcttaatcag
      961 tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcct gactccccgt
     1021 cgtgtagata actacgatac gggagggctt accatctggc cccagtgctg caatgatacc
     1081 gcgagaccca cgctcaccgg ctccagattt atcagcaata aaccagccag ccggaagggc
     1141 cgagcgcaga agtggtcctg caactttatc cgcctccatc cagtctatta attgttgccg
     1201 ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg ccattgctac
     1261 aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg gttcccaacg
     1321 atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc
     1381 tccgatcgtt gtcagaagta agttggccgc agtgttatca ctcatggtta tggcagcact
     1441 gcataattct cttactgtca tgccatccgt aagatgcttt tctgtgactg gtgagtactc
     1501 aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat
     1561 acgggataat accgcgccac atagcagaac tttaaaagtg ctcatcattg gaaaacgttc
     1621 ttcggggcga aaactctcaa ggatcttacc gctgttgaga tccagttcga tgtaacccac
     1681 tcgtgcaccc aactgatctt cagcatcttt tactttcacc agcgtttctg ggtgagcaaa
     1741 aacaggaagg caaaatgccg caaaaaaggg aataagggcg acacggaaat gttgaatact
     1801 catactcttc ctttttcaat attattgaag catttatcag ggttattgtc tcatgagcgg
     1861 atacatattt gaatgtattt agaaaaataa acaaataggg gttccgcgca catttccccg
     1921 aaaagtgcca cctgacgtcg acggatcggg agatctcccg atcccctatg gtgcactctc
     1981 agtacaatct gctctgatgc cgcatagtta agccagtatc tgctccctgc ttgtgtgttg
     2041 gaggtcgctg agtagtgcgc gagcaaaatt taagctacaa caaggcaagg cttgaccgac
     2101 aattgcatga agaatctgct tagggttagg cgttttgcgc tgcttcgcga tgtacgggcc
     2161 agatatacgc gttgacattg attattgact agttattaat agtaatcaat tacggggtca
     2221 ttagttcata gcccatatat ggagttccgc gttacataac ttacggtaaa tggcccgcct
     2281 ggctgaccgc ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt tcccatagta
     2341 acgccaatag ggactttcca ttgacgtcaa tgggtggagt atttacggta aactgcccac
     2401 ttggcagtac atcaagtgta tcatatgcca agtacgcccc ctattgacgt caatgacggt
     2461 aaatggcccg cctggcatta tgcccagtac atgaccttat gggactttcc tacttggcag
     2521 tacatctacg tattagtcat cgctattacc atggtgatgc ggttttggca gtacatcaat
     2581 gggcgtggat agcggtttga ctcacgggga tttccaagtc tccaccccat tgacgtcaat
     2641 gggagtttgt tttggcacca aaatcaacgg gactttccaa aatgtcgtaa caactccgcc
     2701 ccattgacgc aaatgggcgg taggcgtgta cggtgggagg tctatataag cagagctctc
     2761 tggctaacta gagaacccac tgcttactgg cttatcgaaa ttaatacgac tcactatagg
     2821 gagacccgaa ttcgccgcca ccatggatta caaggatgat gatgataaga tgatgtccag
     2881 atacctgctg cgcgatgtcc aggctgtatt acgcggagtt cgcaaagtgg ccgagagtag
     2941 cttaaagctg gagacggaga aagtcagtct gcggcttggt gactttcggt cacagccttc
     3001 ccttcgcagt gtgcctgctt ccctcacaag tcgatcacag gcatttagcc tacaggagat
     3061 agctgctcgt gccggagttg ttctgcgagg agtgcaacaa cagttccgaa acgtcactgg
     3121 agtgaatgcc gctcctgttg tagcctttga taatggatca gttctataca gtgaaagaat
     3181 ccactcgcag agttcgcaga agcaggcccc gactacagta ccaacaggat ccgtcagcaa
     3241 ttcccctcaa ccggaaggaa aggcaaacga agctgctgaa cgcgcaaaac agtttatgaa
     3301 tcctccagtt gcgccaatgg atcctgtcga caagaatgaa tttgtcgcca tgccggagat
     3361 gggtcgtagt aatggaaatg gagaaaacaa acaagctgct gatttcatga aaaaccaagg
     3421 tgacaccgat atggattccc agtacgcgcc tgattcatcg aagaacacga aatcggttcc
     3481 cacgaaggaa atcgttgctg aagatggttc gatgagcatt gaggatatca agaaagctac
     3541 gcaggttact cctggagttg cagttaaaaa cgagggtgtt taggggtgtt tgtggttttg
     3601 ttgtgtggta ttgtgatcag cacgtgttga caattaatca tcggcatagt atatcggcat
     3661 agtataatac gacaaggtga ggaactaaac catggccaag ttgaccagtg ccgttccggt
     3721 gctcaccgcg cgcgacgtcg ccggagcggt cgagttctgg accgaccggc tcgggttctc
     3781 ccgggacttc gtggaggacg acttcgccgg tgtggtccgg gacgacgtga ccctgttcat
     3841 cagcgcggtc caggaccagg tggtgccgga caacaccctg gcctgggtgt gggtgcgcgg
     3901 cctggacgag ctgtacgccg agtggtcgga ggtcgtgtcc acgaacttcc gggacgcctc
     3961 cgggccggcc atgaccgaga tcggcgagca gccgtggggg cgggagttcg ccctgcgcga
     4021 cccggccggc aactgcgtgc acttcgtggc cgaggagcag gactgacacg tgctacgaga
     4081 tttcgattcc accgccgcct tctatgaaag gttgggcttc ggaatcgttt tccgggacgc
     4141 cggctggatg atcctccagc gcggggatct catgctggag ttcttcgccc accccaactt
     4201 gtttattgca gcttataatg gttacaaata aagcaatagc atcacaaatt tcacaaataa
     4261 agcatttttt tcactgcatt ctagttgtgg tttgtccaaa ctcatcaatg tatcttatca
     4321 tgtctgtata ccgtcgacct ctagctagag cttggcgtaa tcatggtcat agctgtttcc
     4381 tgtgtgaaat tgttatccgc tcacaattcc acacaacata cgagccggaa gcataaagtg
     4441 taaagcctgg ggtgcctaat gagtgagcta acttacatta attgcgttgc gctcactgcc
     4501 cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg
     4561 gagaggcggt ttgcgtattg ggcg
//
