LOCUS       sequence-366644-        5660 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  For the mammalian expression of the tardigrade protein CAHS1_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 5660)
  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 5660)
  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: pST_CAHS1_RAMVA (pBS0267)
FEATURES             Location/Qualifiers
     source          1..5660
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(64..83)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        255..634
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        635..838
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     789..809
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     883..902
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        883..901
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             1648..1671
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     primer_bind     complement(1688..1705)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1694..1918
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      1964..2392
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2051..2070)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2261..2282
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2401..2421)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2406..2735
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2586..2721
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     promoter        2783..2830
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             2849..3223
                     /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    3353..3474
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3390..3409)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3444..3463
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3523..3539)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3523..3539)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3536..3558)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3547..3563
                     /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(3571..3577,3578..3595,3596..3601))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     primer_bind     complement(3754..3771)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3925..4513)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4005..4024)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4684..5475,5476..5544))
                     /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     5307..5326
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5545..5649)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gaaaagtgcc acctgacgtc gacggatcgg gagatctccc gatcccctat ggtgcactct
       61 cagtacaatc tgctctgatg ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt
      121 ggaggtcgct gagtagtgcg cgagcaaaat ttaagctaca acaaggcaag gcttgaccga
      181 caattgcatg aagaatctgc ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc
      241 cagatatacg cgttgacatt gattattgac tagttattaa tagtaatcaa ttacggggtc
      301 attagttcat agcccatata tggagttccg cgttacataa cttacggtaa atggcccgcc
      361 tggctgaccg cccaacgacc cccgcccatt gacgtcaata atgacgtatg ttcccatagt
      421 aacgccaata gggactttcc attgacgtca atgggtggag tatttacggt aaactgccca
      481 cttggcagta catcaagtgt atcatatgcc aagtacgccc cctattgacg tcaatgacgg
      541 taaatggccc gcctggcatt atgcccagta catgacctta tgggactttc ctacttggca
      601 gtacatctac gtattagtca tcgctattac catggtgatg cggttttggc agtacatcaa
      661 tgggcgtgga tagcggtttg actcacgggg atttccaagt ctccacccca ttgacgtcaa
      721 tgggagtttg ttttggcacc aaaatcaacg ggactttcca aaatgtcgta acaactccgc
      781 cccattgacg caaatgggcg gtaggcgtgt acggtgggag gtctatataa gcagagctct
      841 ctggctaact agagaaccca ctgcttactg gcttatcgaa attaatacga ctcactatag
      901 ggagacccaa gctggctagc caccatgccc tacgaaaagc acgtggagca gacggtcgtg
      961 gagaaaaccg agcagccagg gcactccagt acccaccacg cccccgctca gcggaccgtg
     1021 gcccgggaac aggaggaagt ggtgcacaag gagtttaccc acactgatat ccgggtacca
     1081 cacattgacg ctcccccccc cattattgca gccagcgccg tggggctggc cgaggaaatc
     1141 gttagccacg ggttccaggc cagcgctgct cggatcagcg gagcctccac cgaggtcgat
     1201 atgagaccta gccctaagct ggccgaagag gccagacggg acgccgagcg gtatcagaag
     1261 gagcacgaga tgatcaacag acaggccgaa gccacactgc agaaaaaggc cgaggagtac
     1321 cggcaccaga ccgaggccga ggccgagaaa atccgcagag agttggagaa gcagcacgag
     1381 agggatattc agttcagaaa ggacctgatt gatcagacta tcgagaagca gaagagagaa
     1441 gtggatctgg aggctaaaat ggccaagcgg gagttagaca gagaagctca gttggcaaaa
     1501 gaggccttag aacgtagtcg aatggcaact aacgttgaag tcacgttgga taccgccgca
     1561 ggacacacgg tcagcggtgg gacaactgtc tcaagtgtgg acaaggtcga aacggtccgt
     1621 gagcggaaac atcattaagg atctggggac tacaaggacg acgacgacaa gtgaacccgc
     1681 tgatcagcct cgactgtgcc ttctagttgc cagccatctg ttgtttgccc ctcccccgtg
     1741 ccttccttga ccctggaagg tgccactccc actgtccttt cctaataaaa tgaggaaatt
     1801 gcatcgcatt gtctgagtag gtgtcattct attctggggg gtggggtggg gcaggacagc
     1861 aagggggagg attgggaaga caatagcagg catgctgggg atgcggtggg ctctatggct
     1921 tctgaggcgg aaagaaccag ctggggctct agggggtatc cccacgcgcc ctgtagcggc
     1981 gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga ccgctacact tgccagcgcc
     2041 ctagcgcccg ctcctttcgc tttcttccct tcctttctcg ccacgttcgc cggctttccc
     2101 cgtcaagctc taaatcgggg gctcccttta gggttccgat ttagtgcttt acggcacctc
     2161 gaccccaaaa aacttgatta gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg
     2221 gtttttcgcc ctttgacgtt ggagtccacg ttctttaata gtggactctt gttccaaact
     2281 ggaacaacac tcaaccctat ctcggtctat tcttttgatt tataagggat tttgccgatt
     2341 tcggcctatt ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa ttaattctgt
     2401 ggaatgtgtg tcagttaggg tgtggaaagt ccccaggctc cccagcaggc agaagtatgc
     2461 aaagcatgca tctcaattag tcagcaacca ggtgtggaaa gtccccaggc tccccagcag
     2521 gcagaagtat gcaaagcatg catctcaatt agtcagcaac catagtcccg cccctaactc
     2581 cgcccatccc gcccctaact ccgcccagtt ccgcccattc tccgccccat ggctgactaa
     2641 ttttttttat ttatgcagag gccgaggccg cctctgcctc tgagctattc cagaagtagt
     2701 gaggaggctt ttttggaggc ctaggctttt gcaaaaagct cccgggagct tgtatatcca
     2761 ttttcggatc tgatcagcac gtgttgacaa ttaatcatcg gcatagtata tcggcatagt
     2821 ataatacgac aaggtgagga actaaaccat ggccaagttg accagtgccg ttccggtgct
     2881 caccgcgcgc gacgtcgccg gagcggtcga gttctggacc gaccggctcg ggttctcccg
     2941 ggacttcgtg gaggacgact tcgccggtgt ggtccgggac gacgtgaccc tgttcatcag
     3001 cgcggtccag gaccaggtgg tgccggacaa caccctggcc tgggtgtggg tgcgcggcct
     3061 ggacgagctg tacgccgagt ggtcggaggt cgtgtccacg aacttccggg acgcctccgg
     3121 gccggccatg accgagatcg gcgagcagcc gtgggggcgg gagttcgccc tgcgcgaccc
     3181 ggccggcaac tgcgtgcact tcgtggccga ggagcaggac tgacacgtgc tacgagattt
     3241 cgattccacc gccgccttct atgaaaggtt gggcttcgga atcgttttcc gggacgccgg
     3301 ctggatgatc ctccagcgcg gggatctcat gctggagttc ttcgcccacc ccaacttgtt
     3361 tattgcagct tataatggtt acaaataaag caatagcatc acaaatttca caaataaagc
     3421 atttttttca ctgcattcta gttgtggttt gtccaaactc atcaatgtat cttatcatgt
     3481 ctgtataccg tcgacctcta gctagagctt ggcgtaatca tggtcatagc tgtttcctgt
     3541 gtgaaattgt tatccgctca caattccaca caacatacga gccggaagca taaagtgtaa
     3601 agcctggggt gcctaatgag tgagctaact tacattaatt gcgttgcgct cactgcccgc
     3661 tttccagtcg ggaaacctgt cgtgccagct gcattaatga atcggccaac gcgcggggag
     3721 aggcggtttg cgtattgggc gctcttccgc ttcctcgctc actgactcgc tgcgctcggt
     3781 cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt tatccacaga
     3841 atcaggggat aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg
     3901 taaaaaggcc gcgttgctgg cgtttttcca taggctccgc ccccctgacg agcatcacaa
     3961 aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga ctataaagat accaggcgtt
     4021 tccccctgga agctccctcg tgcgctctcc tgttccgacc ctgccgctta ccggatacct
     4081 gtccgccttt ctcccttcgg gaagcgtggc gctttctcat agctcacgct gtaggtatct
     4141 cagttcggtg taggtcgttc gctccaagct gggctgtgtg cacgaacccc ccgttcagcc
     4201 cgaccgctgc gccttatccg gtaactatcg tcttgagtcc aacccggtaa gacacgactt
     4261 atcgccactg gcagcagcca ctggtaacag gattagcaga gcgaggtatg taggcggtgc
     4321 tacagagttc ttgaagtggt ggcctaacta cggctacact agaagaacag tatttggtat
     4381 ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa
     4441 acaaaccacc gctggtagcg gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa
     4501 aaaaggatct caagaagatc ctttgatctt ttctacgggg tctgacgctc agtggaacga
     4561 aaactcacgt taagggattt tggtcatgag attatcaaaa aggatcttca cctagatcct
     4621 tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa cttggtctga
     4681 cagttaccaa tgcttaatca gtgaggcacc tatctcagcg atctgtctat ttcgttcatc
     4741 catagttgcc tgactccccg tcgtgtagat aactacgata cgggagggct taccatctgg
     4801 ccccagtgct gcaatgatac cgcgagaccc acgctcaccg gctccagatt tatcagcaat
     4861 aaaccagcca gccggaaggg ccgagcgcag aagtggtcct gcaactttat ccgcctccat
     4921 ccagtctatt aattgttgcc gggaagctag agtaagtagt tcgccagtta atagtttgcg
     4981 caacgttgtt gccattgcta caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc
     5041 attcagctcc ggttcccaac gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa
     5101 agcggttagc tccttcggtc ctccgatcgt tgtcagaagt aagttggccg cagtgttatc
     5161 actcatggtt atggcagcac tgcataattc tcttactgtc atgccatccg taagatgctt
     5221 ttctgtgact ggtgagtact caaccaagtc attctgagaa tagtgtatgc ggcgaccgag
     5281 ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca catagcagaa ctttaaaagt
     5341 gctcatcatt ggaaaacgtt cttcggggcg aaaactctca aggatcttac cgctgttgag
     5401 atccagttcg atgtaaccca ctcgtgcacc caactgatct tcagcatctt ttactttcac
     5461 cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc
     5521 gacacggaaa tgttgaatac tcatactctt cctttttcaa tattattgaa gcatttatca
     5581 gggttattgt ctcatgagcg gatacatatt tgaatgtatt tagaaaaata aacaaatagg
     5641 ggttccgcgc acatttcccc
//
