LOCUS       sequence-366748-        7268 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  For the mammalian expression of the C. elegans protein H2FLK4_CAEEL.
            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 7268)
  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 7268)
  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_H2FLK4_CAEEL (pBS0314)
FEATURES             Location/Qualifiers
     source          1..7268
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(45..64)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        236..615
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        616..819
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     770..790
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     864..883
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        864..882
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             3237..3260
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     primer_bind     complement(3277..3294)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3283..3507
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3553..3981
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3640..3659)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3850..3871
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3990..4010)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3995..4324
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4175..4310
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4237..4256
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        4372..4419
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             4438..4812
                     /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    4942..5063
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4979..4998)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5033..5052
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5112..5128)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5112..5128)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5125..5147)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5136..5152
                     /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(5160..5166,5167..5184,5185..5190))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     primer_bind     complement(5343..5360)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5514..6102)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5594..5613)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6273..7064,7065..7133))
                     /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     6896..6915
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7134..7238)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 cgacggatcg ggagatctcc cgatccccta tggtgcactc tcagtacaat ctgctctgat
       61 gccgcatagt taagccagta tctgctccct gcttgtgtgt tggaggtcgc tgagtagtgc
      121 gcgagcaaaa tttaagctac aacaaggcaa ggcttgaccg acaattgcat gaagaatctg
      181 cttagggtta ggcgttttgc gctgcttcgc gatgtacggg ccagatatac gcgttgacat
      241 tgattattga ctagttatta atagtaatca attacggggt cattagttca tagcccatat
      301 atggagttcc gcgttacata acttacggta aatggcccgc ctggctgacc gcccaacgac
      361 ccccgcccat tgacgtcaat aatgacgtat gttcccatag taacgccaat agggactttc
      421 cattgacgtc aatgggtgga gtatttacgg taaactgccc acttggcagt acatcaagtg
      481 tatcatatgc caagtacgcc ccctattgac gtcaatgacg gtaaatggcc cgcctggcat
      541 tatgcccagt acatgacctt atgggacttt cctacttggc agtacatcta cgtattagtc
      601 atcgctatta ccatggtgat gcggttttgg cagtacatca atgggcgtgg atagcggttt
      661 gactcacggg gatttccaag tctccacccc attgacgtca atgggagttt gttttggcac
      721 caaaatcaac gggactttcc aaaatgtcgt aacaactccg ccccattgac gcaaatgggc
      781 ggtaggcgtg tacggtggga ggtctatata agcagagctc tctggctaac tagagaaccc
      841 actgcttact ggcttatcga aattaatacg actcactata gggagaccca agctggctag
      901 ccaccatgtt gagtaatccg aatttgaaac ggctggtaat gaaccaatgg cggcccgcac
      961 aaatattgtt gctggaacaa ccggggccaa gcagaagatt tgttgaaaac actacgcaca
     1021 acaaaatgag ctttatggat aaggtcaaag acacggctaa tagtgcatat gaaagtgttg
     1081 agaacacagc tgttggcgcc aaagatgggg tcgttgtagc tttcaacact gtaggagaga
     1141 aaattggtgg ggcgtacgac tccacgaaac aatatgcgtc tgataaagtc agtgcagccg
     1201 gcgattacgc aagtgaagct gggaagaaag tgggcgatgc agaagactct atgatacaag
     1261 gggcttatga tgccaaggat tccgctaaag aatctgccag taatgcaggc aataagatta
     1321 gcaatgcctt tacaagcctc aaagatagca ttaccggcga taatgctcat aaaaccggtg
     1381 aagcgcttaa tgataactat gaatcagtga aggaaagcgc gagcgaagca gccgatcgac
     1441 tcaaggaacc ggcccaaaac accgctgata atgcaaagga tctggccaat caaaccgggg
     1501 aagaaatgtc cgacgccttt gaaagcgtca agcaacttac atccgataca gcagaaaatg
     1561 caaaggagaa tgcacatgat gcggccgata ctgccaagag ttataaagat aaggcgggag
     1621 aaaaggcttc aggtattctc gactccttta aagcgcacgc ttcagatgcc aaagacagcg
     1681 tcgaaaataa ggctagcgat gcttataaca gcgccaaaga taaagcggga gatgcatggg
     1741 attctactaa agatgcagcg ggtgatgcca aagacaaaac caaatccttt accgaaaagg
     1801 ttggggacaa aataagtgga gcatatgatt ctgttaaaga gaaagccagc gacgtcgcag
     1861 attctttcaa ggcccatagt accgactcaa aagataatgt cgaaaacaaa gcagccgatg
     1921 cttacaacac ggctaaagat aaggcttccg atgcttggga caaaactaaa gataaggctg
     1981 gtgaagcaaa agacaagatg ggtgatgcgt gggatactac gaaagataaa gcgggtgatg
     2041 cctgggacac aacgaaagat aaagctgggg atggaaaagg aaaagccgga gatgcatggg
     2101 atactactaa agacaaggca agcgatgcct gggatacaac taaagataaa gccggcgaag
     2161 caaaggacaa aatgggtgaa gcatgggatc acactaaaga caaagccggt gaagctaaag
     2221 ataaagcgag cgatgccgct gacgatgccc agggcaaaag caaatcaatg actgagaaga
     2281 tcggggacag catctcaggc gcttgggagt ctaccaaaga cacagctgtt tcagccaaag
     2341 ataaaaccgc ggaagccgca ggtaaggtgg gagatagtat ctccggcgcg tatgatacgg
     2401 ttaaggaaaa ggccagcgac attgctgata gcttcaaagc tcatagcaca gattctaaag
     2461 ataatgttga aaataaagcc gcggatgcct ataactccgc taaggataaa gcttccgatg
     2521 cttgggataa gactaaagat aaggctggcg atgcaaaaga taaagccgcg gatgcctggg
     2581 ataccacaaa agacaaggcg ggcgatgcgt gggactccac taaggatcat gcagcagatg
     2641 ctaaggataa ggcaagtgat gccgcgggag atgccaaaga taaaagcaag tctctcacag
     2701 agaaagctgg ggatgcgatc tctggagcat acgatagtgt caaagagaaa gcttctgaca
     2761 ttgcagattc ctttaaagct cacagcacca acagcaaaga caatgttgaa aataaagctt
     2821 cagacgctta taactccgcg aaggataaag catcagacgc ctgggataaa actaaagata
     2881 aagcaggtga agctaaagat aaagctgggg atgcttggga caatacaaaa gataaagccg
     2941 ggaatgcctg ggattctact aaagacaaag catctgatgc gtgggataca acaaaagaca
     3001 aagcttctga tgccaaggaa agcgcgggag atgcagcaga ttcagccaag gataaatcaa
     3061 agtcaattac agaaaccatt ggcgataaaa ttagcggagc atatgatagc gtcaaagaaa
     3121 aggcgtcaga tgtggcagat tccttcaaag cacacagcag cgacgcgcaa gataccgtcg
     3181 aagtgtccca tcataaatat tttaatgttt tctccagctt cgaataagga tctggggact
     3241 acaaggacga cgacgacaag tgaacccgct gatcagcctc gactgtgcct tctagttgcc
     3301 agccatctgt tgtttgcccc tcccccgtgc cttccttgac cctggaaggt gccactccca
     3361 ctgtcctttc ctaataaaat gaggaaattg catcgcattg tctgagtagg tgtcattcta
     3421 ttctgggggg tggggtgggg caggacagca agggggagga ttgggaagac aatagcaggc
     3481 atgctgggga tgcggtgggc tctatggctt ctgaggcgga aagaaccagc tggggctcta
     3541 gggggtatcc ccacgcgccc tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc
     3601 gcagcgtgac cgctacactt gccagcgccc tagcgcccgc tcctttcgct ttcttccctt
     3661 cctttctcgc cacgttcgcc ggctttcccc gtcaagctct aaatcggggg ctccctttag
     3721 ggttccgatt tagtgcttta cggcacctcg accccaaaaa acttgattag ggtgatggtt
     3781 cacgtagtgg gccatcgccc tgatagacgg tttttcgccc tttgacgttg gagtccacgt
     3841 tctttaatag tggactcttg ttccaaactg gaacaacact caaccctatc tcggtctatt
     3901 cttttgattt ataagggatt ttgccgattt cggcctattg gttaaaaaat gagctgattt
     3961 aacaaaaatt taacgcgaat taattctgtg gaatgtgtgt cagttagggt gtggaaagtc
     4021 cccaggctcc ccagcaggca gaagtatgca aagcatgcat ctcaattagt cagcaaccag
     4081 gtgtggaaag tccccaggct ccccagcagg cagaagtatg caaagcatgc atctcaatta
     4141 gtcagcaacc atagtcccgc ccctaactcc gcccatcccg cccctaactc cgcccagttc
     4201 cgcccattct ccgccccatg gctgactaat tttttttatt tatgcagagg ccgaggccgc
     4261 ctctgcctct gagctattcc agaagtagtg aggaggcttt tttggaggcc taggcttttg
     4321 caaaaagctc ccgggagctt gtatatccat tttcggatct gatcagcacg tgttgacaat
     4381 taatcatcgg catagtatat cggcatagta taatacgaca aggtgaggaa ctaaaccatg
     4441 gccaagttga ccagtgccgt tccggtgctc accgcgcgcg acgtcgccgg agcggtcgag
     4501 ttctggaccg accggctcgg gttctcccgg gacttcgtgg aggacgactt cgccggtgtg
     4561 gtccgggacg acgtgaccct gttcatcagc gcggtccagg accaggtggt gccggacaac
     4621 accctggcct gggtgtgggt gcgcggcctg gacgagctgt acgccgagtg gtcggaggtc
     4681 gtgtccacga acttccggga cgcctccggg ccggccatga ccgagatcgg cgagcagccg
     4741 tgggggcggg agttcgccct gcgcgacccg gccggcaact gcgtgcactt cgtggccgag
     4801 gagcaggact gacacgtgct acgagatttc gattccaccg ccgccttcta tgaaaggttg
     4861 ggcttcggaa tcgttttccg ggacgccggc tggatgatcc tccagcgcgg ggatctcatg
     4921 ctggagttct tcgcccaccc caacttgttt attgcagctt ataatggtta caaataaagc
     4981 aatagcatca caaatttcac aaataaagca tttttttcac tgcattctag ttgtggtttg
     5041 tccaaactca tcaatgtatc ttatcatgtc tgtataccgt cgacctctag ctagagcttg
     5101 gcgtaatcat ggtcatagct gtttcctgtg tgaaattgtt atccgctcac aattccacac
     5161 aacatacgag ccggaagcat aaagtgtaaa gcctggggtg cctaatgagt gagctaactt
     5221 acattaattg cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc gtgccagctg
     5281 cattaatgaa tcggccaacg cgcggggaga ggcggtttgc gtattgggcg ctcttccgct
     5341 tcctcgctca ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt atcagctcac
     5401 tcaaaggcgg taatacggtt atccacagaa tcaggggata acgcaggaaa gaacatgtga
     5461 gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc gtttttccat
     5521 aggctccgcc cccctgacga gcatcacaaa aatcgacgct caagtcagag gtggcgaaac
     5581 ccgacaggac tataaagata ccaggcgttt ccccctggaa gctccctcgt gcgctctcct
     5641 gttccgaccc tgccgcttac cggatacctg tccgcctttc tcccttcggg aagcgtggcg
     5701 ctttctcata gctcacgctg taggtatctc agttcggtgt aggtcgttcg ctccaagctg
     5761 ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg ccttatccgg taactatcgt
     5821 cttgagtcca acccggtaag acacgactta tcgccactgg cagcagccac tggtaacagg
     5881 attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg gcctaactac
     5941 ggctacacta gaagaacagt atttggtatc tgcgctctgc tgaagccagt taccttcgga
     6001 aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg tggttttttt
     6061 gtttgcaagc agcagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt
     6121 tctacggggt ctgacgctca gtggaacgaa aactcacgtt aagggatttt ggtcatgaga
     6181 ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa aatgaagttt taaatcaatc
     6241 taaagtatat atgagtaaac ttggtctgac agttaccaat gcttaatcag tgaggcacct
     6301 atctcagcga tctgtctatt tcgttcatcc atagttgcct gactccccgt cgtgtagata
     6361 actacgatac gggagggctt accatctggc cccagtgctg caatgatacc gcgagaccca
     6421 cgctcaccgg ctccagattt atcagcaata aaccagccag ccggaagggc cgagcgcaga
     6481 agtggtcctg caactttatc cgcctccatc cagtctatta attgttgccg ggaagctaga
     6541 gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg ccattgctac aggcatcgtg
     6601 gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg gttcccaacg atcaaggcga
     6661 gttacatgat cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt
     6721 gtcagaagta agttggccgc agtgttatca ctcatggtta tggcagcact gcataattct
     6781 cttactgtca tgccatccgt aagatgcttt tctgtgactg gtgagtactc aaccaagtca
     6841 ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat acgggataat
     6901 accgcgccac atagcagaac tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga
     6961 aaactctcaa ggatcttacc gctgttgaga tccagttcga tgtaacccac tcgtgcaccc
     7021 aactgatctt cagcatcttt tactttcacc agcgtttctg ggtgagcaaa aacaggaagg
     7081 caaaatgccg caaaaaaggg aataagggcg acacggaaat gttgaatact catactcttc
     7141 ctttttcaat attattgaag catttatcag ggttattgtc tcatgagcgg atacatattt
     7201 gaatgtattt agaaaaataa acaaataggg gttccgcgca catttccccg aaaagtgcca
     7261 cctgacgt
//
