LOCUS       sequence-458887-        6117 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Transient overexpression of mouse FEM1A in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6117)
  AUTHORS   Manford AG, Mena EL, Shih KY, Gee CL, McMinimy R, Martinez-Gonzalez 
            B, Sherriff R, Lew B, Zoltek M, Rodriguez-Perez F, Woldesenbet M, 
            Kuriyan J, Rape M
  TITLE     Structural basis and regulation of the reductive stress response.
  JOURNAL   Cell. 2021 Oct 14;184(21):5375-5390.e16. doi: 
            10.1016/j.cell.2021.09.002. Epub 2021 Sep 24.
  PUBMED    34562363
REFERENCE   2  (bases 1 to 6117)
  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: 
            "10.1016/j.cell.2021.09.002"; journalName: "Cell"; date: 
            "2021-10-14- 14"; volume: "184"; issue: "21"; pages: "5375-5390.e16"
COMMENT     Sequence Label: pCS2 3xFLAG FEM1A
FEATURES             Location/Qualifiers
     source          1..6117
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        35..53
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     35..52
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     regulatory      85..94
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             94..159
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     polyA_signal    2169..2303
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2193..2212)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     2247..2266
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(2416..2436)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(2416..2434)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(2455..2471)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2455..2471)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2468..2490)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2479..2495
                     /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(2503..2509,2510..2527,2528..2533))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2548..2569
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2686..2703)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2857..3445)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2937..2956)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3616..4407,4408..4476))
                     /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     4239..4258
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4477..4581)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(4607..5062)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4744..4765)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4956..4975
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        5133..6117
                     /label=CMV IE94 promoter
                     /note="enhancer/promoter region of simian cytomegalovirus 
                     major immediate early transcription unit IE94"
ORIGIN
        1 cgccattctg cctggggacg tcggagcaag cttgatttag gtgacactat agaatacaag
       61 ctacttgttc tttttgcagg atccgccacc atggactaca aagaccatga cggtgattat
      121 aaagatcatg acatcgatta caaggatgac gatgacaagg gcggaggagt cgacatggat
      181 ctgcacacag cggtgtacaa tgcggcgcac gacggcaagc tgccgctgct gcagaagctg
      241 ctggccggcc gtgggcgtga ggagctggag gagctgctgg gtgaggtggc cggcggtggc
      301 acgccactgc taatcgcagc gcgccgtgga cacctagacg tggtggagta tctggtggac
      361 cactgcggcg ccagcgtgga ggcgagcggc tccgtgcact tcgatggcga gaccatcgag
      421 ggcgcgccgc cgctctgggc agcgtcggcc gcgggtcacc tggccgtggt gcgtagcctt
      481 ctgcgccgag gcgcctcggt caaccgcacc acgcgcacca actccacgcc gctgcgcgcc
      541 gcctgcttcg acggccacct ggacgtggta cgctacctgg tgggcgagca caaggcggac
      601 ctggaggtgg ccaatcgcca cggccacacg tgccttatga tctcctgcta caagggccac
      661 cgcgagatcg cgcgctacct gctggagcgc ggcgcacagg tgaaccggcg cagcgccaag
      721 ggcaacaccg ccctgcacga ctgtgcagaa tctggcagcc tggagatcct acagctgctg
      781 ctgggctgcc acgcgcgcat ggagcgcgac ggctatggca tgaccccgct gctggccgcc
      841 agcgtcaccg ggcacaccaa catcgtggag tacctcatcc aggaacagcc gggccacgag
      901 cagctctcag ggacagagct tcccggggaa ggctcctccc aggtggcagg aaaccactgc
      961 tccaccccag aagaagcaga accgtatgag agctgctgcc ctaccagccg ggaagcagct
     1021 gtggaagctt tggagctgct gggtgccacc tacgtggata agaaaaggga tctgcttgga
     1081 gccctgaagc actggagaag ggcgatggaa ctccgtcacc agggtggagg ctacctccct
     1141 aagccggagc cccagcaact ggttctggcc tatgactatt ccagggaggt gactacaccc
     1201 caagagttgg aagccctcat tacagatcct gatgaaatgc ggatgcaggc gctgctgata
     1261 cgggagagaa tcttgggccc ctcacatcct gacacttcat actacatacg gtaccggggt
     1321 gctgtctatg cggactccgg gaatttcgag cgctgtatcc gtctgtggaa gtatgccttg
     1381 gacatgcaac aaaacaactt ggagcccctg agccccatga ccgccagcag cttcctgtcg
     1441 tttgcagagc tcttctccta cgtgctgcag gaccgctcgg ccaagggcaa cctgggcatg
     1501 cagcttggct ttgccgacct catgggtgtg ctcagcaaag gggttcggga agtggagcgg
     1561 gctctgcagt tgcccaagga accgggtgac tccgcacagt tcaccaaagc cattgccatc
     1621 atcctccacc ttctgtacct attggagaag gtggagtgca cccccagcca ggaacacctc
     1681 aagcaccaga cagtctaccg cctgctcaag tgtgcccctc gcggtaaaaa cggcttcacc
     1741 ccactacaca tggcggtgga caaggagacc accaacgtgg gccggtatcg tgtgggcgtc
     1801 ttcccatcgc tgcaggtggt caaggtgctg ttggactgcg gagcagaccc tgacagcagg
     1861 gatttcgaca acaacacccc actgcacatc gctgcccaga ataattgccc ggccatcatg
     1921 gatgctctca tcgaagctgg ggcccacatg gacgccacca atgccttcaa aaagactgcc
     1981 tacgagctgc tggactcgaa gctgttggct aagagcaccg tgcagccttt caattatgtg
     2041 acactgcagt gcctagctgc ccgtgccctg gacaggaaca aggtccctta taagggcttc
     2101 atcccagagg agctggaggc cttcatccag ttgcattgat ctagaactat agtgagtcgt
     2161 attacgtaga tccagacatg ataagataca ttgatgagtt tggacaaacc acaactagaa
     2221 tgcagtgaaa aaaatgcttt atttgtgaaa tttgtgatgc tattgcttta tttgtaacca
     2281 ttataagctg caataaacaa gttaacaaca acaattgcat tcattttatg tttcaggttc
     2341 agggggaggt gtgggaggtt ttttaattcg cggccgcggc gccaatgcat tgggcccggt
     2401 acccagcttt tgttcccttt agtgagggtt aattgcgcgc ttggcgtaat catggtcata
     2461 gctgtttcct gtgtgaaatt gttatccgct cacaattcca cacaacatac gagccgggag
     2521 cataaagtgt aaagcctggg gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg
     2581 ctcactgccc gctttccagt cgggaaacct gtcgtgccag ctgcattaat gaatcggcca
     2641 acgcgcgggg agaggcggtt tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc
     2701 gctgcgctcg gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg cggtaatacg
     2761 gttatccaca gaatcagggg ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa
     2821 ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga
     2881 cgagcatcac aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag
     2941 ataccaggcg tttccccctg gaagctccct cgtgcgctct cctgttccga ccctgccgct
     3001 taccggatac ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc atagctcacg
     3061 ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc
     3121 ccccgttcag cccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccggt
     3181 aagacacgac ttatcgccac tggcagcagc cactggtaac aggattagca gagcgaggta
     3241 tgtaggcggt gctacagagt tcttgaagtg gtggcctaac tacggctaca ctagaagaac
     3301 agtatttggt atctgcgctc tgctgaagcc agttaccttc ggaaaaagag ttggtagctc
     3361 ttgatccggc aaacaaacca ccgctggtag cggtggtttt tttgtttgca agcagcagat
     3421 tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc
     3481 tcagtggaac gaaaactcac gttaagggat tttggtcatg agattatcaa aaaggatctt
     3541 cacctagatc cttttaaatt aaaaatgaag ttttaaatca atctaaagta tatatgagta
     3601 aacttggtct gacagttacc aatgcttaat cagtgaggca cctatctcag cgatctgtct
     3661 atttcgttca tccatagttg cctgactccc cgtcgtgtag ataactacga tacgggaggg
     3721 cttaccatct ggccccagtg ctgcaatgat accgcgagac ccacgctcac cggctccaga
     3781 tttatcagca ataaaccagc cagccggaag ggccgagcgc agaagtggtc ctgcaacttt
     3841 atccgcctcc atccagtcta ttaattgttg ccgggaagct agagtaagta gttcgccagt
     3901 taatagtttg cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt
     3961 tggtatggct tcattcagct ccggttccca acgatcaagg cgagttacat gatcccccat
     4021 gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc gttgtcagaa gtaagttggc
     4081 cgcagtgtta tcactcatgg ttatggcagc actgcataat tctcttactg tcatgccatc
     4141 cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag tcattctgag aatagtgtat
     4201 gcggcgaccg agttgctctt gcccggcgtc aatacgggat aataccgcgc cacatagcag
     4261 aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct caaggatctt
     4321 accgctgttg agatccagtt cgatgtaacc cactcgtgca cccaactgat cttcagcatc
     4381 ttttactttc accagcgttt ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa
     4441 gggaataagg gcgacacgga aatgttgaat actcatactc ttcctttttc aatattattg
     4501 aagcatttat cagggttatt gtctcatgag cggatacata tttgaatgta tttagaaaaa
     4561 taaacaaata ggggttccgc gcacatttcc ccgaaaagtg ccacctaaat tgtaagcgtt
     4621 aatattttgt taaaattcgc gttaaatttt tgttaaatca gctcattttt taaccaatag
     4681 gccgaaatcg gcaaaatccc ttataaatca aaagaataga ccgagatagg gttgagtgtt
     4741 gttccagttt ggaacaagag tccactatta aagaacgtgg actccaacgt caaagggcga
     4801 aaaaccgtct atcagggcga tggcccacta cgtgaaccat caccctaatc aagttttttg
     4861 gggtcgaggt gccgtaaagc actaaatcgg aaccctaaag ggagcccccg atttagagct
     4921 tgacggggaa agccggcgaa cgtggcgaga aaggaaggga agaaagcgaa aggagcgggc
     4981 gctagggcgc tggcaagtgt agcggtcacg ctgcgcgtaa ccaccacacc cgccgcgctt
     5041 aatgcgccgc tacagggcgc gtcccattcg ccattcaggc tgcgcaactg ttgggaaggg
     5101 cgatcggtgc gggcctcttc gctattacgc cagtcgaccg ccaattcaat atggcgtata
     5161 tggactcatg ccaattcaat atggtggatc tggacctgtg ccaattcaat atggcgtata
     5221 tggactcgtg ccaattcaat atggtggatc tggaccccag ccaattcaat atggcggact
     5281 tggcaccatg ccaattcaat atggcggacc tggcactgtg ccaactgggg aggggtctac
     5341 ttggcacggt gccaagtttg aggaggggtc ttggccctgt gccaagtccg ccatattgaa
     5401 ttggcatggt gccaataatg gcggccatat tggctatatg ccaggatcaa tatataggca
     5461 atatccaata tggccctatg ccaatatggc tattggccag gttcaatact atgtattggc
     5521 cctatgccat atagtattcc atatatgggt tttcctattg acgtagatag cccctcccaa
     5581 tgggcggtcc catataccat atatggggct tcctaatacc gcccatagcc actcccccat
     5641 tgacgtcaat ggtctctata tatggtcttt cctattgacg tcatatgggc ggtcctattg
     5701 acgtatatgg cgcctccccc attgacgtca attacggtaa atggcccgcc tggctcaatg
     5761 cccattgacg tcaataggac cacccaccat tgacgtcaat gggatggctc attgcccatt
     5821 catatccgtt ctcacgcccc ctattgacgt caatgacggt aaatggccca cttggcagta
     5881 catcaatatc tattaatagt aacttggcaa gtacattact attggaagta cgccagggta
     5941 cattggcagt actcccattg acgtcaatgg cggtaaatgg cccgcgatgg ctgccaagta
     6001 catccccatt gacgtcaatg gggaggggca atgacgcaaa tgggcgttcc attgacgtaa
     6061 atgggcggta ggcgtgccta atgggaggtc tatataagca atgctcgttt agggaac
//
