LOCUS       sequence-198376-        7518 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Suppression of NFS1.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7518)
  AUTHORS   Alvarez SW, Sviderskiy VO, Terzi EM, Papagiannakopoulos T, Moreira 
            AL, Adams S, Sabatini DM, Birsoy K, Possemato R
  TITLE     NFS1 undergoes positive selection in lung tumours and protects cells
            from ferroptosis.
  JOURNAL   Nature. 2017 Nov 22. doi: 10.1038/nature24637.
  PUBMED    29168506
REFERENCE   2  (bases 1 to 7518)
  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: "Nature. 
            2017 Nov 22. doi: 10.1038/nature24637."
COMMENT     Sequence Label: pLKO.1P shNFS1_1
FEATURES             Location/Qualifiers
     source          1..7518
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     28..45
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    162..183
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(198..203,204..221,222..228)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    236..252
                     /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)."
     primer_bind     241..263
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     260..276
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     260..276
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     295..315
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        297..315
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        343..569
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             570..750
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    797..922
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1415..1648
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             1833..1877
                     /codon_start=1
                     /product="antigenic peptide corresponding to amino acids 
                     655 to 669 of the HIV envelope protein gp41 (Lutje Hulsik 
                     et al., 2013)"
                     /label=gp41 peptide
                     /note="recognized by the 2H10 single-chain llama nanobody"
                     /translation="KNEQELLELDKWASL"
     promoter        2055..2303
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     2055..2075
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     2226..2245
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_feature    2400..2517
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        2566..3076
                     /label=hPGK promoter
                     /note="human phosphoglycerate kinase 1 promoter"
     primer_bind     2995..3013
                     /label=hPGK-F
                     /note="Human PGK promoter, forward primer"
     CDS             3098..3697
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(3098..3117)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     3594..3614
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     misc_feature    3713..4301
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3766..3786)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(4184..4195)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             4373..4606
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    4684..4805
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4721..4740)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4775..4794
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      4845..4980
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(5000..5019)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(5001..5019)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(5029..5046)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(5029..5045)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5038..5060)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      5187..5642
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5274..5293)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5484..5505
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        5668..5772
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5773..5841,5842..6633)
                     /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     complement(5991..6010)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6804..7392
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7293..7312
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 gctcgccgca gccgaacgac cgagcgcagc gagtcagtga gcgaggaagc ggaagagcgc
       61 ccaatacgca aaccgcctct ccccgcgcgt tggccgattc attaatgcag ctggcacgac
      121 aggtttcccg actggaaagc gggcagtgag cgcaacgcaa ttaatgtgag ttagctcact
      181 cattaggcac cccaggcttt acactttatg cttccggctc gtatgttgtg tggaattgtg
      241 agcggataac aatttcacac aggaaacagc tatgaccatg attacgccaa gcgcgcaatt
      301 aaccctcact aaagggaaca aaagctggag ctgcaagctt aatgtagtct tatgcaatac
      361 tcttgtagtc ttgcaacatg gtaacgatga gttagcaaca tgccttacaa ggagagaaaa
      421 agcaccgtgc atgccgattg gtggaagtaa ggtggtacga tcgtgcctta ttaggaaggc
      481 aacagacggg tctgacatgg attggacgaa ccactgaatt gccgcattgc agagatattg
      541 tatttaagtg cctagctcga tacataaacg ggtctctctg gttagaccag atctgagcct
      601 gggagctctc tggctaacta gggaacccac tgcttaagcc tcaataaagc ttgccttgag
      661 tgcttcaagt agtgtgtgcc cgtctgttgt gtgactctgg taactagaga tccctcagac
      721 ccttttagtc agtgtggaaa atctctagca gtggcgcccg aacagggact tgaaagcgaa
      781 agggaaacca gaggagctct ctcgacgcag gactcggctt gctgaagcgc gcacggcaag
      841 aggcgagggg cggcgactgg tgagtacgcc aaaaattttg actagcggag gctagaagga
      901 gagagatggg tgcgagagcg tcagtattaa gcgggggaga attagatcgc gatgggaaaa
      961 aattcggtta aggccagggg gaaagaaaaa atataaatta aaacatatag tatgggcaag
     1021 cagggagcta gaacgattcg cagttaatcc tggcctgtta gaaacatcag aaggctgtag
     1081 acaaatactg ggacagctac aaccatccct tcagacagga tcagaagaac ttagatcatt
     1141 atataataca gtagcaaccc tctattgtgt gcatcaaagg atagagataa aagacaccaa
     1201 ggaagcttta gacaagatag aggaagagca aaacaaaagt aagaccaccg cacagcaagc
     1261 ggccgctgat cttcagacct ggaggaggag atatgaggga caattggaga agtgaattat
     1321 ataaatataa agtagtaaaa attgaaccat taggagtagc acccaccaag gcaaagagaa
     1381 gagtggtgca gagagaaaaa agagcagtgg gaataggagc tttgttcctt gggttcttgg
     1441 gagcagcagg aagcactatg ggcgcagcgt caatgacgct gacggtacag gccagacaat
     1501 tattgtctgg tatagtgcag cagcagaaca atttgctgag ggctattgag gcgcaacagc
     1561 atctgttgca actcacagtc tggggcatca agcagctcca ggcaagaatc ctggctgtgg
     1621 aaagatacct aaaggatcaa cagctcctgg ggatttgggg ttgctctgga aaactcattt
     1681 gcaccactgc tgtgccttgg aatgctagtt ggagtaataa atctctggaa cagatttgga
     1741 atcacacgac ctggatggag tgggacagag aaattaacaa ttacacaagc ttaatacact
     1801 ccttaattga agaatcgcaa aaccagcaag aaaagaatga acaagaatta ttggaattag
     1861 ataaatgggc aagtttgtgg aattggttta acataacaaa ttggctgtgg tatataaaat
     1921 tattcataat gatagtagga ggcttggtag gtttaagaat agtttttgct gtactttcta
     1981 tagtgaatag agttaggcag ggatattcac cattatcgtt tcagacccac ctcccaaccc
     2041 cgaggggacc cagagagggc ctatttccca tgattccttc atatttgcat atacgataca
     2101 aggctgttag agagataatt agaattaatt tgactgtaaa cacaaagata ttagtacaaa
     2161 atacgtgacg tagaaagtaa taatttcttg ggtagtttgc agttttaaaa ttatgtttta
     2221 aaatggacta tcatatgctt accgtaactt gaaagtattt cgatttcttg gctttatata
     2281 tcttgtggaa aggacgaggt accggcagtt ccagaaaggt atatttctcg agaaatatac
     2341 ctttctggaa ctgtttttga attctagatc ttgagacaaa tggcagtatt catccacaat
     2401 tttaaaagaa aaggggggat tggggggtac agtgcagggg aaagaatagt agacataata
     2461 gcaacagaca tacaaactaa agaattacaa aaacaaatta caaaaattca aaattttcgg
     2521 gtttattaca gggacagcag agatccactt tggcgccggc tcgagggggc ccgggttgcg
     2581 ccttttccaa ggcagccctg ggtttgcgca gggacgcggc tgctctgggc gtggttccgg
     2641 gaaacgcagc ggcgccgacc ctgggtctcg cacattcttc acgtccgttc gcagcgtcac
     2701 ccggatcttc gccgctaccc ttgtgggccc cccggcgacg cttcctgctc cgcccctaag
     2761 tcgggaaggt tccttgcggt tcgcggcgtg ccggacgtga caaacggaag ccgcacgtct
     2821 cactagtacc ctcgcagacg gacagcgcca gggagcaatg gcagcgcgcc gaccgcgatg
     2881 ggctgtggcc aatagcggct gctcagcagg gcgcgccgag agcagcggcc gggaaggggc
     2941 ggtgcgggag gcggggtgtg gggcggtagt gtgggccctg ttcctgcccg cgcggtgttc
     3001 cgcattctgc aagcctccgg agcgcacgtc ggcagtcggc tccctcgttg accgaatcac
     3061 cgacctctct ccccaggggg atccaccgga gcttaccatg accgagtaca agcccacggt
     3121 gcgcctcgcc acccgcgacg acgtccccag ggccgtacgc accctcgccg ccgcgttcgc
     3181 cgactacccc gccacgcgcc acaccgtcga tccggaccgc cacatcgagc gggtcaccga
     3241 gctgcaagaa ctcttcctca cgcgcgtcgg gctcgacatc ggcaaggtgt gggtcgcgga
     3301 cgacggcgcc gccgtggcgg tctggaccac gccggagagc gtcgaagcgg gggcggtgtt
     3361 cgccgagatc ggcccgcgca tggccgagtt gagcggttcc cggctggccg cgcagcaaca
     3421 gatggaaggc ctcctggcgc cgcaccggcc caaggagccc gcgtggttcc tggccaccgt
     3481 cggcgtctcg cccgaccacc agggcaaggg tctgggcagc gccgtcgtgc tccccggagt
     3541 ggaggcggcc gagcgcgccg gggtgcccgc cttcctggag acctccgcgc cccgcaacct
     3601 ccccttctac gagcggctcg gcttcaccgt caccgccgac gtcgaggtgc ccgaaggacc
     3661 gcgcacctgg tgcatgaccc gcaagcccgg tgcctgaacg cgttaagtcg acaatcaacc
     3721 tctggattac aaaatttgtg aaagattgac tggtattctt aactatgttg ctccttttac
     3781 gctatgtgga tacgctgctt taatgccttt gtatcatgct attgcttccc gtatggcttt
     3841 cattttctcc tccttgtata aatcctggtt gctgtctctt tatgaggagt tgtggcccgt
     3901 tgtcaggcaa cgtggcgtgg tgtgcactgt gtttgctgac gcaaccccca ctggttgggg
     3961 cattgccacc acctgtcagc tcctttccgg gactttcgct ttccccctcc ctattgccac
     4021 ggcggaactc atcgccgcct gccttgcccg ctgctggaca ggggctcggc tgttgggcac
     4081 tgacaattcc gtggtgttgt cggggaaatc atcgtccttt ccttggctgc tcgcctgtgt
     4141 tgccacctgg attctgcgcg ggacgtcctt ctgctacgtc ccttcggccc tcaatccagc
     4201 ggaccttcct tcccgcggcc tgctgccggc tctgcggcct cttccgcgtc ttcgccttcg
     4261 ccctcagacg agtcggatct ccctttgggc cgcctccccg cgtcgacttt aagaccaatg
     4321 acttacaagg cagctgtaga tcttagccac tttttaaaag aaaagggggg actggaaggg
     4381 ctaattcact cccaacgaag acaagatctg ctttttgctt gtactgggtc tctctggtta
     4441 gaccagatct gagcctggga gctctctggc taactaggga acccactgct taagcctcaa
     4501 taaagcttgc cttgagtgct tcaagtagtg tgtgcccgtc tgttgtgtga ctctggtaac
     4561 tagagatccc tcagaccctt ttagtcagtg tggaaaatct ctagcagtac gtatagtagt
     4621 tcatgtcatc ttattattca gtatttataa cttgcaaaga aatgaatatc agagagtgag
     4681 aggaacttgt ttattgcagc ttataatggt tacaaataaa gcaatagcat cacaaatttc
     4741 acaaataaag catttttttc actgcattct agttgtggtt tgtccaaact catcaatgta
     4801 tcttatcatg tctggctcta gctatcccgc ccctaactcc gcccatcccg cccctaactc
     4861 cgcccagttc cgcccattct ccgccccatg gctgactaat tttttttatt tatgcagagg
     4921 ccgaggccgc ctcggcctct gagctattcc agaagtagtg aggaggcttt tttggaggcc
     4981 tagggacgta cccaattcgc cctatagtga gtcgtattac gcgcgctcac tggccgtcgt
     5041 tttacaacgt cgtgactggg aaaaccctgg cgttacccaa cttaatcgcc ttgcagcaca
     5101 tccccctttc gccagctggc gtaatagcga agaggcccgc accgatcgcc cttcccaaca
     5161 gttgcgcagc ctgaatggcg aatgggacgc gccctgtagc ggcgcattaa gcgcggcggg
     5221 tgtggtggtt acgcgcagcg tgaccgctac acttgccagc gccctagcgc ccgctccttt
     5281 cgctttcttc ccttcctttc tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg
     5341 ggggctccct ttagggttcc gatttagtgc tttacggcac ctcgacccca aaaaacttga
     5401 ttagggtgat ggttcacgta gtgggccatc gccctgatag acggtttttc gccctttgac
     5461 gttggagtcc acgttcttta atagtggact cttgttccaa actggaacaa cactcaaccc
     5521 tatctcggtc tattcttttg atttataagg gattttgccg atttcggcct attggttaaa
     5581 aaatgagctg atttaacaaa aatttaacgc gaattttaac aaaatattaa cgcttacaat
     5641 ttaggtggca cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt tttctaaata
     5701 cattcaaata tgtatccgct catgagacaa taaccctgat aaatgcttca ataatattga
     5761 aaaaggaaga gtatgagtat tcaacatttc cgtgtcgccc ttattccctt ttttgcggca
     5821 ttttgccttc ctgtttttgc tcacccagaa acgctggtga aagtaaaaga tgctgaagat
     5881 cagttgggtg cacgagtggg ttacatcgaa ctggatctca acagcggtaa gatccttgag
     5941 agttttcgcc ccgaagaacg ttttccaatg atgagcactt ttaaagttct gctatgtggc
     6001 gcggtattat cccgtattga cgccgggcaa gagcaactcg gtcgccgcat acactattct
     6061 cagaatgact tggttgagta ctcaccagtc acagaaaagc atcttacgga tggcatgaca
     6121 gtaagagaat tatgcagtgc tgccataacc atgagtgata acactgcggc caacttactt
     6181 ctgacaacga tcggaggacc gaaggagcta accgcttttt tgcacaacat gggggatcat
     6241 gtaactcgcc ttgatcgttg ggaaccggag ctgaatgaag ccataccaaa cgacgagcgt
     6301 gacaccacga tgcctgtagc aatggcaaca acgttgcgca aactattaac tggcgaacta
     6361 cttactctag cttcccggca acaattaata gactggatgg aggcggataa agttgcagga
     6421 ccacttctgc gctcggccct tccggctggc tggtttattg ctgataaatc tggagccggt
     6481 gagcgtgggt ctcgcggtat cattgcagca ctggggccag atggtaagcc ctcccgtatc
     6541 gtagttatct acacgacggg gagtcaggca actatggatg aacgaaatag acagatcgct
     6601 gagataggtg cctcactgat taagcattgg taactgtcag accaagttta ctcatatata
     6661 ctttagattg atttaaaact tcatttttaa tttaaaagga tctaggtgaa gatccttttt
     6721 gataatctca tgaccaaaat cccttaacgt gagttttcgt tccactgagc gtcagacccc
     6781 gtagaaaaga tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg
     6841 caaacaaaaa aaccaccgct accagcggtg gtttgtttgc cggatcaaga gctaccaact
     6901 ctttttccga aggtaactgg cttcagcaga gcgcagatac caaatactgt tcttctagtg
     6961 tagccgtagt taggccacca cttcaagaac tctgtagcac cgcctacata cctcgctctg
     7021 ctaatcctgt taccagtggc tgctgccagt ggcgataagt cgtgtcttac cgggttggac
     7081 tcaagacgat agttaccgga taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca
     7141 cagcccagct tggagcgaac gacctacacc gaactgagat acctacagcg tgagctatga
     7201 gaaagcgcca cgcttcccga agggagaaag gcggacaggt atccggtaag cggcagggtc
     7261 ggaacaggag agcgcacgag ggagcttcca gggggaaacg cctggtatct ttatagtcct
     7321 gtcgggtttc gccacctctg acttgagcgt cgatttttgt gatgctcgtc aggggggcgg
     7381 agcctatgga aaaacgccag caacgcggcc tttttacggt tcctggcctt ttgctggcct
     7441 tttgctcaca tgttctttcc tgcgttatcc cctgattctg tggataaccg tattaccgcc
     7501 tttgagtgag ctgatacc
//
