LOCUS       sequence-198943-        7083 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Suppression of FXN.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7083)
  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 7083)
  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 shFXN_2
FEATURES             Location/Qualifiers
     source          1..7083
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        16..526
                     /label=hPGK promoter
                     /note="human phosphoglycerate kinase 1 promoter"
     primer_bind     445..463
                     /label=hPGK-F
                     /note="Human PGK promoter, forward primer"
     CDS             548..1147
                     /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(548..567)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     1044..1064
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     LTR             1275..1508
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    1580..1701
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1617..1636)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1671..1690
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      1741..1876
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(1896..1915)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1897..1915)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1925..1942)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(1925..1941)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1934..1956)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      2083..2538
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2170..2189)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2380..2401
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        2564..2668
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2669..2737,2738..3529)
                     /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(2887..2906)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3700..4288
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4189..4208
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4442..4459
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    4576..4597
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(4612..4617,4618..4635,4636..4642)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4650..4666
                     /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     4655..4677
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     4674..4690
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     4674..4690
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     4709..4729
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        4711..4729
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        4757..4983
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             4984..5164
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    5211..5336
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    5829..6062
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             6247..6291
                     /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        6589..6837
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     6589..6609
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     6760..6779
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_feature    6933..7050
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
ORIGIN
        1 tggccgcggc tcgagggggt tggggttgcg ccttttccaa ggcagccctg ggtttgcgca
       61 gggacgcggc tgctctgggc gtggttccgg gaaacgcagc ggcgccgacc ctgggtctcg
      121 cacattcttc acgtccgttc gcagcgtcac ccggatcttc gccgctaccc ttgtgggccc
      181 cccggcgacg cttcctgctc cgcccctaag tcgggaaggt tccttgcggt tcgcggcgtg
      241 ccggacgtga caaacggaag ccgcacgtct cactagtacc ctcgcagacg gacagcgcca
      301 gggagcaatg gcagcgcgcc gaccgcgatg ggctgtggcc aatagcggct gctcagcagg
      361 gcgcgccgag agcagcggcc gggaaggggc ggtgcgggag gcggggtgtg gggcggtagt
      421 gtgggccctg ttcctgcccg cgcggtgttc cgcattctgc aagcctccgg agcgcacgtc
      481 ggcagtcggc tccctcgttg accgaatcac cgacctctct ccccaggggg atccaccgga
      541 gcttaccatg accgagtaca agcccacggt gcgcctcgcc acccgcgacg acgtccccag
      601 ggccgtacgc accctcgccg ccgcgttcgc cgactacccc gccacgcgcc acaccgtcga
      661 tccggaccgc cacatcgagc gggtcaccga gctgcaagaa ctcttcctca cgcgcgtcgg
      721 gctcgacatc ggcaaggtgt gggtcgcgga cgacggcgcc gcggtggcgg tctggaccac
      781 gccggagagc gtcgaagcgg gggcggtgtt cgccgagatc ggcccgcgca tggccgagtt
      841 gagcggttcc cggctggccg cgcagcaaca gatggaaggc ctcctggcgc cgcaccggcc
      901 caaggagccc gcgtggttcc tggccaccgt cggcgtctcg cccgaccacc agggcaaggg
      961 tctgggcagc gccgtcgtgc tccccggagt ggaggcggcc gagcgcgccg gggtgcccgc
     1021 cttcctggag acctccgcgc cccgcaacct ccccttctac gagcggctcg gcttcaccgt
     1081 caccgccgac gtcgaggtgc ccgaaggacc gcgcacctgg tgcatgaccc gcaagcccgg
     1141 tgcctgacgc ccgccccacg acccgcagcg cccgaccgaa aggagcgcac gaccccatgc
     1201 atcggtacct ttaagaccaa tgacttacaa ggcagctgta gatcttagcc actttttaaa
     1261 agaaaagggg ggactggaag ggctaattca ctcccaacga agacaagatc tgctttttgc
     1321 ttgtactggg tctctctggt tagaccagat ctgagcctgg gagctctctg gctaactagg
     1381 gaacccactg cttaagcctc aataaagctt gccttgagtg cttcaagtag tgtgtgcccg
     1441 tctgttgtgt gactctggta actagagatc cctcagaccc ttttagtcag tgtggaaaat
     1501 ctctagcagt agtagttcat gtcatcttat tattcagtat ttataacttg caaagaaatg
     1561 aatatcagag agtgagagga acttgtttat tgcagcttat aatggttaca aataaagcaa
     1621 tagcatcaca aatttcacaa ataaagcatt tttttcactg cattctagtt gtggtttgtc
     1681 caaactcatc aatgtatctt atcatgtctg gctctagcta tcccgcccct aactccgccc
     1741 atcccgcccc taactccgcc cagttccgcc cattctccgc cccatggctg actaattttt
     1801 tttatttatg cagaggccga ggccgcctcg gcctctgagc tattccagaa gtagtgagga
     1861 ggcttttttg gaggcctagg gacgtaccca attcgcccta tagtgagtcg tattacgcgc
     1921 gctcactggc cgtcgtttta caacgtcgtg actgggaaaa ccctggcgtt acccaactta
     1981 atcgccttgc agcacatccc cctttcgcca gctggcgtaa tagcgaagag gcccgcaccg
     2041 atcgcccttc ccaacagttg cgcagcctga atggcgaatg ggacgcgccc tgtagcggcg
     2101 cattaagcgc ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt gccagcgccc
     2161 tagcgcccgc tcctttcgct ttcttccctt cctttctcgc cacgttcgcc ggctttcccc
     2221 gtcaagctct aaatcggggg ctccctttag ggttccgatt tagtgcttta cggcacctcg
     2281 accccaaaaa acttgattag ggtgatggtt cacgtagtgg gccatcgccc tgatagacgg
     2341 tttttcgccc tttgacgttg gagtccacgt tctttaatag tggactcttg ttccaaactg
     2401 gaacaacact caaccctatc tcggtctatt cttttgattt ataagggatt ttgccgattt
     2461 cggcctattg gttaaaaaat gagctgattt aacaaaaatt taacgcgaat tttaacaaaa
     2521 tattaacgct tacaatttag gtggcacttt tcggggaaat gtgcgcggaa cccctatttg
     2581 tttatttttc taaatacatt caaatatgta tccgctcatg agacaataac cctgataaat
     2641 gcttcaataa tattgaaaaa ggaagagtat gagtattcaa catttccgtg tcgcccttat
     2701 tccctttttt gcggcatttt gccttcctgt ttttgctcac ccagaaacgc tggtgaaagt
     2761 aaaagatgct gaagatcagt tgggtgcacg agtgggttac atcgaactgg atctcaacag
     2821 cggtaagatc cttgagagtt ttcgccccga agaacgtttt ccaatgatga gcacttttaa
     2881 agttctgcta tgtggcgcgg tattatcccg tattgacgcc gggcaagagc aactcggtcg
     2941 ccgcatacac tattctcaga atgacttggt tgagtactca ccagtcacag aaaagcatct
     3001 tacggatggc atgacagtaa gagaattatg cagtgctgcc ataaccatga gtgataacac
     3061 tgcggccaac ttacttctga caacgatcgg aggaccgaag gagctaaccg cttttttgca
     3121 caacatgggg gatcatgtaa ctcgccttga tcgttgggaa ccggagctga atgaagccat
     3181 accaaacgac gagcgtgaca ccacgatgcc tgtagcaatg gcaacaacgt tgcgcaaact
     3241 attaactggc gaactactta ctctagcttc ccggcaacaa ttaatagact ggatggaggc
     3301 ggataaagtt gcaggaccac ttctgcgctc ggcccttccg gctggctggt ttattgctga
     3361 taaatctgga gccggtgagc gtgggtctcg cggtatcatt gcagcactgg ggccagatgg
     3421 taagccctcc cgtatcgtag ttatctacac gacggggagt caggcaacta tggatgaacg
     3481 aaatagacag atcgctgaga taggtgcctc actgattaag cattggtaac tgtcagacca
     3541 agtttactca tatatacttt agattgattt aaaacttcat ttttaattta aaaggatcta
     3601 ggtgaagatc ctttttgata atctcatgac caaaatccct taacgtgagt tttcgttcca
     3661 ctgagcgtca gaccccgtag aaaagatcaa aggatcttct tgagatcctt tttttctgcg
     3721 cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca gcggtggttt gtttgccgga
     3781 tcaagagcta ccaactcttt ttccgaaggt aactggcttc agcagagcgc agataccaaa
     3841 tactgttctt ctagtgtagc cgtagttagg ccaccacttc aagaactctg tagcaccgcc
     3901 tacatacctc gctctgctaa tcctgttacc agtggctgct gccagtggcg ataagtcgtg
     3961 tcttaccggg ttggactcaa gacgatagtt accggataag gcgcagcggt cgggctgaac
     4021 ggggggttcg tgcacacagc ccagcttgga gcgaacgacc tacaccgaac tgagatacct
     4081 acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc
     4141 ggtaagcggc agggtcggaa caggagagcg cacgagggag cttccagggg gaaacgcctg
     4201 gtatctttat agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg
     4261 ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac gcggcctttt tacggttcct
     4321 ggccttttgc tggccttttg ctcacatgtt ctttcctgcg ttatcccctg attctgtgga
     4381 taaccgtatt accgcctttg agtgagctga taccgctcgc cgcagccgaa cgaccgagcg
     4441 cagcgagtca gtgagcgagg aagcggaaga gcgcccaata cgcaaaccgc ctctccccgc
     4501 gcgttggccg attcattaat gcagctggca cgacaggttt cccgactgga aagcgggcag
     4561 tgagcgcaac gcaattaatg tgagttagct cactcattag gcaccccagg ctttacactt
     4621 tatgcttccg gctcgtatgt tgtgtggaat tgtgagcgga taacaatttc acacaggaaa
     4681 cagctatgac catgattacg ccaagcgcgc aattaaccct cactaaaggg aacaaaagct
     4741 ggagctgcaa gcttaatgta gtcttatgca atactcttgt agtcttgcaa catggtaacg
     4801 atgagttagc aacatgcctt acaaggagag aaaaagcacc gtgcatgccg attggtggaa
     4861 gtaaggtggt acgatcgtgc cttattagga aggcaacaga cgggtctgac atggattgga
     4921 cgaaccactg aattgccgca ttgcagagat attgtattta agtgcctagc tcgatacata
     4981 aacgggtctc tctggttaga ccagatctga gcctgggagc tctctggcta actagggaac
     5041 ccactgctta agcctcaata aagcttgcct tgagtgcttc aagtagtgtg tgcccgtctg
     5101 ttgtgtgact ctggtaacta gagatccctc agaccctttt agtcagtgtg gaaaatctct
     5161 agcagtggcg cccgaacagg gacttgaaag cgaaagggaa accagaggag ctctctcgac
     5221 gcaggactcg gcttgctgaa gcgcgcacgg caagaggcga ggggcggcga ctggtgagta
     5281 cgccaaaaat tttgactagc ggaggctaga aggagagaga tgggtgcgag agcgtcagta
     5341 ttaagcgggg gagaattaga tcgcgatggg aaaaaattcg gttaaggcca gggggaaaga
     5401 aaaaatataa attaaaacat atagtatggg caagcaggga gctagaacga ttcgcagtta
     5461 atcctggcct gttagaaaca tcagaaggct gtagacaaat actgggacag ctacaaccat
     5521 cccttcagac aggatcagaa gaacttagat cattatataa tacagtagca accctctatt
     5581 gtgtgcatca aaggatagag ataaaagaca ccaaggaagc tttagacaag atagaggaag
     5641 agcaaaacaa aagtaagacc accgcacagc aagcggccgc tgatcttcag acctggagga
     5701 ggagatatga gggacaattg gagaagtgaa ttatataaat ataaagtagt aaaaattgaa
     5761 ccattaggag tagcacccac caaggcaaag agaagagtgg tgcagagaga aaaaagagca
     5821 gtgggaatag gagctttgtt ccttgggttc ttgggagcag caggaagcac tatgggcgca
     5881 gcgtcaatga cgctgacggt acaggccaga caattattgt ctggtatagt gcagcagcag
     5941 aacaatttgc tgagggctat tgaggcgcaa cagcatctgt tgcaactcac agtctggggc
     6001 atcaagcagc tccaggcaag aatcctggct gtggaaagat acctaaagga tcaacagctc
     6061 ctggggattt ggggttgctc tggaaaactc atttgcacca ctgctgtgcc ttggaatgct
     6121 agttggagta ataaatctct ggaacagatt tggaatcaca cgacctggat ggagtgggac
     6181 agagaaatta acaattacac aagcttaata cactccttaa ttgaagaatc gcaaaaccag
     6241 caagaaaaga atgaacaaga attattggaa ttagataaat gggcaagttt gtggaattgg
     6301 tttaacataa caaattggct gtggtatata aaattattca taatgatagt aggaggcttg
     6361 gtaggtttaa gaatagtttt tgctgtactt tctatagtga atagagttag gcagggatat
     6421 tcaccattat cgtttcagac ccacctccca accccgaggg gacccgacag gcccgaagga
     6481 atagaagaag aaggtggaga gagagacaga gacagatcca ttcgattagt gaacggatct
     6541 cgacggtatc gatcacgaga ctagcctcga gcggccgccc ccttcaccga gggcctattt
     6601 cccatgattc cttcatattt gcatatacga tacaaggctg ttagagagat aattggaatt
     6661 aatttgactg taaacacaaa gatattagta caaaatacgt gacgtagaaa gtaataattt
     6721 cttgggtagt ttgcagtttt aaaattatgt tttaaaatgg actatcatat gcttaccgta
     6781 acttgaaagt atttcgattt cttggcttta tatatcttgt ggaaaggacg aaacaccggg
     6841 cagacgccaa acaagcaaat ctcgagattt gcttgtttgg cgtctgcttt ttaattctcg
     6901 acctcgagac aaatggcagt attcatccac aattttaaaa gaaaaggggg gattgggggg
     6961 tacagtgcag gggaaagaat agtagacata atagcaacag acatacaaac taaagaatta
     7021 caaaaacaaa ttacaaaaat tcaaaatttt cgggtttatt acagggacag cagagatcca
     7081 ctt
//
