LOCUS       sequence-333096-        7084 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Suppress TRFC.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7084)
  AUTHORS   Terzi EM, Sviderskiy VO, Alvarez SW, Whiten GC, Possemato R
  TITLE     Iron-sulfur cluster deficiency can be sensed by IRP2 and regulates 
            iron homeostasis and sensitivity to ferroptosis independent of IRP1 
            and FBXL5.
  JOURNAL   Sci Adv. 2021 May 26;7(22). pii: 7/22/eabg4302. doi: 
            10.1126/sciadv.abg4302. Print 2021 May.
  PUBMED    34039609
REFERENCE   2  (bases 1 to 7084)
  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: "Sci Adv."; 
            date: "2021-05-26"; volume: "7(22). pii: 7/22/eabg4302. doi"; pages:
            " 10.1126/sciadv.abg4302. Print 2021 May"
COMMENT     Sequence Label: pLKO.1P shTFRC
FEATURES             Location/Qualifiers
     source          1..7084
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        4..514
                     /label=hPGK promoter
                     /note="human phosphoglycerate kinase 1 promoter"
     primer_bind     433..451
                     /label=hPGK-F
                     /note="Human PGK promoter, forward primer"
     CDS             536..1135
                     /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(536..555)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     1032..1052
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     LTR             1263..1496
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    1568..1689
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1605..1624)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1659..1678
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      1729..1864
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(1884..1903)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1885..1903)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1913..1930)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(1913..1929)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1922..1944)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      2071..2526
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2158..2177)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2368..2389
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        2552..2656
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2657..2725,2726..3517)
                     /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(2875..2894)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3688..4276
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4177..4196
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4430..4447
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    4564..4585
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(4600..4605,4606..4623,4624..4630)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4638..4654
                     /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     4643..4665
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     4662..4678
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     4662..4678
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     4697..4717
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        4699..4717
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        4745..4971
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             4972..5152
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    5199..5324
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    5817..6050
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             6235..6279
                     /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        6577..6825
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     6577..6597
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     6748..6767
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_feature    6922..7039
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
ORIGIN
        1 gagggggttg gggttgcgcc ttttccaagg cagccctggg tttgcgcagg gacgcggctg
       61 ctctgggcgt ggttccggga aacgcagcgg cgccgaccct gggtctcgca cattcttcac
      121 gtccgttcgc agcgtcaccc ggatcttcgc cgctaccctt gtgggccccc cggcgacgct
      181 tcctgctccg cccctaagtc gggaaggttc cttgcggttc gcggcgtgcc ggacgtgaca
      241 aacggaagcc gcacgtctca ctagtaccct cgcagacgga cagcgccagg gagcaatggc
      301 agcgcgccga ccgcgatggg ctgtggccaa tagcggctgc tcagcagggc gcgccgagag
      361 cagcggccgg gaaggggcgg tgcgggaggc ggggtgtggg gcggtagtgt gggccctgtt
      421 cctgcccgcg cggtgttccg cattctgcaa gcctccggag cgcacgtcgg cagtcggctc
      481 cctcgttgac cgaatcaccg acctctctcc ccagggggat ccaccggagc ttaccatgac
      541 cgagtacaag cccacggtgc gcctcgccac ccgcgacgac gtccccaggg ccgtacgcac
      601 cctcgccgcc gcgttcgccg actaccccgc cacgcgccac accgtcgatc cggaccgcca
      661 catcgagcgg gtcaccgagc tgcaagaact cttcctcacg cgcgtcgggc tcgacatcgg
      721 caaggtgtgg gtcgcggacg acggcgccgc ggtggcggtc tggaccacgc cggagagcgt
      781 cgaagcgggg gcggtgttcg ccgagatcgg cccgcgcatg gccgagttga gcggttcccg
      841 gctggccgcg cagcaacaga tggaaggcct cctggcgccg caccggccca aggagcccgc
      901 gtggttcctg gccaccgtcg gcgtctcgcc cgaccaccag ggcaagggtc tgggcagcgc
      961 cgtcgtgctc cccggagtgg aggcggccga gcgcgccggg gtgcccgcct tcctggagac
     1021 ctccgcgccc cgcaacctcc ccttctacga gcggctcggc ttcaccgtca ccgccgacgt
     1081 cgaggtgccc gaaggaccgc gcacctggtg catgacccgc aagcccggtg cctgacgccc
     1141 gccccacgac ccgcagcgcc cgaccgaaag gagcgcacga ccccatgcat cggtaccttt
     1201 aagaccaatg acttacaagg cagctgtaga tcttagccac tttttaaaag aaaagggggg
     1261 actggaaggg ctaattcact cccaacgaag acaagatctg ctttttgctt gtactgggtc
     1321 tctctggtta gaccagatct gagcctggga gctctctggc taactaggga acccactgct
     1381 taagcctcaa taaagcttgc cttgagtgct tcaagtagtg tgtgcccgtc tgttgtgtga
     1441 ctctggtaac tagagatccc tcagaccctt ttagtcagtg tggaaaatct ctagcagtag
     1501 tagttcatgt catcttatta ttcagtattt ataacttgca aagaaatgaa tatcagagag
     1561 tgagaggaac ttgtttattg cagcttataa tggttacaaa taaagcaata gcatcacaaa
     1621 tttcacaaat aaagcatttt tttcactgca ttctagttgt ggtttgtcca aactcatcaa
     1681 tgtatcttat catgtctggc tctagctatc ccgcccctaa ctccgcccat cccgccccta
     1741 actccgccca gttccgccca ttctccgccc catggctgac taattttttt tatttatgca
     1801 gaggccgagg ccgcctcggc ctctgagcta ttccagaagt agtgaggagg cttttttgga
     1861 ggcctaggga cgtacccaat tcgccctata gtgagtcgta ttacgcgcgc tcactggccg
     1921 tcgttttaca acgtcgtgac tgggaaaacc ctggcgttac ccaacttaat cgccttgcag
     1981 cacatccccc tttcgccagc tggcgtaata gcgaagaggc ccgcaccgat cgcccttccc
     2041 aacagttgcg cagcctgaat ggcgaatggg acgcgccctg tagcggcgca ttaagcgcgg
     2101 cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc
     2161 ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg ctttccccgt caagctctaa
     2221 atcgggggct ccctttaggg ttccgattta gtgctttacg gcacctcgac cccaaaaaac
     2281 ttgattaggg tgatggttca cgtagtgggc catcgccctg atagacggtt tttcgccctt
     2341 tgacgttgga gtccacgttc tttaatagtg gactcttgtt ccaaactgga acaacactca
     2401 accctatctc ggtctattct tttgatttat aagggatttt gccgatttcg gcctattggt
     2461 taaaaaatga gctgatttaa caaaaattta acgcgaattt taacaaaata ttaacgctta
     2521 caatttaggt ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt tatttttcta
     2581 aatacattca aatatgtatc cgctcatgag acaataaccc tgataaatgc ttcaataata
     2641 ttgaaaaagg aagagtatga gtattcaaca tttccgtgtc gcccttattc ccttttttgc
     2701 ggcattttgc cttcctgttt ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga
     2761 agatcagttg ggtgcacgag tgggttacat cgaactggat ctcaacagcg gtaagatcct
     2821 tgagagtttt cgccccgaag aacgttttcc aatgatgagc acttttaaag ttctgctatg
     2881 tggcgcggta ttatcccgta ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta
     2941 ttctcagaat gacttggttg agtactcacc agtcacagaa aagcatctta cggatggcat
     3001 gacagtaaga gaattatgca gtgctgccat aaccatgagt gataacactg cggccaactt
     3061 acttctgaca acgatcggag gaccgaagga gctaaccgct tttttgcaca acatggggga
     3121 tcatgtaact cgccttgatc gttgggaacc ggagctgaat gaagccatac caaacgacga
     3181 gcgtgacacc acgatgcctg tagcaatggc aacaacgttg cgcaaactat taactggcga
     3241 actacttact ctagcttccc ggcaacaatt aatagactgg atggaggcgg ataaagttgc
     3301 aggaccactt ctgcgctcgg cccttccggc tggctggttt attgctgata aatctggagc
     3361 cggtgagcgt gggtctcgcg gtatcattgc agcactgggg ccagatggta agccctcccg
     3421 tatcgtagtt atctacacga cggggagtca ggcaactatg gatgaacgaa atagacagat
     3481 cgctgagata ggtgcctcac tgattaagca ttggtaactg tcagaccaag tttactcata
     3541 tatactttag attgatttaa aacttcattt ttaatttaaa aggatctagg tgaagatcct
     3601 ttttgataat ctcatgacca aaatccctta acgtgagttt tcgttccact gagcgtcaga
     3661 ccccgtagaa aagatcaaag gatcttcttg agatcctttt tttctgcgcg taatctgctg
     3721 cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc aagagctacc
     3781 aactcttttt ccgaaggtaa ctggcttcag cagagcgcag ataccaaata ctgttcttct
     3841 agtgtagccg tagttaggcc accacttcaa gaactctgta gcaccgccta catacctcgc
     3901 tctgctaatc ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt
     3961 ggactcaaga cgatagttac cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg
     4021 cacacagccc agcttggagc gaacgaccta caccgaactg agatacctac agcgtgagct
     4081 atgagaaagc gccacgcttc ccgaagggag aaaggcggac aggtatccgg taagcggcag
     4141 ggtcggaaca ggagagcgca cgagggagct tccaggggga aacgcctggt atctttatag
     4201 tcctgtcggg tttcgccacc tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg
     4261 gcggagccta tggaaaaacg ccagcaacgc ggccttttta cggttcctgg ccttttgctg
     4321 gccttttgct cacatgttct ttcctgcgtt atcccctgat tctgtggata accgtattac
     4381 cgcctttgag tgagctgata ccgctcgccg cagccgaacg accgagcgca gcgagtcagt
     4441 gagcgaggaa gcggaagagc gcccaatacg caaaccgcct ctccccgcgc gttggccgat
     4501 tcattaatgc agctggcacg acaggtttcc cgactggaaa gcgggcagtg agcgcaacgc
     4561 aattaatgtg agttagctca ctcattaggc accccaggct ttacacttta tgcttccggc
     4621 tcgtatgttg tgtggaattg tgagcggata acaatttcac acaggaaaca gctatgacca
     4681 tgattacgcc aagcgcgcaa ttaaccctca ctaaagggaa caaaagctgg agctgcaagc
     4741 ttaatgtagt cttatgcaat actcttgtag tcttgcaaca tggtaacgat gagttagcaa
     4801 catgccttac aaggagagaa aaagcaccgt gcatgccgat tggtggaagt aaggtggtac
     4861 gatcgtgcct tattaggaag gcaacagacg ggtctgacat ggattggacg aaccactgaa
     4921 ttgccgcatt gcagagatat tgtatttaag tgcctagctc gatacataaa cgggtctctc
     4981 tggttagacc agatctgagc ctgggagctc tctggctaac tagggaaccc actgcttaag
     5041 cctcaataaa gcttgccttg agtgcttcaa gtagtgtgtg cccgtctgtt gtgtgactct
     5101 ggtaactaga gatccctcag acccttttag tcagtgtgga aaatctctag cagtggcgcc
     5161 cgaacaggga cttgaaagcg aaagggaaac cagaggagct ctctcgacgc aggactcggc
     5221 ttgctgaagc gcgcacggca agaggcgagg ggcggcgact ggtgagtacg ccaaaaattt
     5281 tgactagcgg aggctagaag gagagagatg ggtgcgagag cgtcagtatt aagcggggga
     5341 gaattagatc gcgatgggaa aaaattcggt taaggccagg gggaaagaaa aaatataaat
     5401 taaaacatat agtatgggca agcagggagc tagaacgatt cgcagttaat cctggcctgt
     5461 tagaaacatc agaaggctgt agacaaatac tgggacagct acaaccatcc cttcagacag
     5521 gatcagaaga acttagatca ttatataata cagtagcaac cctctattgt gtgcatcaaa
     5581 ggatagagat aaaagacacc aaggaagctt tagacaagat agaggaagag caaaacaaaa
     5641 gtaagaccac cgcacagcaa gcggccgctg atcttcagac ctggaggagg agatatgagg
     5701 gacaattgga gaagtgaatt atataaatat aaagtagtaa aaattgaacc attaggagta
     5761 gcacccacca aggcaaagag aagagtggtg cagagagaaa aaagagcagt gggaatagga
     5821 gctttgttcc ttgggttctt gggagcagca ggaagcacta tgggcgcagc gtcaatgacg
     5881 ctgacggtac aggccagaca attattgtct ggtatagtgc agcagcagaa caatttgctg
     5941 agggctattg aggcgcaaca gcatctgttg caactcacag tctggggcat caagcagctc
     6001 caggcaagaa tcctggctgt ggaaagatac ctaaaggatc aacagctcct ggggatttgg
     6061 ggttgctctg gaaaactcat ttgcaccact gctgtgcctt ggaatgctag ttggagtaat
     6121 aaatctctgg aacagatttg gaatcacacg acctggatgg agtgggacag agaaattaac
     6181 aattacacaa gcttaataca ctccttaatt gaagaatcgc aaaaccagca agaaaagaat
     6241 gaacaagaat tattggaatt agataaatgg gcaagtttgt ggaattggtt taacataaca
     6301 aattggctgt ggtatataaa attattcata atgatagtag gaggcttggt aggtttaaga
     6361 atagtttttg ctgtactttc tatagtgaat agagttaggc agggatattc accattatcg
     6421 tttcagaccc acctcccaac cccgagggga cccgacaggc ccgaaggaat agaagaagaa
     6481 ggtggagaga gagacagaga cagatccatt cgattagtga acggatctcg acggtatcga
     6541 tcacgagact agcctcgagc ggccgccccc ttcaccgagg gcctatttcc catgattcct
     6601 tcatatttgc atatacgata caaggctgtt agagagataa ttggaattaa tttgactgta
     6661 aacacaaaga tattagtaca aaatacgtga cgtagaaagt aataatttct tgggtagttt
     6721 gcagttttaa aattatgttt taaaatggac tatcatatgc ttaccgtaac ttgaaagtat
     6781 ttcgatttct tggctttata tatcttgtgg aaaggacgaa acaccgggcc agctttactg
     6841 gagaacttct cgagaagttc tccagtaaag ctggcttttt gaattctcga cctcgagaca
     6901 aatggcagta ttcatccaca attttaaaag aaaagggggg attggggggt acagtgcagg
     6961 ggaaagaata gtagacataa tagcaacaga catacaaact aaagaattac aaaaacaaat
     7021 tacaaaaatt caaaattttc gggtttatta cagggacagc agagatccac tttggccgcg
     7081 gctc
//
