LOCUS       sequence-193107-        7084 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Lentiviral expression of shRNA targeting BNIP3.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7084)
  TITLE     Gordon Lab plasmids
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"
COMMENT     Sequence Label: pLKO-sh-BNIP3-FL
FEATURES             Location/Qualifiers
     source          1..7084
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      21..609
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     510..529
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     763..780
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    897..918
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(933..938,939..956,957..963)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    971..987
                     /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     976..998
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     995..1011
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     995..1011
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     1030..1050
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        1032..1050
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        1078..1304
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             1305..1485
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    1532..1657
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    2150..2383
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             2568..2612
                     /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        2910..3158
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     2910..2930
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     3081..3100
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_feature    3255..3372
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        3421..3931
                     /label=hPGK promoter
                     /note="human phosphoglycerate kinase 1 promoter"
     primer_bind     3850..3868
                     /label=hPGK-F
                     /note="Human PGK promoter, forward primer"
     CDS             3953..4552
                     /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(3953..3972)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     4449..4469
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     LTR             4680..4913
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    4985..5106
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5022..5041)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5076..5095
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      5146..5281
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(5301..5320)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(5302..5320)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(5330..5347)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(5330..5346)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5339..5361)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      5488..5943
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5575..5594)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5785..5806
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        5969..6073
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(6074..6142,6143..6934)
                     /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(6292..6311)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 gaaaagatca aaggatcttc ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa
       61 acaaaaaaac caccgctacc agcggtggtt tgtttgccgg atcaagagct accaactctt
      121 tttccgaagg taactggctt cagcagagcg cagataccaa atactgttct tctagtgtag
      181 ccgtagttag gccaccactt caagaactct gtagcaccgc ctacatacct cgctctgcta
      241 atcctgttac cagtggctgc tgccagtggc gataagtcgt gtcttaccgg gttggactca
      301 agacgatagt taccggataa ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag
      361 cccagcttgg agcgaacgac ctacaccgaa ctgagatacc tacagcgtga gctatgagaa
      421 agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga
      481 acaggagagc gcacgaggga gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc
      541 gggtttcgcc acctctgact tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc
      601 ctatggaaaa acgccagcaa cgcggccttt ttacggttcc tggccttttg ctggcctttt
      661 gctcacatgt tctttcctgc gttatcccct gattctgtgg ataaccgtat taccgccttt
      721 gagtgagctg ataccgctcg ccgcagccga acgaccgagc gcagcgagtc agtgagcgag
      781 gaagcggaag agcgcccaat acgcaaaccg cctctccccg cgcgttggcc gattcattaa
      841 tgcagctggc acgacaggtt tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat
      901 gtgagttagc tcactcatta ggcaccccag gctttacact ttatgcttcc ggctcgtatg
      961 ttgtgtggaa ttgtgagcgg ataacaattt cacacaggaa acagctatga ccatgattac
     1021 gccaagcgcg caattaaccc tcactaaagg gaacaaaagc tggagctgca agcttaatgt
     1081 agtcttatgc aatactcttg tagtcttgca acatggtaac gatgagttag caacatgcct
     1141 tacaaggaga gaaaaagcac cgtgcatgcc gattggtgga agtaaggtgg tacgatcgtg
     1201 ccttattagg aaggcaacag acgggtctga catggattgg acgaaccact gaattgccgc
     1261 attgcagaga tattgtattt aagtgcctag ctcgatacat aaacgggtct ctctggttag
     1321 accagatctg agcctgggag ctctctggct aactagggaa cccactgctt aagcctcaat
     1381 aaagcttgcc ttgagtgctt caagtagtgt gtgcccgtct gttgtgtgac tctggtaact
     1441 agagatccct cagacccttt tagtcagtgt ggaaaatctc tagcagtggc gcccgaacag
     1501 ggacttgaaa gcgaaaggga aaccagagga gctctctcga cgcaggactc ggcttgctga
     1561 agcgcgcacg gcaagaggcg aggggcggcg actggtgagt acgccaaaaa ttttgactag
     1621 cggaggctag aaggagagag atgggtgcga gagcgtcagt attaagcggg ggagaattag
     1681 atcgcgatgg gaaaaaattc ggttaaggcc agggggaaag aaaaaatata aattaaaaca
     1741 tatagtatgg gcaagcaggg agctagaacg attcgcagtt aatcctggcc tgttagaaac
     1801 atcagaaggc tgtagacaaa tactgggaca gctacaacca tcccttcaga caggatcaga
     1861 agaacttaga tcattatata atacagtagc aaccctctat tgtgtgcatc aaaggataga
     1921 gataaaagac accaaggaag ctttagacaa gatagaggaa gagcaaaaca aaagtaagac
     1981 caccgcacag caagcggccg ctgatcttca gacctggagg aggagatatg agggacaatt
     2041 ggagaagtga attatataaa tataaagtag taaaaattga accattagga gtagcaccca
     2101 ccaaggcaaa gagaagagtg gtgcagagag aaaaaagagc agtgggaata ggagctttgt
     2161 tccttgggtt cttgggagca gcaggaagca ctatgggcgc agcgtcaatg acgctgacgg
     2221 tacaggccag acaattattg tctggtatag tgcagcagca gaacaatttg ctgagggcta
     2281 ttgaggcgca acagcatctg ttgcaactca cagtctgggg catcaagcag ctccaggcaa
     2341 gaatcctggc tgtggaaaga tacctaaagg atcaacagct cctggggatt tggggttgct
     2401 ctggaaaact catttgcacc actgctgtgc cttggaatgc tagttggagt aataaatctc
     2461 tggaacagat ttggaatcac acgacctgga tggagtggga cagagaaatt aacaattaca
     2521 caagcttaat acactcctta attgaagaat cgcaaaacca gcaagaaaag aatgaacaag
     2581 aattattgga attagataaa tgggcaagtt tgtggaattg gtttaacata acaaattggc
     2641 tgtggtatat aaaattattc ataatgatag taggaggctt ggtaggttta agaatagttt
     2701 ttgctgtact ttctatagtg aatagagtta ggcagggata ttcaccatta tcgtttcaga
     2761 cccacctccc aaccccgagg ggacccgaca ggcccgaagg aatagaagaa gaaggtggag
     2821 agagagacag agacagatcc attcgattag tgaacggatc tcgacggtat cgatcacgag
     2881 actagcctcg agcggccgcc cccttcaccg agggcctatt tcccatgatt ccttcatatt
     2941 tgcatatacg atacaaggct gttagagaga taattggaat taatttgact gtaaacacaa
     3001 agatattagt acaaaatacg tgacgtagaa agtaataatt tcttgggtag tttgcagttt
     3061 taaaattatg ttttaaaatg gactatcata tgcttaccgt aacttgaaag tatttcgatt
     3121 tcttggcttt atatatcttg tggaaaggac gaaacaccgg aagacaccac aagataccaa
     3181 cctcgaggtt ggtatcttgt ggtgtctttt tttgaattct cgacctcgag acaaatggca
     3241 gtattcatcc acaattttaa aagaaaaggg gggattgggg ggtacagtgc aggggaaaga
     3301 atagtagaca taatagcaac agacatacaa actaaagaat tacaaaaaca aattacaaaa
     3361 attcaaaatt ttcgggttta ttacagggac agcagagatc cactttggcc gcggctcgag
     3421 ggggttgggg ttgcgccttt tccaaggcag ccctgggttt gcgcagggac gcggctgctc
     3481 tgggcgtggt tccgggaaac gcagcggcgc cgaccctggg tctcgcacat tcttcacgtc
     3541 cgttcgcagc gtcacccgga tcttcgccgc tacccttgtg ggccccccgg cgacgcttcc
     3601 tgctccgccc ctaagtcggg aaggttcctt gcggttcgcg gcgtgccgga cgtgacaaac
     3661 ggaagccgca cgtctcacta gtaccctcgc agacggacag cgccagggag caatggcagc
     3721 gcgccgaccg cgatgggctg tggccaatag cggctgctca gcagggcgcg ccgagagcag
     3781 cggccgggaa ggggcggtgc gggaggcggg gtgtggggcg gtagtgtggg ccctgttcct
     3841 gcccgcgcgg tgttccgcat tctgcaagcc tccggagcgc acgtcggcag tcggctccct
     3901 cgttgaccga atcaccgacc tctctcccca gggggatcca ccggagctta ccatgaccga
     3961 gtacaagccc acggtgcgcc tcgccacccg cgacgacgtc cccagggccg tacgcaccct
     4021 cgccgccgcg ttcgccgact accccgccac gcgccacacc gtcgatccgg accgccacat
     4081 cgagcgggtc accgagctgc aagaactctt cctcacgcgc gtcgggctcg acatcggcaa
     4141 ggtgtgggtc gcggacgacg gcgccgcggt ggcggtctgg accacgccgg agagcgtcga
     4201 agcgggggcg gtgttcgccg agatcggccc gcgcatggcc gagttgagcg gttcccggct
     4261 ggccgcgcag caacagatgg aaggcctcct ggcgccgcac cggcccaagg agcccgcgtg
     4321 gttcctggcc accgtcggcg tctcgcccga ccaccagggc aagggtctgg gcagcgccgt
     4381 cgtgctcccc ggagtggagg cggccgagcg cgccggggtg cccgccttcc tggagacctc
     4441 cgcgccccgc aacctcccct tctacgagcg gctcggcttc accgtcaccg ccgacgtcga
     4501 ggtgcccgaa ggaccgcgca cctggtgcat gacccgcaag cccggtgcct gacgcccgcc
     4561 ccacgacccg cagcgcccga ccgaaaggag cgcacgaccc catgcatcgg tacctttaag
     4621 accaatgact tacaaggcag ctgtagatct tagccacttt ttaaaagaaa aggggggact
     4681 ggaagggcta attcactccc aacgaagaca agatctgctt tttgcttgta ctgggtctct
     4741 ctggttagac cagatctgag cctgggagct ctctggctaa ctagggaacc cactgcttaa
     4801 gcctcaataa agcttgcctt gagtgcttca agtagtgtgt gcccgtctgt tgtgtgactc
     4861 tggtaactag agatccctca gaccctttta gtcagtgtgg aaaatctcta gcagtagtag
     4921 ttcatgtcat cttattattc agtatttata acttgcaaag aaatgaatat cagagagtga
     4981 gaggaacttg tttattgcag cttataatgg ttacaaataa agcaatagca tcacaaattt
     5041 cacaaataaa gcattttttt cactgcattc tagttgtggt ttgtccaaac tcatcaatgt
     5101 atcttatcat gtctggctct agctatcccg cccctaactc cgcccatccc gcccctaact
     5161 ccgcccagtt ccgcccattc tccgccccat ggctgactaa ttttttttat ttatgcagag
     5221 gccgaggccg cctcggcctc tgagctattc cagaagtagt gaggaggctt ttttggaggc
     5281 ctagggacgt acccaattcg ccctatagtg agtcgtatta cgcgcgctca ctggccgtcg
     5341 ttttacaacg tcgtgactgg gaaaaccctg gcgttaccca acttaatcgc cttgcagcac
     5401 atcccccttt cgccagctgg cgtaatagcg aagaggcccg caccgatcgc ccttcccaac
     5461 agttgcgcag cctgaatggc gaatgggacg cgccctgtag cggcgcatta agcgcggcgg
     5521 gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag cgccctagcg cccgctcctt
     5581 tcgctttctt cccttccttt ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc
     5641 gggggctccc tttagggttc cgatttagtg ctttacggca cctcgacccc aaaaaacttg
     5701 attagggtga tggttcacgt agtgggccat cgccctgata gacggttttt cgccctttga
     5761 cgttggagtc cacgttcttt aatagtggac tcttgttcca aactggaaca acactcaacc
     5821 ctatctcggt ctattctttt gatttataag ggattttgcc gatttcggcc tattggttaa
     5881 aaaatgagct gatttaacaa aaatttaacg cgaattttaa caaaatatta acgcttacaa
     5941 tttaggtggc acttttcggg gaaatgtgcg cggaacccct atttgtttat ttttctaaat
     6001 acattcaaat atgtatccgc tcatgagaca ataaccctga taaatgcttc aataatattg
     6061 aaaaaggaag agtatgagta ttcaacattt ccgtgtcgcc cttattccct tttttgcggc
     6121 attttgcctt cctgtttttg ctcacccaga aacgctggtg aaagtaaaag atgctgaaga
     6181 tcagttgggt gcacgagtgg gttacatcga actggatctc aacagcggta agatccttga
     6241 gagttttcgc cccgaagaac gttttccaat gatgagcact tttaaagttc tgctatgtgg
     6301 cgcggtatta tcccgtattg acgccgggca agagcaactc ggtcgccgca tacactattc
     6361 tcagaatgac ttggttgagt actcaccagt cacagaaaag catcttacgg atggcatgac
     6421 agtaagagaa ttatgcagtg ctgccataac catgagtgat aacactgcgg ccaacttact
     6481 tctgacaacg atcggaggac cgaaggagct aaccgctttt ttgcacaaca tgggggatca
     6541 tgtaactcgc cttgatcgtt gggaaccgga gctgaatgaa gccataccaa acgacgagcg
     6601 tgacaccacg atgcctgtag caatggcaac aacgttgcgc aaactattaa ctggcgaact
     6661 acttactcta gcttcccggc aacaattaat agactggatg gaggcggata aagttgcagg
     6721 accacttctg cgctcggccc ttccggctgg ctggtttatt gctgataaat ctggagccgg
     6781 tgagcgtggg tctcgcggta tcattgcagc actggggcca gatggtaagc cctcccgtat
     6841 cgtagttatc tacacgacgg ggagtcaggc aactatggat gaacgaaata gacagatcgc
     6901 tgagataggt gcctcactga ttaagcattg gtaactgtca gaccaagttt actcatatat
     6961 actttagatt gatttaaaac ttcattttta atttaaaagg atctaggtga agatcctttt
     7021 tgataatctc atgaccaaaa tcccttaacg tgagttttcg ttccactgag cgtcagaccc
     7081 cgta
//
