LOCUS       sequence-364242-        6494 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Control lentivector that can be used to express inserts from the PGK
            promoter and/or shRNA cassettes from U6..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6494)
  AUTHORS   Greiner D, Scott TM, Olson GS, Aderem A, Roh-Johnson M, Johnson JS
  TITLE     Genetic Modification of Primary Human Myeloid Cells to Study Cell 
            Migration, Activation, and Organelle Dynamics.
  JOURNAL   Curr Protoc. 2022 Aug;2(8):e514. doi: 10.1002/cpz1.514.
  PUBMED    36018279
REFERENCE   2  (bases 1 to 6494)
  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: "Curr 
            Protoc."; date: "2022-08"; volume: "2(8):e514. doi"; pages: " 
            10.1002/cpz1.514"
COMMENT     Sequence Label: pLKO-MCS
FEATURES             Location/Qualifiers
     source          1..6494
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     426..444
                     /label=hPGK-F
                     /note="Human PGK promoter, forward primer"
     LTR             700..933
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    1005..1126
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1042..1061)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1096..1115
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      1166..1301
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(1321..1340)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1322..1340)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1350..1367)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(1350..1366)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1359..1381)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      1508..1963
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1595..1614)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1805..1826
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        1989..2093
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2094..2162,2163..2954)
                     /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(2312..2331)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3125..3713
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3614..3633
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3867..3884
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    4001..4022
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(4037..4042,4043..4060,4061..4067)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4075..4091
                     /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     4080..4102
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     4099..4115
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     4099..4115
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     4134..4154
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        4136..4154
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        4182..4408
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             4409..4589
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    4636..4761
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    5254..5487
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             5672..5716
                     /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        6014..6262
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     6014..6034
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     6185..6204
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_feature    6325..6442
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        join(6491..6494,1..507)
                     /label=hPGK promoter
                     /note="human phosphoglycerate kinase 1 promoter"
ORIGIN
        1 ttggggttgc gccttttcca aggcagccct gggtttgcgc agggacgcgg ctgctctggg
       61 cgtggttccg ggaaacgcag cggcgccgac cctgggtctc gcacattctt cacgtccgtt
      121 cgcagcgtca cccggatctt cgccgctacc cttgtgggcc ccccggcgac gcttcctgct
      181 ccgcccctaa gtcgggaagg ttccttgcgg ttcgcggcgt gccggacgtg acaaacggaa
      241 gccgcacgtc tcactagtac cctcgcagac ggacagcgcc agggagcaat ggcagcgcgc
      301 cgaccgcgat gggctgtggc caatagcggc tgctcagcag ggcgcgccga gagcagcggc
      361 cgggaagggg cggtgcggga ggcggggtgt ggggcggtag tgtgggccct gttcctgccc
      421 gcgcggtgtt ccgcattctg caagcctccg gagcgcacgt cggcagtcgg ctccctcgtt
      481 gaccgaatca ccgacctctc tccccagggg gatccgacgt cttaattaac ctgcagggtc
      541 gacacgcgtc gtacggatat ctgtacaagt aacgcccgcc ccacgacccg cagcgcccga
      601 ccgaaaggag cgcacgaccc catgcatcgg tacctttaag accaatgact tacaaggcag
      661 ctgtagatct tagccacttt ttaaaagaaa aggggggact ggaagggcta attcactccc
      721 aacgaagaca agatctgctt tttgcttgta ctgggtctct ctggttagac cagatctgag
      781 cctgggagct ctctggctaa ctagggaacc cactgcttaa gcctcaataa agcttgcctt
      841 gagtgcttca agtagtgtgt gcccgtctgt tgtgtgactc tggtaactag agatccctca
      901 gaccctttta gtcagtgtgg aaaatctcta gcagtagtag ttcatgtcat cttattattc
      961 agtatttata acttgcaaag aaatgaatat cagagagtga gaggaacttg tttattgcag
     1021 cttataatgg ttacaaataa agcaatagca tcacaaattt cacaaataaa gcattttttt
     1081 cactgcattc tagttgtggt ttgtccaaac tcatcaatgt atcttatcat gtctggctct
     1141 agctatcccg cccctaactc cgcccatccc gcccctaact ccgcccagtt ccgcccattc
     1201 tccgccccat ggctgactaa ttttttttat ttatgcagag gccgaggccg cctcggcctc
     1261 tgagctattc cagaagtagt gaggaggctt ttttggaggc ctagggacgt acccaattcg
     1321 ccctatagtg agtcgtatta cgcgcgctca ctggccgtcg ttttacaacg tcgtgactgg
     1381 gaaaaccctg gcgttaccca acttaatcgc cttgcagcac atcccccttt cgccagctgg
     1441 cgtaatagcg aagaggcccg caccgatcgc ccttcccaac agttgcgcag cctgaatggc
     1501 gaatgggacg cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc
     1561 gtgaccgcta cacttgccag cgccctagcg cccgctcctt tcgctttctt cccttccttt
     1621 ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc gggggctccc tttagggttc
     1681 cgatttagtg ctttacggca cctcgacccc aaaaaacttg attagggtga tggttcacgt
     1741 agtgggccat cgccctgata gacggttttt cgccctttga cgttggagtc cacgttcttt
     1801 aatagtggac tcttgttcca aactggaaca acactcaacc ctatctcggt ctattctttt
     1861 gatttataag ggattttgcc gatttcggcc tattggttaa aaaatgagct gatttaacaa
     1921 aaatttaacg cgaattttaa caaaatatta acgcttacaa tttaggtggc acttttcggg
     1981 gaaatgtgcg cggaacccct atttgtttat ttttctaaat acattcaaat atgtatccgc
     2041 tcatgagaca ataaccctga taaatgcttc aataatattg aaaaaggaag agtatgagta
     2101 ttcaacattt ccgtgtcgcc cttattccct tttttgcggc attttgcctt cctgtttttg
     2161 ctcacccaga aacgctggtg aaagtaaaag atgctgaaga tcagttgggt gcacgagtgg
     2221 gttacatcga actggatctc aacagcggta agatccttga gagttttcgc cccgaagaac
     2281 gttttccaat gatgagcact tttaaagttc tgctatgtgg cgcggtatta tcccgtattg
     2341 acgccgggca agagcaactc ggtcgccgca tacactattc tcagaatgac ttggttgagt
     2401 actcaccagt cacagaaaag catcttacgg atggcatgac agtaagagaa ttatgcagtg
     2461 ctgccataac catgagtgat aacactgcgg ccaacttact tctgacaacg atcggaggac
     2521 cgaaggagct aaccgctttt ttgcacaaca tgggggatca tgtaactcgc cttgatcgtt
     2581 gggaaccgga gctgaatgaa gccataccaa acgacgagcg tgacaccacg atgcctgtag
     2641 caatggcaac aacgttgcgc aaactattaa ctggcgaact acttactcta gcttcccggc
     2701 aacaattaat agactggatg gaggcggata aagttgcagg accacttctg cgctcggccc
     2761 ttccggctgg ctggtttatt gctgataaat ctggagccgg tgagcgtggg tctcgcggta
     2821 tcattgcagc actggggcca gatggtaagc cctcccgtat cgtagttatc tacacgacgg
     2881 ggagtcaggc aactatggat gaacgaaata gacagatcgc tgagataggt gcctcactga
     2941 ttaagcattg gtaactgtca gaccaagttt actcatatat actttagatt gatttaaaac
     3001 ttcattttta atttaaaagg atctaggtga agatcctttt tgataatctc atgaccaaaa
     3061 tcccttaacg tgagttttcg ttccactgag cgtcagaccc cgtagaaaag atcaaaggat
     3121 cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc
     3181 taccagcggt ggtttgtttg ccggatcaag agctaccaac tctttttccg aaggtaactg
     3241 gcttcagcag agcgcagata ccaaatactg ttcttctagt gtagccgtag ttaggccacc
     3301 acttcaagaa ctctgtagca ccgcctacat acctcgctct gctaatcctg ttaccagtgg
     3361 ctgctgccag tggcgataag tcgtgtctta ccgggttgga ctcaagacga tagttaccgg
     3421 ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa
     3481 cgacctacac cgaactgaga tacctacagc gtgagctatg agaaagcgcc acgcttcccg
     3541 aagggagaaa ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga
     3601 gggagcttcc agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct
     3661 gacttgagcg tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaaaacgcca
     3721 gcaacgcggc ctttttacgg ttcctggcct tttgctggcc ttttgctcac atgttctttc
     3781 ctgcgttatc ccctgattct gtggataacc gtattaccgc ctttgagtga gctgataccg
     3841 ctcgccgcag ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg gaagagcgcc
     3901 caatacgcaa accgcctctc cccgcgcgtt ggccgattca ttaatgcagc tggcacgaca
     3961 ggtttcccga ctggaaagcg ggcagtgagc gcaacgcaat taatgtgagt tagctcactc
     4021 attaggcacc ccaggcttta cactttatgc ttccggctcg tatgttgtgt ggaattgtga
     4081 gcggataaca atttcacaca ggaaacagct atgaccatga ttacgccaag cgcgcaatta
     4141 accctcacta aagggaacaa aagctggagc tgcaagctta atgtagtctt atgcaatact
     4201 cttgtagtct tgcaacatgg taacgatgag ttagcaacat gccttacaag gagagaaaaa
     4261 gcaccgtgca tgccgattgg tggaagtaag gtggtacgat cgtgccttat taggaaggca
     4321 acagacgggt ctgacatgga ttggacgaac cactgaattg ccgcattgca gagatattgt
     4381 atttaagtgc ctagctcgat acataaacgg gtctctctgg ttagaccaga tctgagcctg
     4441 ggagctctct ggctaactag ggaacccact gcttaagcct caataaagct tgccttgagt
     4501 gcttcaagta gtgtgtgccc gtctgttgtg tgactctggt aactagagat ccctcagacc
     4561 cttttagtca gtgtggaaaa tctctagcag tggcgcccga acagggactt gaaagcgaaa
     4621 gggaaaccag aggagctctc tcgacgcagg actcggcttg ctgaagcgcg cacggcaaga
     4681 ggcgaggggc ggcgactggt gagtacgcca aaaattttga ctagcggagg ctagaaggag
     4741 agagatgggt gcgagagcgt cagtattaag cgggggagaa ttagatcgcg atgggaaaaa
     4801 attcggttaa ggccaggggg aaagaaaaaa tataaattaa aacatatagt atgggcaagc
     4861 agggagctag aacgattcgc agttaatcct ggcctgttag aaacatcaga aggctgtaga
     4921 caaatactgg gacagctaca accatccctt cagacaggat cagaagaact tagatcatta
     4981 tataatacag tagcaaccct ctattgtgtg catcaaagga tagagataaa agacaccaag
     5041 gaagctttag acaagataga ggaagagcaa aacaaaagta agaccaccgc acagcaagcg
     5101 gccgctgatc ttcagacctg gaggaggaga tatgagggac aattggagaa gtgaattata
     5161 taaatataaa gtagtaaaaa ttgaaccatt aggagtagca cccaccaagg caaagagaag
     5221 agtggtgcag agagaaaaaa gagcagtggg aataggagct ttgttccttg ggttcttggg
     5281 agcagcagga agcactatgg gcgcagcgtc aatgacgctg acggtacagg ccagacaatt
     5341 attgtctggt atagtgcagc agcagaacaa tttgctgagg gctattgagg cgcaacagca
     5401 tctgttgcaa ctcacagtct ggggcatcaa gcagctccag gcaagaatcc tggctgtgga
     5461 aagataccta aaggatcaac agctcctggg gatttggggt tgctctggaa aactcatttg
     5521 caccactgct gtgccttgga atgctagttg gagtaataaa tctctggaac agatttggaa
     5581 tcacacgacc tggatggagt gggacagaga aattaacaat tacacaagct taatacactc
     5641 cttaattgaa gaatcgcaaa accagcaaga aaagaatgaa caagaattat tggaattaga
     5701 taaatgggca agtttgtgga attggtttaa cataacaaat tggctgtggt atataaaatt
     5761 attcataatg atagtaggag gcttggtagg tttaagaata gtttttgctg tactttctat
     5821 agtgaataga gttaggcagg gatattcacc attatcgttt cagacccacc tcccaacccc
     5881 gaggggaccc gacaggcccg aaggaataga agaagaaggt ggagagagag acagagacag
     5941 atccattcga ttagtgaacg gatctcgacg gtatcgatca cgagactagc ctcgagcggc
     6001 cgcccccttc accgagggcc tatttcccat gattccttca tatttgcata tacgatacaa
     6061 ggctgttaga gagataattg gaattaattt gactgtaaac acaaagatat tagtacaaaa
     6121 tacgtgacgt agaaagtaat aatttcttgg gtagtttgca gttttaaaat tatgttttaa
     6181 aatggactat catatgctta ccgtaacttg aaagtatttc gatttcttgg ctttatatat
     6241 cttgtggaaa ggacgaaaca ccggtccgca ggtatgcacg cgtgaattct cgacctcgag
     6301 acaaatggca gtattcatcc acaattttaa aagaaaaggg gggattgggg ggtacagtgc
     6361 aggggaaaga atagtagaca taatagcaac agacatacaa actaaagaat tacaaaaaca
     6421 aattacaaaa attcaaaatt ttcgggttta ttacagggac agcagagatc cactttggcc
     6481 gcggctcgag gggg
//
