LOCUS       sequence-171420-        7486 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Cre-regulated lentiviral shRNA vector. Cre addition causes the EGFP 
            gene to be recombined out of the construct, activating shRNA 
            expression..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7486)
  AUTHORS   Ventura A, Meissner A, Dillon CP, McManus M, Sharp PA, Van Parijs L,
            Jaenisch R, Jacks T
  TITLE     Cre-lox-regulated conditional RNA interference from transgenes.
  JOURNAL   Proc Natl Acad Sci U S A. 2004 Jul 13. 101(28):10380-5.
  PUBMED    15240889
REFERENCE   2  (bases 1 to 7486)
  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: "Proc Natl 
            Acad Sci U S A. 2004 Jul 13. 101(28):10380-5."
COMMENT     Sequence Label: pSico PGK GFP
FEATURES             Location/Qualifiers
     source          1..7486
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     40..59
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     regulatory      135..144
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(141..143,144..146,147..860)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(141..143,144..146,147..857)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(186..207)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(447..466)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     794..815
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     protein_bind    complement(868..901)
                     /label=loxP511
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATAC) (Shaw et al., 2021). loxP511 sites are 
                     compatible with each other, but incompatible with wild-type
                     loxP sites."
     primer_bind     928..944
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     misc_feature    1019..1607
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(1072..1092)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(1490..1501)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(1610..1626)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     LTR             2136..2316
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     rep_origin      complement(2378..2966)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2458..2477)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3137..3928,3929..3997))
                     /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     3760..3779
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3998..4102)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(4177..4196)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        4368..4747
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        4749..4947
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     4902..4922
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     LTR             4965..5145
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    5192..5317
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    5814..6047
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             6232..6276
                     /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"
     misc_feature    6574..6691
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     primer_bind     6981..7001
                     /label=mU6-F
                     /note="Mouse U6 promoter, forward primer"
     protein_bind    complement(7019..7052)
                     /label=loxP511
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATAC) (Shaw et al., 2021). loxP511 sites are 
                     compatible with each other, but incompatible with wild-type
                     loxP sites."
     promoter        join(7094..7486,1..107)
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     complement(7129..7152)
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
ORIGIN
        1 ggggcggggc gggcgcccga aggtcctccg gaggcccggc attctgcacg cttcaaaagc
       61 gcacgtctgc cgcgctgttc tcctcttcct catctccggg cctttcgacc tgcatccatc
      121 tagatctccc ggtcgccacc atggtgagca agggcgagga gctgttcacc ggggtggtgc
      181 ccatcctggt cgagctggac ggcgacgtaa acggccacaa gttcagcgtg tccggcgagg
      241 gcgagggcga tgccacctac ggcaagctga ccctgaagtt catctgcacc accggcaagc
      301 tgcccgtgcc ctggcccacc ctcgtgacca ccctgaccta cggcgtgcag tgcttcagcc
      361 gctaccccga ccacatgaag cagcacgact tcttcaagtc cgccatgccc gaaggctacg
      421 tccaggagcg caccatcttc ttcaaggacg acggcaacta caagacccgc gccgaggtga
      481 agttcgaggg cgacaccctg gtgaaccgca tcgagctgaa gggcatcgac ttcaaggagg
      541 acggcaacat cctggggcac aagctggagt acaactacaa cagccacaac gtctatatca
      601 tggccgacaa gcagaagaac ggcatcaagg tgaacttcaa gatccgccac aacatcgagg
      661 acggcagcgt gcagctcgcc gaccactacc agcagaacac ccccatcggc gacggccccg
      721 tgctgctgcc cgacaaccac tacctgagca cccagtccgc cctgagcaaa gaccccaacg
      781 agaagcgcga tcacatggtc ctgctggagt tcgtgaccgc cgccgggatc actctcggca
      841 tggacgagct gtacaagtag cggccgcata acttcgtata gtataaatta tacgaagtta
      901 taagccttgt taacgcgcgg tgaccctcga ggtcgacggt atcgataagc tcgcttcacg
      961 agatcatgtt taagggttcc ggttccacta ggtacaattc gatatcaagc ttatcgataa
     1021 tcaacctctg gattacaaaa tttgtgaaag attgactggt attcttaact atgttgctcc
     1081 ttttacgcta tgtggatacg ctgctttaat gcctttgtat catgctattg cttcccgtat
     1141 ggctttcatt ttctcctcct tgtataaatc ctggttgctg tctctttatg aggagttgtg
     1201 gcccgttgtc aggcaacgtg gcgtggtgtg cactgtgttt gctgacgcaa cccccactgg
     1261 ttggggcatt gccaccacct gtcagctcct ttccgggact ttcgctttcc ccctccctat
     1321 tgccacggcg gaactcatcg ccgcctgcct tgcccgctgc tggacagggg ctcggctgtt
     1381 gggcactgac aattccgtgg tgttgtcggg gaaatcatcg tcctttcctt ggctgctcgc
     1441 ctgtgttgcc acctggattc tgcgcgggac gtccttctgc tacgtccctt cggccctcaa
     1501 tccagcggac cttccttccc gcggcctgct gccggctctg cggcctcttc cgcgtcttcg
     1561 ccttcgccct cagacgagtc ggatctccct ttgggccgcc tccccgcatc gataccgtcg
     1621 acctcgatcg agacctagaa aaacatggag caatcacaag tagcaataca gcagctacca
     1681 atgctgattg tgcctggcta gaagcacaag aggaggagga ggtgggtttt ccagtcacac
     1741 ctcaggtacc tttaagacca atgacttaca aggcagctgt agatcttagc cactttttaa
     1801 aagaaaaggg gggactggaa gggctaattc actcccaacg aagacaagat atccttgatc
     1861 tgtggatcta ccacacacaa ggctacttcc ctgattggca gaactacaca ccagggccag
     1921 ggatcagata tccactgacc tttggatggt gctacaagct agtaccagtt gagcaagaga
     1981 aggtagaaga agccaatgaa ggagagaaca cccgcttgtt acaccctgtg agcctgcatg
     2041 ggatggatga cccggagaga gaagtattag agtggaggtt tgacagccgc ctagcatttc
     2101 atcacatggc ccgagagctg catccggact gtactgggtc tctctggtta gaccagatct
     2161 gagcctggga gctctctggc taactaggga acccactgct taagcctcaa taaagcttgc
     2221 cttgagtgct tcaagtagtg tgtgcccgtc tgttgtgtga ctctggtaac tagagatccc
     2281 tcagaccctt ttagtcagtg tggaaaatct ctagcagcat gtgagcaaaa ggccagcaaa
     2341 aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc cgcccccctg
     2401 acgagcatca caaaaatcga cgctcaagtc agaggtggcg aaacccgaca ggactataaa
     2461 gataccaggc gtttccccct ggaagctccc tcgtgcgctc tcctgttccg accctgccgc
     2521 ttaccggata cctgtccgcc tttctccctt cgggaagcgt ggcgctttct catagctcac
     2581 gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac
     2641 cccccgttca gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag tccaacccgg
     2701 taagacacga cttatcgcca ctggcagcag ccactggtaa caggattagc agagcgaggt
     2761 atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa ctacggctac actagaagaa
     2821 cagtatttgg tatctgcgct ctgctgaagc cagttacctt cggaaaaaga gttggtagct
     2881 cttgatccgg caaacaaacc accgctggta gcggtggttt ttttgtttgc aagcagcaga
     2941 ttacgcgcag aaaaaaagga tctcaagaag atcctttgat cttttctacg gggtctgacg
     3001 ctcagtggaa cgaaaactca cgttaaggga ttttggtcat gagattatca aaaaggatct
     3061 tcacctagat ccttttaaat taaaaatgaa gttttaaatc aatctaaagt atatatgagt
     3121 aaacttggtc tgacagttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc
     3181 tatttcgttc atccatagtt gcctgactcc ccgtcgtgta gataactacg atacgggagg
     3241 gcttaccatc tggccccagt gctgcaatga taccgcgaga cccacgctca ccggctccag
     3301 atttatcagc aataaaccag ccagccggaa gggccgagcg cagaagtggt cctgcaactt
     3361 tatccgcctc catccagtct attaattgtt gccgggaagc tagagtaagt agttcgccag
     3421 ttaatagttt gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt
     3481 ttggtatggc ttcattcagc tccggttccc aacgatcaag gcgagttaca tgatccccca
     3541 tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat cgttgtcaga agtaagttgg
     3601 ccgcagtgtt atcactcatg gttatggcag cactgcataa ttctcttact gtcatgccat
     3661 ccgtaagatg cttttctgtg actggtgagt actcaaccaa gtcattctga gaatagtgta
     3721 tgcggcgacc gagttgctct tgcccggcgt caatacggga taataccgcg ccacatagca
     3781 gaactttaaa agtgctcatc attggaaaac gttcttcggg gcgaaaactc tcaaggatct
     3841 taccgctgtt gagatccagt tcgatgtaac ccactcgtgc acccaactga tcttcagcat
     3901 cttttacttt caccagcgtt tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa
     3961 agggaataag ggcgacacgg aaatgttgaa tactcatact cttccttttt caatattatt
     4021 gaagcattta tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa
     4081 ataaacaaat aggggttccg cgcacatttc cccgaaaagt gccacctgac gtcgacggat
     4141 cgggagatct cccgatcccc tatggtgcac tctcagtaca atctgctctg atgccgcata
     4201 gttaagccag tatctgctcc ctgcttgtgt gttggaggtc gctgagtagt gcgcgagcaa
     4261 aatttaagct acaacaaggc aaggcttgac cgacaattgc atgaagaatc tgcttagggt
     4321 taggcgtttt gcgctgcttc gcgatgtacg ggccagatat acgcgttgac attgattatt
     4381 gactagttat taatagtaat caattacggg gtcattagtt catagcccat atatggagtt
     4441 ccgcgttaca taacttacgg taaatggccc gcctggctga ccgcccaacg acccccgccc
     4501 attgacgtca ataatgacgt atgttcccat agtaacgcca atagggactt tccattgacg
     4561 tcaatgggtg gagtatttac ggtaaactgc ccacttggca gtacatcaag tgtatcatat
     4621 gccaagtacg ccccctattg acgtcaatga cggtaaatgg cccgcctggc attatgccca
     4681 gtacatgacc ttatgggact ttcctacttg gcagtacatc tacgtattag tcatcgctat
     4741 taccatggtg atgcggtttt ggcagtacat caatgggcgt ggatagcggt ttgactcacg
     4801 gggatttcca agtctccacc ccattgacgt caatgggagt ttgttttggc accaaaatca
     4861 acgggacttt ccaaaatgtc gtaacaactc cgccccattg acgcaaatgg gcggtaggcg
     4921 tgtacggtgg gaggtctata taagcagcgc gttttgcctg tactgggtct ctctggttag
     4981 accagatctg agcctgggag ctctctggct aactagggaa cccactgctt aagcctcaat
     5041 aaagcttgcc ttgagtgctt caagtagtgt gtgcccgtct gttgtgtgac tctggtaact
     5101 agagatccct cagacccttt tagtcagtgt ggaaaatctc tagcagtggc gcccgaacag
     5161 ggacttgaaa gcgaaaggga aaccagagga gctctctcga cgcaggactc ggcttgctga
     5221 agcgcgcacg gcaagaggcg aggggcggcg actggtgagt acgccaaaaa ttttgactag
     5281 cggaggctag aaggagagag atgggtgcga gagcgtcagt attaagcggg ggagaattag
     5341 atcgcgatgg gaaaaaattc ggttaaggcc agggggaaag aaaaaatata aattaaaaca
     5401 tatagtatgg gcaagcaggg agctagaacg attcgcagtt aatcctggcc tgttagaaac
     5461 atcagaaggc tgtagacaaa tactgggaca gctacaacca tcccttcaga caggatcaga
     5521 agaacttaga tcattatata atacagtagc aaccctctat tgtgtgcatc aaaggataga
     5581 gataaaagac accaaggaag ctttagacaa gatagaggaa gagcaaaaca aaagtaagac
     5641 caccgcacag caagcggccg gccgctgatc ttcagacctg gaggaggaga tatgagggac
     5701 aattggagaa gtgaattata taaatataaa gtagtaaaaa ttgaaccatt aggagtagca
     5761 cccaccaagg caaagagaag agtggtgcag agagaaaaaa gagcagtggg aataggagct
     5821 ttgttccttg ggttcttggg agcagcagga agcactatgg gcgcagcgtc aatgacgctg
     5881 acggtacagg ccagacaatt attgtctggt atagtgcagc agcagaacaa tttgctgagg
     5941 gctattgagg cgcaacagca tctgttgcaa ctcacagtct ggggcatcaa gcagctccag
     6001 gcaagaatcc tggctgtgga aagataccta aaggatcaac agctcctggg gatttggggt
     6061 tgctctggaa aactcatttg caccactgct gtgccttgga atgctagttg gagtaataaa
     6121 tctctggaac agatttggaa tcacacgacc tggatggagt gggacagaga aattaacaat
     6181 tacacaagct taatacactc cttaattgaa gaatcgcaaa accagcaaga aaagaatgaa
     6241 caagaattat tggaattaga taaatgggca agtttgtgga attggtttaa cataacaaat
     6301 tggctgtggt atataaaatt attcataatg atagtaggag gcttggtagg tttaagaata
     6361 gtttttgctg tactttctat agtgaataga gttaggcagg gatattcacc attatcgttt
     6421 cagacccacc tcccaacccc gaggggaccc gacaggcccg aaggaataga agaagaaggt
     6481 ggagagagag acagagacag atccattcga ttagtgaacg gatcggcact gcgtgcgcca
     6541 attctgcaga caaatggcag tattcatcca caattttaaa agaaaagggg ggattggggg
     6601 gtacagtgca ggggaaagaa tagtagacat aatagcaaca gacatacaaa ctaaagaatt
     6661 acaaaaacaa attacaaaaa ttcaaaattt tcgggtttat tacagggaca gcagagatcc
     6721 agtttggtta gtaccgggcc cgctctagag atccgacgcg ccatctctag gcccgcgccg
     6781 gccccctcgc acagacttgt gggagaagct cggctactcc cctgccccgg ttaatttgca
     6841 tataatattt cctagtaact atagaggctt aatgtgcgat aaaagacaga taatctgttc
     6901 tttttaatac tagctacatt ttacatgata ggcttggatt tctataagag atacaaatac
     6961 taaattatta ttttaaaaaa cagcacaaaa ggaaactcac cctaactgta aagtaattat
     7021 aacttcgtat agtataaatt atacgaagtt ataagccttg gttttttgaa ttcgtcgacc
     7081 tcgaaattct accgggtagg ggaggcgctt ttcccaaggc agtctggagc atgcgcttta
     7141 gcagccccgc tgggcacttg gcgctacaca agtggcctct ggcctcgcac acattccaca
     7201 tccaccggta ggcgccaacc ggctccgttc tttggtggcc ccttcgcgcc accttctact
     7261 cctcccctag tcaggaagtt cccccccgcc ccgcagctcg cgtcgtgcag gacgtgacaa
     7321 atggaagtag cacgtctcac tagtctcgtg cagatggaca gcaccgctga gcaatggaag
     7381 cgggtaggcc tttggggcag cggccaatag cagctttgct ccttcgcttt ctgggctcag
     7441 aggctgggaa ggggtgggtc cgggggcggg ctcaggggcg ggctca
//
