LOCUS       sequence-194345-        8130 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Cre-regulated lentiviral shRNA vector. Cre addition causes both 
            mCherry and shRNA to be recombined out of the construct, turning OFF
            shRNA expression. Includes puromycin selection..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8130)
  AUTHORS   Salomonis N, Schlieve CR, Pereira L, Wahlquist C, Colas A, Zambon 
            AC, Vranizan K, Spindler MJ, Pico AR, Cline MS, Clark TA, Williams 
            A, Blume JE, Samal E, Mercola M, Merrill BJ, Conklin BR
  TITLE     Alternative splicing regulates mouse embryonic stem cell 
            pluripotency and differentiation.
  JOURNAL   Proc Natl Acad Sci U S A. 2010 Jun 8;107(23):10514-9. Epub 2010 May 
            24.
  PUBMED    20498046
REFERENCE   2  (bases 1 to 8130)
  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."; date: "2010-06-8"; volume: "107(23)"; pages: 
            "10514-9. Epub 2010 May 24"
COMMENT     Sequence Label: pSicoR-Ef1a-mCh-Puro
FEATURES             Location/Qualifiers
     source          1..8130
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        137..516
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        518..716
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     671..691
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     LTR             734..914
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    961..1086
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1583..1816
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             2001..2045
                     /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    2343..2460
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     protein_bind    complement(2713..2746)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     promoter        2906..3117
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     LTR             3130..3398
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from human 
                     T-cell leukemia virus (HTLV) type 1"
     regulatory      3436..3445
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             3442..4149
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(3587..3605)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     3855..3874
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     CDS             4156..4209
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     CDS             4213..4809
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="TEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIERV
                     TELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLAA
                     QQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETSA
                     PRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     4706..4726
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     protein_bind    complement(4829..4862)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     misc_feature    4918..5506
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(4971..4991)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(5389..5400)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(5509..5525)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5510..5526)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     LTR             6035..6215
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     rep_origin      complement(6277..6865)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6357..6376)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(7036..7827,7828..7896))
                     /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     7659..7678
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7897..8001)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(8076..8095)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 ttggaggtcg ctgagtagtg cgcgagcaaa atttaagcta caacaaggca aggcttgacc
       61 gacaattgca tgaagaatct gcttagggtt aggcgttttg cgctgcttcg cgatgtacgg
      121 gccagatata cgcgttgaca ttgattattg actagttatt aatagtaatc aattacgggg
      181 tcattagttc atagcccata tatggagttc cgcgttacat aacttacggt aaatggcccg
      241 cctggctgac cgcccaacga cccccgccca ttgacgtcaa taatgacgta tgttcccata
      301 gtaacgccaa tagggacttt ccattgacgt caatgggtgg agtatttacg gtaaactgcc
      361 cacttggcag tacatcaagt gtatcatatg ccaagtacgc cccctattga cgtcaatgac
      421 ggtaaatggc ccgcctggca ttatgcccag tacatgacct tatgggactt tcctacttgg
      481 cagtacatct acgtattagt catcgctatt accatggtga tgcggttttg gcagtacatc
      541 aatgggcgtg gatagcggtt tgactcacgg ggatttccaa gtctccaccc cattgacgtc
      601 aatgggagtt tgttttggca ccaaaatcaa cgggactttc caaaatgtcg taacaactcc
      661 gccccattga cgcaaatggg cggtaggcgt gtacggtggg aggtctatat aagcagcgcg
      721 ttttgcctgt actgggtctc tctggttaga ccagatctga gcctgggagc tctctggcta
      781 actagggaac ccactgctta agcctcaata aagcttgcct tgagtgcttc aagtagtgtg
      841 tgcccgtctg ttgtgtgact ctggtaacta gagatccctc agaccctttt agtcagtgtg
      901 gaaaatctct agcagtggcg cccgaacagg gacttgaaag cgaaagggaa accagaggag
      961 ctctctcgac gcaggactcg gcttgctgaa gcgcgcacgg caagaggcga ggggcggcga
     1021 ctggtgagta cgccaaaaat tttgactagc ggaggctaga aggagagaga tgggtgcgag
     1081 agcgtcagta ttaagcgggg gagaattaga tcgcgatggg aaaaaattcg gttaaggcca
     1141 gggggaaaga aaaaatataa attaaaacat atagtatggg caagcaggga gctagaacga
     1201 ttcgcagtta atcctggcct gttagaaaca tcagaaggct gtagacaaat actgggacag
     1261 ctacaaccat cccttcagac aggatcagaa gaacttagat cattatataa tacagtagca
     1321 accctctatt gtgtgcatca aaggatagag ataaaagaca ccaaggaagc tttagacaag
     1381 atagaggaag agcaaaacaa aagtaagacc accgcacagc aagcggccgg ccgctgatct
     1441 tcagacctgg aggaggagat atgagggaca attggagaag tgaattatat aaatataaag
     1501 tagtaaaaat tgaaccatta ggagtagcac ccaccaaggc aaagagaaga gtggtgcaga
     1561 gagaaaaaag agcagtggga ataggagctt tgttccttgg gttcttggga gcagcaggaa
     1621 gcactatggg cgcagcgtca atgacgctga cggtacaggc cagacaatta ttgtctggta
     1681 tagtgcagca gcagaacaat ttgctgaggg ctattgaggc gcaacagcat ctgttgcaac
     1741 tcacagtctg gggcatcaag cagctccagg caagaatcct ggctgtggaa agatacctaa
     1801 aggatcaaca gctcctgggg atttggggtt gctctggaaa actcatttgc accactgctg
     1861 tgccttggaa tgctagttgg agtaataaat ctctggaaca gatttggaat cacacgacct
     1921 ggatggagtg ggacagagaa attaacaatt acacaagctt aatacactcc ttaattgaag
     1981 aatcgcaaaa ccagcaagaa aagaatgaac aagaattatt ggaattagat aaatgggcaa
     2041 gtttgtggaa ttggtttaac ataacaaatt ggctgtggta tataaaatta ttcataatga
     2101 tagtaggagg cttggtaggt ttaagaatag tttttgctgt actttctata gtgaatagag
     2161 ttaggcaggg atattcacca ttatcgtttc agacccacct cccaaccccg aggggacccg
     2221 acaggcccga aggaatagaa gaagaaggtg gagagagaga cagagacaga tccattcgat
     2281 tagtgaacgg atcggcactg cgtgcgccaa ttctgcagac aaatggcagt attcatccac
     2341 aattttaaaa gaaaaggggg gattgggggg tacagtgcag gggaaagaat agtagacata
     2401 atagcaacag acatacaaac taaagaatta caaaaacaaa ttacaaaaat tcaaaatttt
     2461 cgggtttatt acagggacag cagagatcca gtttggttag taccgggccc gctctagaga
     2521 tccgacgcgc catctctagg cccgcgccgg ccccctcgca cagacttgtg ggagaagctc
     2581 ggctactccc ctgccccggt taatttgcat ataatatttc ctagtaacta tagaggctta
     2641 atgtgcgata aaagacagat aatctgttct ttttaatact agctacattt tacatgatag
     2701 gcttggattt ctataacttc gtatagcata cattatacga agttatacat gtcacaaaag
     2761 gaaactcacc ctaactgtaa agtaattgtg tgttttgaga ctataaatat cccttggaga
     2821 aaagccttgt taacgcgcgg tgaccctcga gtactaggat ccattaggcg gccgcggatc
     2881 tgcgatcgct ccggtgcccg tcagtgggca gagcgcacat cgcccacagt ccccgagaag
     2941 ttggggggag gggtcggcaa ttgaaccggt gcctagagaa ggtggcgcgg ggtaaactgg
     3001 gaaagtgatg tcgtgtactg gctccgcctt tttcccgagg gtgggggaga accgtatata
     3061 agtgcagtag tcgccgtgaa cgttcttttt cgcaacgggt ttgccgccag aacacagctg
     3121 aagcttcgag gggctcgcat ctctccttca cgcgcccgcc gccctacctg aggccgccat
     3181 ccacgccggt tgagtcgcgt tctgccgcct cccgcctgtg gtgcctcctg aactgcgtcc
     3241 gccgtctagg taagtttaaa gctcaggtcg agaccgggcc tttgtccggc gctcccttgg
     3301 agcctaccta gactcagccg gctctccacg ctttgcctga ccctgcttgc tcaactctac
     3361 gtctttgttt cgttttctgt tctgcgccgt tacagatcca agctgtgacc ggcgcctacg
     3421 ctagcgctac cggtcgccac catggtgagc aagggcgagg aggataacat ggccatcatc
     3481 aaggagttca tgcgcttcaa ggtgcacatg gagggctccg tgaacggcca cgagttcgag
     3541 atcgagggcg agggcgaggg ccgcccctac gagggcaccc agaccgccaa gctgaaggtg
     3601 accaagggtg gccccctgcc cttcgcctgg gacatcctgt cccctcagtt catgtacggc
     3661 tccaaggcct acgtgaagca ccccgccgac atccccgact acttgaagct gtccttcccc
     3721 gagggcttca agtgggagcg cgtgatgaac ttcgaggacg gcggcgtggt gaccgtgacc
     3781 caggactcct ccctgcagga cggcgagttc atctacaagg tgaagctgcg cggcaccaac
     3841 ttcccctccg acggccccgt aatgcagaag aagaccatgg gctgggaggc ctcctccgag
     3901 cggatgtacc ccgaggacgg cgccctgaag ggcgagatca agcagaggct gaagctgaag
     3961 gacggcggcc actacgacgc tgaggtcaag accacctaca aggccaagaa gcccgtgcag
     4021 ctgcccggcg cctacaacgt caacatcaag ttggacatca cctcccacaa cgaggactac
     4081 accatcgtgg aacagtacga acgcgccgag ggccgccact ccaccggcgg catggacgag
     4141 ctgtacaagc cccgggaggg cagaggaagt cttctaacat gcggtgacgt ggaggagaat
     4201 cccggccctc gaaccgagta caagcccacg gtgcgcctcg ccacccgcga cgacgtcccc
     4261 agggccgtac gcaccctcgc cgccgcgttc gccgactacc ccgccacgcg ccacaccgtc
     4321 gatccggacc gccacatcga gcgggtcacc gagctgcaag aactcttcct cacgcgcgtc
     4381 gggctcgaca tcggcaaggt gtgggtcgcg gacgacggcg ccgcggtggc ggtctggacc
     4441 acgccggaga gcgtcgaagc gggggcggtg ttcgccgaga tcggcccgcg catggccgag
     4501 ttgagcggtt cccggctggc cgcgcagcaa cagatggaag gcctcctggc gccgcaccgg
     4561 cccaaggagc ccgcgtggtt cctggccacc gtcggcgtct cgcccgacca ccagggcaag
     4621 ggtctgggca gcgccgtcgt gctccccgga gtggaggcgg ccgagcgcgc cggggtgccc
     4681 gccttcctgg agacctccgc gccccgcaac ctccccttct acgagcggct cggcttcacc
     4741 gtcaccgccg acgtcgaggt gcccgaagga ccgcgcacct ggtgcatgac ccgcaagccc
     4801 ggtgcctaag aattcgtcga gggacctaat aacttcgtat agcatacatt atacgaagtt
     4861 atacatgttt aagggttccg gttccactag gtacaattcg atatcaagct tatcgataat
     4921 caacctctgg attacaaaat ttgtgaaaga ttgactggta ttcttaacta tgttgctcct
     4981 tttacgctat gtggatacgc tgctttaatg cctttgtatc atgctattgc ttcccgtatg
     5041 gctttcattt tctcctcctt gtataaatcc tggttgctgt ctctttatga ggagttgtgg
     5101 cccgttgtca ggcaacgtgg cgtggtgtgc actgtgtttg ctgacgcaac ccccactggt
     5161 tggggcattg ccaccacctg tcagctcctt tccgggactt tcgctttccc cctccctatt
     5221 gccacggcgg aactcatcgc cgcctgcctt gcccgctgct ggacaggggc tcggctgttg
     5281 ggcactgaca attccgtggt gttgtcgggg aaatcatcgt cctttccttg gctgctcgcc
     5341 tgtgttgcca cctggattct gcgcgggacg tccttctgct acgtcccttc ggccctcaat
     5401 ccagcggacc ttccttcccg cggcctgctg ccggctctgc ggcctcttcc gcgtcttcgc
     5461 cttcgccctc agacgagtcg gatctccctt tgggccgcct ccccgcatcg ataccgtcga
     5521 cctcgatcga gacctagaaa aacatggagc aatcacaagt agcaatacag cagctaccaa
     5581 tgctgattgt gcctggctag aagcacaaga ggaggaggag gtgggttttc cagtcacacc
     5641 tcaggtacct ttaagaccaa tgacttacaa ggcagctgta gatcttagcc actttttaaa
     5701 agaaaagggg ggactggaag ggctaattca ctcccaacga agacaagata tccttgatct
     5761 gtggatctac cacacacaag gctacttccc tgattggcag aactacacac cagggccagg
     5821 gatcagatat ccactgacct ttggatggtg ctacaagcta gtaccagttg agcaagagaa
     5881 ggtagaagaa gccaatgaag gagagaacac ccgcttgtta caccctgtga gcctgcatgg
     5941 gatggatgac ccggagagag aagtattaga gtggaggttt gacagccgcc tagcatttca
     6001 tcacatggcc cgagagctgc atccggactg tactgggtct ctctggttag accagatctg
     6061 agcctgggag ctctctggct aactagggaa cccactgctt aagcctcaat aaagcttgcc
     6121 ttgagtgctt caagtagtgt gtgcccgtct gttgtgtgac tctggtaact agagatccct
     6181 cagacccttt tagtcagtgt ggaaaatctc tagcagcatg tgagcaaaag gccagcaaaa
     6241 ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga
     6301 cgagcatcac aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag
     6361 ataccaggcg tttccccctg gaagctccct cgtgcgctct cctgttccga ccctgccgct
     6421 taccggatac ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc atagctcacg
     6481 ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc
     6541 ccccgttcag cccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccggt
     6601 aagacacgac ttatcgccac tggcagcagc cactggtaac aggattagca gagcgaggta
     6661 tgtaggcggt gctacagagt tcttgaagtg gtggcctaac tacggctaca ctagaagaac
     6721 agtatttggt atctgcgctc tgctgaagcc agttaccttc ggaaaaagag ttggtagctc
     6781 ttgatccggc aaacaaacca ccgctggtag cggtggtttt tttgtttgca agcagcagat
     6841 tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc
     6901 tcagtggaac gaaaactcac gttaagggat tttggtcatg agattatcaa aaaggatctt
     6961 cacctagatc cttttaaatt aaaaatgaag ttttaaatca atctaaagta tatatgagta
     7021 aacttggtct gacagttacc aatgcttaat cagtgaggca cctatctcag cgatctgtct
     7081 atttcgttca tccatagttg cctgactccc cgtcgtgtag ataactacga tacgggaggg
     7141 cttaccatct ggccccagtg ctgcaatgat accgcgagac ccacgctcac cggctccaga
     7201 tttatcagca ataaaccagc cagccggaag ggccgagcgc agaagtggtc ctgcaacttt
     7261 atccgcctcc atccagtcta ttaattgttg ccgggaagct agagtaagta gttcgccagt
     7321 taatagtttg cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt
     7381 tggtatggct tcattcagct ccggttccca acgatcaagg cgagttacat gatcccccat
     7441 gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc gttgtcagaa gtaagttggc
     7501 cgcagtgtta tcactcatgg ttatggcagc actgcataat tctcttactg tcatgccatc
     7561 cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag tcattctgag aatagtgtat
     7621 gcggcgaccg agttgctctt gcccggcgtc aatacgggat aataccgcgc cacatagcag
     7681 aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct caaggatctt
     7741 accgctgttg agatccagtt cgatgtaacc cactcgtgca cccaactgat cttcagcatc
     7801 ttttactttc accagcgttt ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa
     7861 gggaataagg gcgacacgga aatgttgaat actcatactc ttcctttttc aatattattg
     7921 aagcatttat cagggttatt gtctcatgag cggatacata tttgaatgta tttagaaaaa
     7981 taaacaaata ggggttccgc gcacatttcc ccgaaaagtg ccacctgacg tcgacggatc
     8041 gggagatctc ccgatcccct atggtgcact ctcagtacaa tctgctctga tgccgcatag
     8101 ttaagccagt atctgctccc tgcttgtgtg
//
