LOCUS       sequence-35872-e        8141 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 8141)
  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 8141)
  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..8141
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(56..75)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        247..626
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        628..826
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     781..801
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     LTR             844..1024
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    1071..1196
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1695..1928
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             2113..2157
                     /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    2455..2572
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     protein_bind    complement(2825..2858)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     promoter        3018..3229
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     LTR             3242..3510
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from human 
                     T-cell leukemia virus (HTLV) type 1"
     regulatory      3548..3557
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             3554..4261
                     /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(3699..3717)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     3967..3986
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     CDS             4268..4321
                     /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             4325..4921
                     /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     4818..4838
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     protein_bind    complement(4941..4974)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     misc_feature    5030..5618
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(5083..5103)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(5501..5512)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(5621..5637)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5622..5638)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     LTR             6147..6327
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     rep_origin      complement(6389..6977)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6469..6488)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(7148..7939,7940..8008))
                     /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     7771..7790
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(8009..8113)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gcttaagcgg tcgacggatc gggagatctc ccgatcccct atggtgcact ctcagtacaa
       61 tctgctctga tgccgcatag ttaagccagt atctgctccc tgcttgtgtg ttggaggtcg
      121 ctgagtagtg cgcgagcaaa atttaagcta caacaaggca aggcttgacc gacaattgca
      181 tgaagaatct gcttagggtt aggcgttttg cgctgcttcg cgatgtacgg gccagatata
      241 cgcgttgaca ttgattattg actagttatt aatagtaatc aattacgggg tcattagttc
      301 atagcccata tatggagttc cgcgttacat aacttacggt aaatggcccg cctggctgac
      361 cgcccaacga cccccgccca ttgacgtcaa taatgacgta tgttcccata gtaacgccaa
      421 tagggacttt ccattgacgt caatgggtgg agtatttacg gtaaactgcc cacttggcag
      481 tacatcaagt gtatcatatg ccaagtacgc cccctattga cgtcaatgac ggtaaatggc
      541 ccgcctggca ttatgcccag tacatgacct tatgggactt tcctacttgg cagtacatct
      601 acgtattagt catcgctatt accatggtga tgcggttttg gcagtacatc aatgggcgtg
      661 gatagcggtt tgactcacgg ggatttccaa gtctccaccc cattgacgtc aatgggagtt
      721 tgttttggca ccaaaatcaa cgggactttc caaaatgtcg taacaactcc gccccattga
      781 cgcaaatggg cggtaggcgt gtacggtggg aggtctatat aagcagcgcg ttttgcctgt
      841 actgggtctc tctggttaga ccagatctga gcctgggagc tctctggcta actagggaac
      901 ccactgctta agcctcaata aagcttgcct tgagtgcttc aagtagtgtg tgcccgtctg
      961 ttgtgtgact ctggtaacta gagatccctc agaccctttt agtcagtgtg gaaaatctct
     1021 agcagtggcg cccgaacagg gacttgaaag cgaaagggaa accagaggag ctctctcgac
     1081 gcaggactcg gcttgctgaa gcgcgcacgg caagaggcga ggggcggcga ctggtgagta
     1141 cgccaaaaat tttgactagc ggaggctaga aggagagaga tgggtgcgag agcgtcagta
     1201 ttaagcgggg gagaattaga tcgcgatggg aaaaaattcg gttaaggcca gggggaaaga
     1261 aaaaatataa attaaaacat atagtatggg caagcaggga gctagaacga ttcgcagtta
     1321 atcctggcct gttagaaaca tcagaaggct gtagacaaat actgggacag ctacaaccat
     1381 cccttcagac aggatcagaa gaacttagat cattatataa tacagtagca accctctatt
     1441 gtgtgcatca aaggatagag ataaaagaca ccaaggaagc tttagacaag atagaggaag
     1501 agcaaaacaa aagtaagacc accgcacagc aagcggccgg ccgcgctgat cttcagacct
     1561 ggaggaggag atatgaggga caattggaga agtgaattat ataaatataa agtagtaaaa
     1621 attgaaccat taggagtagc acccaccaag gcaaagagaa gagtggtgca gagagaaaaa
     1681 agagcagtgg gaataggagc tttgttcctt gggttcttgg gagcagcagg aagcactatg
     1741 ggcgcagcgt caatgacgct gacggtacag gccagacaat tattgtctgg tatagtgcag
     1801 cagcagaaca atttgctgag ggctattgag gcgcaacagc atctgttgca actcacagtc
     1861 tggggcatca agcagctcca ggcaagaatc ctggctgtgg aaagatacct aaaggatcaa
     1921 cagctcctgg ggatttgggg ttgctctgga aaactcattt gcaccactgc tgtgccttgg
     1981 aatgctagtt ggagtaataa atctctggaa cagatttgga atcacacgac ctggatggag
     2041 tgggacagag aaattaacaa ttacacaagc ttaatacact ccttaattga agaatcgcaa
     2101 aaccagcaag aaaagaatga acaagaatta ttggaattag ataaatgggc aagtttgtgg
     2161 aattggttta acataacaaa ttggctgtgg tatataaaat tattcataat gatagtagga
     2221 ggcttggtag gtttaagaat agtttttgct gtactttcta tagtgaatag agttaggcag
     2281 ggatattcac cattatcgtt tcagacccac ctcccaaccc cgaggggacc cgacaggccc
     2341 gaaggaatag aagaagaagg tggagagaga gacagagaca gatccattcg attagtgaac
     2401 ggatcggcac tgcgtgcgcc aattctgcag acaaatggca gtattcatcc acaattttaa
     2461 aagaaaaggg gggattgggg ggtacagtgc aggggaaaga atagtagaca taatagcaac
     2521 agacatacaa actaaagaat tacaaaaaca aattacaaaa attcaaaatt ttcgggttta
     2581 ttacagggac agcagagatc cagtttggtt agtaccgggc ccgctctaga gatccgacgc
     2641 gccatctcta ggcccgcgcc ggccccctcg cacagacttg tgggagaagc tcggctactc
     2701 ccctgccccg gttaatttgc atataatatt tcctagtaac tatagaggct taatgtgcga
     2761 taaaagacag ataatctgtt ctttttaata ctagctacat tttacatgat aggcttggat
     2821 ttctataact tcgtatagca tacattatac gaagttatac atgtcacaaa aggaaactca
     2881 ccctaactgt aaagtaattg tgtgttttga gactataaat atcccttgga gaaaagcctt
     2941 gttaacgcgc ggtgaccctc gagtactagg atccattagg cggccgcgga tctgcgatcg
     3001 ctccggtgcc cgtcagtggg cagagcgcac atcgcccaca gtccccgaga agttgggggg
     3061 aggggtcggc aattgaaccg gtgcctagag aaggtggcgc ggggtaaact gggaaagtga
     3121 tgtcgtgtac tggctccgcc tttttcccga gggtggggga gaaccgtata taagtgcagt
     3181 agtcgccgtg aacgttcttt ttcgcaacgg gtttgccgcc agaacacagc tgaagcttcg
     3241 aggggctcgc atctctcctt cacgcgcccg ccgccctacc tgaggccgcc atccacgccg
     3301 gttgagtcgc gttctgccgc ctcccgcctg tggtgcctcc tgaactgcgt ccgccgtcta
     3361 ggtaagttta aagctcaggt cgagaccggg cctttgtccg gcgctccctt ggagcctacc
     3421 tagactcagc cggctctcca cgctttgcct gaccctgctt gctcaactct acgtctttgt
     3481 ttcgttttct gttctgcgcc gttacagatc caagctgtga ccggcgccta cgctagcgct
     3541 accggtcgcc accatggtga gcaagggcga ggaggataac atggccatca tcaaggagtt
     3601 catgcgcttc aaggtgcaca tggagggctc cgtgaacggc cacgagttcg agatcgaggg
     3661 cgagggcgag ggccgcccct acgagggcac ccagaccgcc aagctgaagg tgaccaaggg
     3721 tggccccctg cccttcgcct gggacatcct gtcccctcag ttcatgtacg gctccaaggc
     3781 ctacgtgaag caccccgccg acatccccga ctacttgaag ctgtccttcc ccgagggctt
     3841 caagtgggag cgcgtgatga acttcgagga cggcggcgtg gtgaccgtga cccaggactc
     3901 ctccctgcag gacggcgagt tcatctacaa ggtgaagctg cgcggcacca acttcccctc
     3961 cgacggcccc gtaatgcaga agaagaccat gggctgggag gcctcctccg agcggatgta
     4021 ccccgaggac ggcgccctga agggcgagat caagcagagg ctgaagctga aggacggcgg
     4081 ccactacgac gctgaggtca agaccaccta caaggccaag aagcccgtgc agctgcccgg
     4141 cgcctacaac gtcaacatca agttggacat cacctcccac aacgaggact acaccatcgt
     4201 ggaacagtac gaacgcgccg agggccgcca ctccaccggc ggcatggacg agctgtacaa
     4261 gccccgggag ggcagaggaa gtcttctaac atgcggtgac gtggaggaga atcccggccc
     4321 tcgaaccgag tacaagccca cggtgcgcct cgccacccgc gacgacgtcc ccagggccgt
     4381 acgcaccctc gccgccgcgt tcgccgacta ccccgccacg cgccacaccg tcgatccgga
     4441 ccgccacatc gagcgggtca ccgagctgca agaactcttc ctcacgcgcg tcgggctcga
     4501 catcggcaag gtgtgggtcg cggacgacgg cgccgcggtg gcggtctgga ccacgccgga
     4561 gagcgtcgaa gcgggggcgg tgttcgccga gatcggcccg cgcatggccg agttgagcgg
     4621 ttcccggctg gccgcgcagc aacagatgga aggcctcctg gcgccgcacc ggcccaagga
     4681 gcccgcgtgg ttcctggcca ccgtcggcgt ctcgcccgac caccagggca agggtctggg
     4741 cagcgccgtc gtgctccccg gagtggaggc ggccgagcgc gccggggtgc ccgccttcct
     4801 ggagacctcc gcgccccgca acctcccctt ctacgagcgg ctcggcttca ccgtcaccgc
     4861 cgacgtcgag gtgcccgaag gaccgcgcac ctggtgcatg acccgcaagc ccggtgccta
     4921 agaattcgtc gagggaccta ataacttcgt atagcataca ttatacgaag ttatacatgt
     4981 ttaagggttc cggttccact aggtacaatt cgatatcaag cttatcgata atcaacctct
     5041 ggattacaaa atttgtgaaa gattgactgg tattcttaac tatgttgctc cttttacgct
     5101 atgtggatac gctgctttaa tgcctttgta tcatgctatt gcttcccgta tggctttcat
     5161 tttctcctcc ttgtataaat cctggttgct gtctctttat gaggagttgt ggcccgttgt
     5221 caggcaacgt ggcgtggtgt gcactgtgtt tgctgacgca acccccactg gttggggcat
     5281 tgccaccacc tgtcagctcc tttccgggac tttcgctttc cccctcccta ttgccacggc
     5341 ggaactcatc gccgcctgcc ttgcccgctg ctggacaggg gctcggctgt tgggcactga
     5401 caattccgtg gtgttgtcgg ggaaatcatc gtcctttcct tggctgctcg cctgtgttgc
     5461 cacctggatt ctgcgcggga cgtccttctg ctacgtccct tcggccctca atccagcgga
     5521 ccttccttcc cgcggcctgc tgccggctct gcggcctctt ccgcgtcttc gccttcgccc
     5581 tcagacgagt cggatctccc tttgggccgc ctccccgcat cgataccgtc gacctcgatc
     5641 gagacctaga aaaacatgga gcaatcacaa gtagcaatac agcagctacc aatgctgatt
     5701 gtgcctggct agaagcacaa gaggaggagg aggtgggttt tccagtcaca cctcaggtac
     5761 ctttaagacc aatgacttac aaggcagctg tagatcttag ccacttttta aaagaaaagg
     5821 ggggactgga agggctaatt cactcccaac gaagacaaga tatccttgat ctgtggatct
     5881 accacacaca aggctacttc cctgattggc agaactacac accagggcca gggatcagat
     5941 atccactgac ctttggatgg tgctacaagc tagtaccagt tgagcaagag aaggtagaag
     6001 aagccaatga aggagagaac acccgcttgt tacaccctgt gagcctgcat gggatggatg
     6061 acccggagag agaagtatta gagtggaggt ttgacagccg cctagcattt catcacatgg
     6121 cccgagagct gcatccggac tgtactgggt ctctctggtt agaccagatc tgagcctggg
     6181 agctctctgg ctaactaggg aacccactgc ttaagcctca ataaagcttg ccttgagtgc
     6241 ttcaagtagt gtgtgcccgt ctgttgtgtg actctggtaa ctagagatcc ctcagaccct
     6301 tttagtcagt gtggaaaatc tctagcagca tgtgagcaaa aggccagcaa aaggccagga
     6361 accgtaaaaa ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc
     6421 acaaaaatcg acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg
     6481 cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat
     6541 acctgtccgc ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt
     6601 atctcagttc ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc
     6661 agcccgaccg ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg
     6721 acttatcgcc actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg
     6781 gtgctacaga gttcttgaag tggtggccta actacggcta cactagaaga acagtatttg
     6841 gtatctgcgc tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg
     6901 gcaaacaaac caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca
     6961 gaaaaaaagg atctcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga
     7021 acgaaaactc acgttaaggg attttggtca tgagattatc aaaaaggatc ttcacctaga
     7081 tccttttaaa ttaaaaatga agttttaaat caatctaaag tatatatgag taaacttggt
     7141 ctgacagtta ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt
     7201 catccatagt tgcctgactc cccgtcgtgt agataactac gatacgggag ggcttaccat
     7261 ctggccccag tgctgcaatg ataccgcgag acccacgctc accggctcca gatttatcag
     7321 caataaacca gccagccgga agggccgagc gcagaagtgg tcctgcaact ttatccgcct
     7381 ccatccagtc tattaattgt tgccgggaag ctagagtaag tagttcgcca gttaatagtt
     7441 tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg
     7501 cttcattcag ctccggttcc caacgatcaa ggcgagttac atgatccccc atgttgtgca
     7561 aaaaagcggt tagctccttc ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt
     7621 tatcactcat ggttatggca gcactgcata attctcttac tgtcatgcca tccgtaagat
     7681 gcttttctgt gactggtgag tactcaacca agtcattctg agaatagtgt atgcggcgac
     7741 cgagttgctc ttgcccggcg tcaatacggg ataataccgc gccacatagc agaactttaa
     7801 aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt
     7861 tgagatccag ttcgatgtaa cccactcgtg cacccaactg atcttcagca tcttttactt
     7921 tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa
     7981 gggcgacacg gaaatgttga atactcatac tcttcctttt tcaatattat tgaagcattt
     8041 atcagggtta ttgtctcatg agcggataca tatttgaatg tatttagaaa aataaacaaa
     8101 taggggttcc gcgcacattt ccccgaaaag tgccacctga c
//
