LOCUS       Exported                8181 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    pSicoR-Ef1a-mCh-Puro-Luci
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8181)
  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 8181)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..8181
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     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    2825..2858
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (GCATACAT)."
     promoter        3058..3269
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     LTR             3282..3550
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from human 
                     T-cell leukemia virus (HTLV) type 1"
     regulatory      3588..3597
                     /regulatory_class="other"
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     CDS             3594..4301
                     /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(3739..3757)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     4007..4026
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     CDS             4308..4361
                     /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             4365..4961
                     /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     4858..4878
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     protein_bind    4981..5014
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (GCATACAT)."
     misc_feature    5070..5658
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(5123..5143)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(5541..5552)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(5661..5677)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5662..5678)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     LTR             6187..6367
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     rep_origin      complement(6429..7017)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6509..6528)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(7188..8048)
                     /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     7811..7830
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(8049..8153)
                     /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 gtttcttacg ctgagtactt cgattcaaga gatcgaagta ctcagcgtaa gttttttctc
     3001 gagtactagg atccattagg cggccgcgga tctgcgatcg ctccggtgcc cgtcagtggg
     3061 cagagcgcac atcgcccaca gtccccgaga agttgggggg aggggtcggc aattgaaccg
     3121 gtgcctagag aaggtggcgc ggggtaaact gggaaagtga tgtcgtgtac tggctccgcc
     3181 tttttcccga gggtggggga gaaccgtata taagtgcagt agtcgccgtg aacgttcttt
     3241 ttcgcaacgg gtttgccgcc agaacacagc tgaagcttcg aggggctcgc atctctcctt
     3301 cacgcgcccg ccgccctacc tgaggccgcc atccacgccg gttgagtcgc gttctgccgc
     3361 ctcccgcctg tggtgcctcc tgaactgcgt ccgccgtcta ggtaagttta aagctcaggt
     3421 cgagaccggg cctttgtccg gcgctccctt ggagcctacc tagactcagc cggctctcca
     3481 cgctttgcct gaccctgctt gctcaactct acgtctttgt ttcgttttct gttctgcgcc
     3541 gttacagatc caagctgtga ccggcgccta cgctagcgct accggtcgcc accatggtga
     3601 gcaagggcga ggaggataac atggccatca tcaaggagtt catgcgcttc aaggtgcaca
     3661 tggagggctc cgtgaacggc cacgagttcg agatcgaggg cgagggcgag ggccgcccct
     3721 acgagggcac ccagaccgcc aagctgaagg tgaccaaggg tggccccctg cccttcgcct
     3781 gggacatcct gtcccctcag ttcatgtacg gctccaaggc ctacgtgaag caccccgccg
     3841 acatccccga ctacttgaag ctgtccttcc ccgagggctt caagtgggag cgcgtgatga
     3901 acttcgagga cggcggcgtg gtgaccgtga cccaggactc ctccctgcag gacggcgagt
     3961 tcatctacaa ggtgaagctg cgcggcacca acttcccctc cgacggcccc gtaatgcaga
     4021 agaagaccat gggctgggag gcctcctccg agcggatgta ccccgaggac ggcgccctga
     4081 agggcgagat caagcagagg ctgaagctga aggacggcgg ccactacgac gctgaggtca
     4141 agaccaccta caaggccaag aagcccgtgc agctgcccgg cgcctacaac gtcaacatca
     4201 agttggacat cacctcccac aacgaggact acaccatcgt ggaacagtac gaacgcgccg
     4261 agggccgcca ctccaccggc ggcatggacg agctgtacaa gccccgggag ggcagaggaa
     4321 gtcttctaac atgcggtgac gtggaggaga atcccggccc tcgaaccgag tacaagccca
     4381 cggtgcgcct cgccacccgc gacgacgtcc ccagggccgt acgcaccctc gccgccgcgt
     4441 tcgccgacta ccccgccacg cgccacaccg tcgatccgga ccgccacatc gagcgggtca
     4501 ccgagctgca agaactcttc ctcacgcgcg tcgggctcga catcggcaag gtgtgggtcg
     4561 cggacgacgg cgccgcggtg gcggtctgga ccacgccgga gagcgtcgaa gcgggggcgg
     4621 tgttcgccga gatcggcccg cgcatggccg agttgagcgg ttcccggctg gccgcgcagc
     4681 aacagatgga aggcctcctg gcgccgcacc ggcccaagga gcccgcgtgg ttcctggcca
     4741 ccgtcggcgt ctcgcccgac caccagggca agggtctggg cagcgccgtc gtgctccccg
     4801 gagtggaggc ggccgagcgc gccggggtgc ccgccttcct ggagacctcc gcgccccgca
     4861 acctcccctt ctacgagcgg ctcggcttca ccgtcaccgc cgacgtcgag gtgcccgaag
     4921 gaccgcgcac ctggtgcatg acccgcaagc ccggtgccta agaattcgtc gagggaccta
     4981 ataacttcgt atagcataca ttatacgaag ttatacatgt ttaagggttc cggttccact
     5041 aggtacaatt cgatatcaag cttatcgata atcaacctct ggattacaaa atttgtgaaa
     5101 gattgactgg tattcttaac tatgttgctc cttttacgct atgtggatac gctgctttaa
     5161 tgcctttgta tcatgctatt gcttcccgta tggctttcat tttctcctcc ttgtataaat
     5221 cctggttgct gtctctttat gaggagttgt ggcccgttgt caggcaacgt ggcgtggtgt
     5281 gcactgtgtt tgctgacgca acccccactg gttggggcat tgccaccacc tgtcagctcc
     5341 tttccgggac tttcgctttc cccctcccta ttgccacggc ggaactcatc gccgcctgcc
     5401 ttgcccgctg ctggacaggg gctcggctgt tgggcactga caattccgtg gtgttgtcgg
     5461 ggaaatcatc gtcctttcct tggctgctcg cctgtgttgc cacctggatt ctgcgcggga
     5521 cgtccttctg ctacgtccct tcggccctca atccagcgga ccttccttcc cgcggcctgc
     5581 tgccggctct gcggcctctt ccgcgtcttc gccttcgccc tcagacgagt cggatctccc
     5641 tttgggccgc ctccccgcat cgataccgtc gacctcgatc gagacctaga aaaacatgga
     5701 gcaatcacaa gtagcaatac agcagctacc aatgctgatt gtgcctggct agaagcacaa
     5761 gaggaggagg aggtgggttt tccagtcaca cctcaggtac ctttaagacc aatgacttac
     5821 aaggcagctg tagatcttag ccacttttta aaagaaaagg ggggactgga agggctaatt
     5881 cactcccaac gaagacaaga tatccttgat ctgtggatct accacacaca aggctacttc
     5941 cctgattggc agaactacac accagggcca gggatcagat atccactgac ctttggatgg
     6001 tgctacaagc tagtaccagt tgagcaagag aaggtagaag aagccaatga aggagagaac
     6061 acccgcttgt tacaccctgt gagcctgcat gggatggatg acccggagag agaagtatta
     6121 gagtggaggt ttgacagccg cctagcattt catcacatgg cccgagagct gcatccggac
     6181 tgtactgggt ctctctggtt agaccagatc tgagcctggg agctctctgg ctaactaggg
     6241 aacccactgc ttaagcctca ataaagcttg ccttgagtgc ttcaagtagt gtgtgcccgt
     6301 ctgttgtgtg actctggtaa ctagagatcc ctcagaccct tttagtcagt gtggaaaatc
     6361 tctagcagca tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa ggccgcgttg
     6421 ctggcgtttt tccataggct ccgcccccct gacgagcatc acaaaaatcg acgctcaagt
     6481 cagaggtggc gaaacccgac aggactataa agataccagg cgtttccccc tggaagctcc
     6541 ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat acctgtccgc ctttctccct
     6601 tcgggaagcg tggcgctttc tcatagctca cgctgtaggt atctcagttc ggtgtaggtc
     6661 gttcgctcca agctgggctg tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta
     6721 tccggtaact atcgtcttga gtccaacccg gtaagacacg acttatcgcc actggcagca
     6781 gccactggta acaggattag cagagcgagg tatgtaggcg gtgctacaga gttcttgaag
     6841 tggtggccta actacggcta cactagaaga acagtatttg gtatctgcgc tctgctgaag
     6901 ccagttacct tcggaaaaag agttggtagc tcttgatccg gcaaacaaac caccgctggt
     6961 agcggtggtt tttttgtttg caagcagcag attacgcgca gaaaaaaagg atctcaagaa
     7021 gatcctttga tcttttctac ggggtctgac gctcagtgga acgaaaactc acgttaaggg
     7081 attttggtca tgagattatc aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga
     7141 agttttaaat caatctaaag tatatatgag taaacttggt ctgacagtta ccaatgctta
     7201 atcagtgagg cacctatctc agcgatctgt ctatttcgtt catccatagt tgcctgactc
     7261 cccgtcgtgt agataactac gatacgggag ggcttaccat ctggccccag tgctgcaatg
     7321 ataccgcgag acccacgctc accggctcca gatttatcag caataaacca gccagccgga
     7381 agggccgagc gcagaagtgg tcctgcaact ttatccgcct ccatccagtc tattaattgt
     7441 tgccgggaag ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt
     7501 gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag ctccggttcc
     7561 caacgatcaa ggcgagttac atgatccccc atgttgtgca aaaaagcggt tagctccttc
     7621 ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt tatcactcat ggttatggca
     7681 gcactgcata attctcttac tgtcatgcca tccgtaagat gcttttctgt gactggtgag
     7741 tactcaacca agtcattctg agaatagtgt atgcggcgac cgagttgctc ttgcccggcg
     7801 tcaatacggg ataataccgc gccacatagc agaactttaa aagtgctcat cattggaaaa
     7861 cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa
     7921 cccactcgtg cacccaactg atcttcagca tcttttactt tcaccagcgt ttctgggtga
     7981 gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg gaaatgttga
     8041 atactcatac tcttcctttt tcaatattat tgaagcattt atcagggtta ttgtctcatg
     8101 agcggataca tatttgaatg tatttagaaa aataaacaaa taggggttcc gcgcacattt
     8161 ccccgaaaag tgccacctga c
//
