LOCUS       sequence-463643-        8424 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  For strong lentivirus-mediated expression of GFP.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8424)
  AUTHORS   Shin D, Kim CN, Ross J, Hennick KM, Wu SR, Paranjape N, Leonard R, 
            Wang JC, Keefe MG, Pavlovic BJ, Donohue KC, Moreau C, Wigdor EM, 
            Larson HH, Allen DE, Cadwell CR, Bhaduri A, Popova G, Bearden CE, 
            Pollen AA, Jacquemont S, Sanders SJ, Haussler D, Wiita AP, Frost NA,
            Sohal VS, Nowakowski TJ
  TITLE     Thalamocortical organoids enable in vitro modeling of 22q11.2 
            microdeletion associated with neuropsychiatric disorders.
  JOURNAL   Cell Stem Cell. 2024 Mar 7;31(3):421-432.e8. doi: 
            10.1016/j.stem.2024.01.010. Epub 2024 Feb 20.
  PUBMED    38382530
REFERENCE   2  (bases 1 to 8424)
  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"; doi: 
            "10.1016/j.stem.2024.01.010"; journalName: "Cell Stem Cell"; date: 
            "2024-03-7- 7"; volume: "31"; issue: "3"; pages: "421-432.e8"
COMMENT     Sequence Label: pSicoFlipped-CAG-GFP-WPRE-bGHpA
FEATURES             Location/Qualifiers
     source          1..8424
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             337..381
                     /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    679..796
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     polyA_signal    854..1078
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     misc_feature    1203..1791
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     CDS             1309..1320
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     1718..1738
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(join(1826..2539,2540..2542,2543..2545))
                     /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             complement(join(1829..2539,2540..2542,2543..2545))
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1871..1892)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     2220..2239
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2479..2500
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     regulatory      2542..2551
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     intron          2583..3599
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     complement(2637..2659)
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     promoter        complement(3600..3875)
                     /label=chicken beta-actin promoter
     enhancer        3877..4256
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     LTR             4665..4845
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     rep_origin      complement(4907..5495)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4987..5006)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5666..6457,6458..6526))
                     /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     6289..6308
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6527..6631)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(6706..6725)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        6897..7276
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        7278..7476
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     7431..7451
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     LTR             7494..7674
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    7721..7846
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    join(8343..8424,1..152)
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
ORIGIN
        1 caattattgt ctggtatagt gcagcagcag aacaatttgc tgagggctat tgaggcgcaa
       61 cagcatctgt tgcaactcac agtctggggc atcaagcagc tccaggcaag aatcctggct
      121 gtggaaagat acctaaagga tcaacagctc ctggggattt ggggttgctc tggaaaactc
      181 atttgcacca ctgctgtgcc ttggaatgct agttggagta ataaatctct ggaacagatt
      241 tggaatcaca cgacctggat ggagtgggac agagaaatta acaattacac aagcttaata
      301 cactccttaa ttgaagaatc gcaaaaccag caagaaaaga atgaacaaga attattggaa
      361 ttagataaat gggcaagttt gtggaattgg tttaacataa caaattggct gtggtatata
      421 aaattattca taatgatagt aggaggcttg gtaggtttaa gaatagtttt tgctgtactt
      481 tctatagtga atagagttag gcagggatat tcaccattat cgtttcagac ccacctccca
      541 accccgaggg gacccgacag gcccgaagga atagaagaag aaggtggaga gagagacaga
      601 gacagatcca ttcgattagt gaacggatcg gcactgcgtg cgccaattct gcagacaaat
      661 ggcagtattc atccacaatt ttaaaagaaa aggggggatt ggggggtaca gtgcagggga
      721 aagaatagta gacataatag caacagacat acaaactaaa gaattacaaa aacaaattac
      781 aaaaattcaa aattttcggg tttattacag ggacagcaga gatccagttt ggttagtacc
      841 gggcccgctc tagccataga gcccaccgca tccccagcat gcctgctatt gtcttcccaa
      901 tcctccccct tgctgtcctg ccccacccca ccccccagaa tagaatgaca cctactcaga
      961 caatgcgatg caatttcctc attttattag gaaaggacag tgggagtggc accttccagg
     1021 gtcaaggaag gcacggggga ggggcaaaca acagatggct ggcaactaga aggcacagct
     1081 cgagccggcc ttagattacc tagcataacc gccctaagtt acctagcata aacgccccaa
     1141 gcgatccggc ctccttcctc ttgaattcgc ttgtgcccca gtttgctagg gaggtccagg
     1201 acgcggggag gcggcccaaa gggagatccg actcgtctga gggcgaaggc gaagacgcgg
     1261 aagaggccgc agagccggca gcaggccgcg ggaaggaagg tccgctggat tgagggccga
     1321 agggacgtag cagaaggacg tcccgcgcag aatccaggtg gcaacacagg cgagcagcca
     1381 aggaaaggac gatgatttcc ccgacaacac cacggaattg tcagtgccca acagccgagc
     1441 ccctgtccag cagcgggcaa ggcaggcggc gatgagttcc gccgtggcaa tagggagggg
     1501 gaaagcgaaa gtcccggaaa ggagctgaca ggtggtggca atgccccaac cagtgggggt
     1561 tgcgtcagca aacacagtgc acaccacgcc acgttgcctg acaacgggcc acaactcctc
     1621 ataaagagac agcaaccagg atttatacaa ggaggagaaa atgaaagcca tacgggaagc
     1681 aatagcatga tacaaaggca ttaaagcagc gtatccacat agcgtaaaag gagcaacata
     1741 gttaagaata ccagtcaatc tttcacaaat tttgtaatcc agaggttgat tggatcgtac
     1801 gtcggcataa gctttgtagc ggccgctact tgtacagctc gtccatgccg agagtgatcc
     1861 cggcggcggt cacgaactcc agcaggacca tgtgatcgcg cttctcgttg gggtctttgc
     1921 tcagggcgga ctgggtgctc aggtagtggt tgtcgggcag cagcacgggg ccgtcgccga
     1981 tgggggtgtt ctgctggtag tggtcggcga gctgcacgct gccgtcctcg atgttgtggc
     2041 ggatcttgaa gttcaccttg atgccgttct tctgcttgtc ggccatgata tagacgttgt
     2101 ggctgttgta gttgtactcc agcttgtgcc ccaggatgtt gccgtcctcc ttgaagtcga
     2161 tgcccttcag ctcgatgcgg ttcaccaggg tgtcgccctc gaacttcacc tcggcgcggg
     2221 tcttgtagtt gccgtcgtcc ttgaagaaga tggtgcgctc ctggacgtag ccttcgggca
     2281 tggcggactt gaagaagtcg tgctgcttca tgtggtcggg gtagcggctg aagcactgca
     2341 cgccgtaggt cagggtggtc acgagggtgg gccagggcac gggcagcttg ccggtggtgc
     2401 agatgaactt cagggtcagc ttgccgtagg tggcatcgcc ctcgccctcg ccggacacgc
     2461 tgaacttgtg gccgtttacg tcgccgtcca gctcgaccag gatgggcacc accccggtga
     2521 acagctcctc gcccttgctc accatggtgg cgaccggtag cgctagttac gactagcagg
     2581 agctgtagga aaaagaagaa ggcatgaaca tggttagcag aggctctaga gccgccggtc
     2641 acacgccaga agccgaaccc cgccctgccc cgtccccccc gaaggcagcc gtccccccgc
     2701 ggacagcccc gaggctggag agggagaagg ggacggcggc gcggcgacgc acgaaggccc
     2761 tccccgccca tttccttcct gccggcgccg caccgcttcg ccccgcgccc gctagagggg
     2821 gtgcggcggc gcctcccaga tttcggctcc gcacagattt gggacaaagg aagtccctgc
     2881 gccctctcgc acgattacca taaaaggcaa tggctgcggc tcgccgcgcc tcgacagccg
     2941 ccggcgctcc gggggccgcc gcgcccctcc cccgagccct ccccggcccg aggcggcccc
     3001 gccccgcccg gcacccccac ctgccgccac cccccgcccg gcacggcgag ccccgcgcca
     3061 cgccccgtac ggagccccgc acccgaagcc gggccgtgct cagcaactcg gggagggggg
     3121 tgcagggggg gttgcagccc gaccgacgcg cccacacccc ctgctcaccc ccccacgcac
     3181 acaccccgca cgcagccttt gttcccctcg cagccccccc cgcaccgcgg ggcaccgccc
     3241 ccggccgcgc tcccctcgcg cacactgcgg agcgcacaaa gccccgcgcc gcgcccgcag
     3301 cgctcacagc cgccgggcag cgcggagccg cacgcggcgc tccccacgca cacacacacg
     3361 cacgcacccc ccgagccgct ccccccgcac aaagggccct cccggagccc ctcaaggctt
     3421 tcacgcagcc acagaaaaga aacaagccgt cattaaacca agcgctaatt acagcccgga
     3481 ggagaagggc cgtcccgccc gctcacctgt gggagtaacg cggtcagtca gagccggggc
     3541 gggcggcgcg aggcggcggc ggagcggggc acggggcgaa ggcagcgcgc agcgactccc
     3601 gcccgccgcg cgcttcgctt tttatagggc cgccgccgcc gccgcctcgc cataaaagga
     3661 aactttcgga gcgcgccgct ctgattggct gccgccgcac ctctccgcct cgccccgccc
     3721 cgcccctcgc cccgccccgc cccgcctggc gcgccccccc cccccccccg cccccatcgc
     3781 tgcacaaaat aattaaaaaa taaataaata caaaattggg ggtggggagg ggggggagat
     3841 ggggagagtg aagcagaacg tggggctcac ctcgaccatg gtaatagcga tgactaatac
     3901 gtagatgtac tgccaagtag gaaagtccca taaggtcatg tactgggcat aatgccaggc
     3961 gggccattta ccgtcattga cgtcaatagg gggcgtactt ggcatatgat acacttgatg
     4021 tactgccaag tgggcagttt accgtaaata ctccacccat tgacgtcaat ggaaagtccc
     4081 tattggcgtt actatgggaa catacgtcat tattgacgtc aatgggcggg ggtcgttggg
     4141 cggtcagcca ggcgggccat ttaccgtaag ttatgtaacg cggaactcca tatatgggct
     4201 atgaactaat gaccccgtaa ttgattacta ttaataacta gtcaataatc aatgtcgagc
     4261 tgatctacga tgtagtacct ttaagaccaa tgacttacaa ggcagctgta gatcttagcc
     4321 actttttaaa agaaaagggg ggactggaag ggctaattca ctcccaacga agacaagata
     4381 tccttgatct gtggatctac cacacacaag gctacttccc tgattggcag aactacacac
     4441 cagggccagg gatcagatat ccactgacct ttggatggtg ctacaagcta gtaccagttg
     4501 agcaagagaa ggtagaagaa gccaatgaag gagagaacac ccgcttgtta caccctgtga
     4561 gcctgcatgg gatggatgac ccggagagag aagtattaga gtggaggttt gacagccgcc
     4621 tagcatttca tcacatggcc cgagagctgc atccggactg tactgggtct ctctggttag
     4681 accagatctg agcctgggag ctctctggct aactagggaa cccactgctt aagcctcaat
     4741 aaagcttgcc ttgagtgctt caagtagtgt gtgcccgtct gttgtgtgac tctggtaact
     4801 agagatccct cagacccttt tagtcagtgt ggaaaatctc tagcagcatg tgagcaaaag
     4861 gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc
     4921 gcccccctga cgagcatcac aaaaatcgac gctcaagtca gaggtggcga aacccgacag
     4981 gactataaag ataccaggcg tttccccctg gaagctccct cgtgcgctct cctgttccga
     5041 ccctgccgct taccggatac ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc
     5101 atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg
     5161 tgcacgaacc ccccgttcag cccgaccgct gcgccttatc cggtaactat cgtcttgagt
     5221 ccaacccggt aagacacgac ttatcgccac tggcagcagc cactggtaac aggattagca
     5281 gagcgaggta tgtaggcggt gctacagagt tcttgaagtg gtggcctaac tacggctaca
     5341 ctagaagaac agtatttggt atctgcgctc tgctgaagcc agttaccttc ggaaaaagag
     5401 ttggtagctc ttgatccggc aaacaaacca ccgctggtag cggtggtttt tttgtttgca
     5461 agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg
     5521 ggtctgacgc tcagtggaac gaaaactcac gttaagggat tttggtcatg agattatcaa
     5581 aaaggatctt cacctagatc cttttaaatt aaaaatgaag ttttaaatca atctaaagta
     5641 tatatgagta aacttggtct gacagttacc aatgcttaat cagtgaggca cctatctcag
     5701 cgatctgtct atttcgttca tccatagttg cctgactccc cgtcgtgtag ataactacga
     5761 tacgggaggg cttaccatct ggccccagtg ctgcaatgat accgcgagac ccacgctcac
     5821 cggctccaga tttatcagca ataaaccagc cagccggaag ggccgagcgc agaagtggtc
     5881 ctgcaacttt atccgcctcc atccagtcta ttaattgttg ccgggaagct agagtaagta
     5941 gttcgccagt taatagtttg cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac
     6001 gctcgtcgtt tggtatggct tcattcagct ccggttccca acgatcaagg cgagttacat
     6061 gatcccccat gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc gttgtcagaa
     6121 gtaagttggc cgcagtgtta tcactcatgg ttatggcagc actgcataat tctcttactg
     6181 tcatgccatc cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag tcattctgag
     6241 aatagtgtat gcggcgaccg agttgctctt gcccggcgtc aatacgggat aataccgcgc
     6301 cacatagcag aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct
     6361 caaggatctt accgctgttg agatccagtt cgatgtaacc cactcgtgca cccaactgat
     6421 cttcagcatc ttttactttc accagcgttt ctgggtgagc aaaaacagga aggcaaaatg
     6481 ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat actcatactc ttcctttttc
     6541 aatattattg aagcatttat cagggttatt gtctcatgag cggatacata tttgaatgta
     6601 tttagaaaaa taaacaaata ggggttccgc gcacatttcc ccgaaaagtg ccacctgacg
     6661 tcgacggatc gggagatctc ccgatcccct atggtgcact ctcagtacaa tctgctctga
     6721 tgccgcatag ttaagccagt atctgctccc tgcttgtgtg ttggaggtcg ctgagtagtg
     6781 cgcgagcaaa atttaagcta caacaaggca aggcttgacc gacaattgca tgaagaatct
     6841 gcttagggtt aggcgttttg cgctgcttcg cgatgtacgg gccagatata cgcgttgaca
     6901 ttgattattg actagttatt aatagtaatc aattacgggg tcattagttc atagcccata
     6961 tatggagttc cgcgttacat aacttacggt aaatggcccg cctggctgac cgcccaacga
     7021 cccccgccca ttgacgtcaa taatgacgta tgttcccata gtaacgccaa tagggacttt
     7081 ccattgacgt caatgggtgg agtatttacg gtaaactgcc cacttggcag tacatcaagt
     7141 gtatcatatg ccaagtacgc cccctattga cgtcaatgac ggtaaatggc ccgcctggca
     7201 ttatgcccag tacatgacct tatgggactt tcctacttgg cagtacatct acgtattagt
     7261 catcgctatt accatggtga tgcggttttg gcagtacatc aatgggcgtg gatagcggtt
     7321 tgactcacgg ggatttccaa gtctccaccc cattgacgtc aatgggagtt tgttttggca
     7381 ccaaaatcaa cgggactttc caaaatgtcg taacaactcc gccccattga cgcaaatggg
     7441 cggtaggcgt gtacggtggg aggtctatat aagcagcgcg ttttgcctgt actgggtctc
     7501 tctggttaga ccagatctga gcctgggagc tctctggcta actagggaac ccactgctta
     7561 agcctcaata aagcttgcct tgagtgcttc aagtagtgtg tgcccgtctg ttgtgtgact
     7621 ctggtaacta gagatccctc agaccctttt agtcagtgtg gaaaatctct agcagtggcg
     7681 cccgaacagg gacttgaaag cgaaagggaa accagaggag ctctctcgac gcaggactcg
     7741 gcttgctgaa gcgcgcacgg caagaggcga ggggcggcga ctggtgagta cgccaaaaat
     7801 tttgactagc ggaggctaga aggagagaga tgggtgcgag agcgtcagta ttaagcgggg
     7861 gagaattaga tcgcgatggg aaaaaattcg gttaaggcca gggggaaaga aaaaatataa
     7921 attaaaacat atagtatggg caagcaggga gctagaacga ttcgcagtta atcctggcct
     7981 gttagaaaca tcagaaggct gtagacaaat actgggacag ctacaaccat cccttcagac
     8041 aggatcagaa gaacttagat cattatataa tacagtagca accctctatt gtgtgcatca
     8101 aaggatagag ataaaagaca ccaaggaagc tttagacaag atagaggaag agcaaaacaa
     8161 aagtaagacc accgcacagc aagcggccgg ccgctgatct tcagacctgg aggaggagat
     8221 atgagggaca attggagaag tgaattatat aaatataaag tagtaaaaat tgaaccatta
     8281 ggagtagcac ccaccaaggc aaagagaaga gtggtgcaga gagaaaaaag agcagtggga
     8341 ataggagctt tgttccttgg gttcttggga gcagcaggaa gcactatggg cgcagcgtca
     8401 atgacgctga cggtacaggc caga
//
