LOCUS       sequence-211558-        7507 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7507)
  AUTHORS   Beronja S, Livshits G, Williams S, Fuchs E
  TITLE     Rapid functional dissection of genetic networks via tissue-specific 
            transduction and RNAi in mouse embryos.
  JOURNAL   Nat Med. 2010 Jul . 16(7):821-7.
  PUBMED    20526348
REFERENCE   2  (bases 1 to 7507)
  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: "Nat Med. 
            2010 Jul . 16(7):821-7."
COMMENT     Sequence Label: LV-GFP
FEATURES             Location/Qualifiers
     source          1..7507
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     426..444
                     /label=hPGK-F
                     /note="Human PGK promoter, forward primer"
     CDS             519..896
                     /codon_start=1
                     /gene="HIST1H2BJ"
                     /product="human histone H2B"
                     /label=H2B
                     /translation="MPEPAKSAPAPKKGSKKAVTKAQKKGGKKRKRSRKESYSIYVYKV
                     LKQVHPDTGISSKAMGIMNSFVNDIFERIAGEASRLAHYNKRSTITSREIQTAVRLLLP
                     GELAKHAVSEGTKAITKYTSAK"
     regulatory      909..918
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(915..917,918..920,921..1634)
                     /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(915..917,918..920,921..1631)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(960..981)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1221..1240)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1568..1589
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     LTR             1713..1946
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    2018..2139
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2055..2074)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2109..2128
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      2179..2314
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(2334..2353)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2335..2353)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2363..2380)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2363..2379)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2372..2394)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      2521..2976
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2608..2627)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2818..2839
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        3002..3106
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3107..3175,3176..3967)
                     /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     complement(3325..3344)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4138..4726
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4627..4646
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4880..4897
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    5014..5035
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(5050..5055,5056..5073,5074..5080)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5088..5104
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     primer_bind     5093..5115
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     5112..5128
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5112..5128
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     5147..5167
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        5149..5167
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        5195..5421
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             5422..5602
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    5649..5774
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    6267..6500
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             6685..6729
                     /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"
     promoter        7027..7275
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     7027..7047
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     7198..7217
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_feature    7338..7455
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        join(7504..7507,1..507)
                     /label=hPGK promoter
                     /note="human phosphoglycerate kinase 1 promoter"
ORIGIN
        1 ttggggttgc gccttttcca aggcagccct gggtttgcgc agggacgcgg ctgctctggg
       61 cgtggttccg ggaaacgcag cggcgccgac cctgggtctc gcacattctt cacgtccgtt
      121 cgcagcgtca cccggatctt cgccgctacc cttgtgggcc ccccggcgac gcttcctgct
      181 ccgcccctaa gtcgggaagg ttccttgcgg ttcgcggcgt gccggacgtg acaaacggaa
      241 gccgcacgtc tcactagtac cctcgcagac ggacagcgcc agggagcaat ggcagcgcgc
      301 cgaccgcgat gggctgtggc caatagcggc tgctcagcag ggcgcgccga gagcagcggc
      361 cgggaagggg cggtgcggga ggcggggtgt ggggcggtag tgtgggccct gttcctgccc
      421 gcgcggtgtt ccgcattctg caagcctccg gagcgcacgt cggcagtcgg ctccctcgtt
      481 gaccgaatca ccgacctctc tccccagggg gatctaccat gccagagcca gcgaagtctg
      541 ctcccgcccc gaaaaagggc tccaagaagg cggtgactaa ggcgcagaag aaaggcggca
      601 agaagcgcaa gcgcagccgc aaggagagct attccatcta tgtgtacaag gttctgaagc
      661 aggtccaccc tgacaccggc atttcgtcca aggccatggg catcatgaac tcgtttgtga
      721 acgacatttt cgagcgcatc gcaggtgagg cttcccgcct ggcgcattac aacaagcgct
      781 cgaccatcac ctccagggag atccagacgg ccgtgcgcct gctgctgcct ggggagttgg
      841 ccaagcacgc cgtgtccgag ggtactaagg ccatcaccaa gtacaccagc gctaaggatc
      901 caccggtcgc caccatggtg agcaagggcg aggagctgtt caccggggtg gtgcccatcc
      961 tggtcgagct ggacggcgac gtaaacggcc acaagttcag cgtgtccggc gagggcgagg
     1021 gcgatgccac ctacggcaag ctgaccctga agttcatctg caccaccggc aagctgcccg
     1081 tgccctggcc caccctcgtg accaccctga cctacggcgt gcagtgcttc agccgctacc
     1141 ccgaccacat gaagcagcac gacttcttca agtccgccat gcccgaaggc tacgtccagg
     1201 agcgcaccat cttcttcaag gacgacggca actacaagac ccgcgccgag gtgaagttcg
     1261 agggcgacac cctggtgaac cgcatcgagc tgaagggcat cgacttcaag gaggacggca
     1321 acatcctggg gcacaagctg gagtacaact acaacagcca caacgtctat atcatggccg
     1381 acaagcagaa gaacggcatc aaggtgaact tcaagatccg ccacaacatc gaggacggca
     1441 gcgtgcagct cgccgaccac taccagcaga acacccccat cggcgacggc cccgtgctgc
     1501 tgcccgacaa ccactacctg agcacccagt ccgccctgag caaagacccc aacgagaagc
     1561 gcgatcacat ggtcctgctg gagttcgtga ccgccgccgg gatcactctc ggcatggacg
     1621 agctgtacaa gtaaatgcat cggtaccttt aagaccaatg acttacaagg cagctgtaga
     1681 tcttagccac tttttaaaag aaaagggggg actggaaggg ctaattcact cccaacgaag
     1741 acaagatctg ctttttgctt gtactgggtc tctctggtta gaccagatct gagcctggga
     1801 gctctctggc taactaggga acccactgct taagcctcaa taaagcttgc cttgagtgct
     1861 tcaagtagtg tgtgcccgtc tgttgtgtga ctctggtaac tagagatccc tcagaccctt
     1921 ttagtcagtg tggaaaatct ctagcagtag tagttcatgt catcttatta ttcagtattt
     1981 ataacttgca aagaaatgaa tatcagagag tgagaggaac ttgtttattg cagcttataa
     2041 tggttacaaa taaagcaata gcatcacaaa tttcacaaat aaagcatttt tttcactgca
     2101 ttctagttgt ggtttgtcca aactcatcaa tgtatcttat catgtctggc tctagctatc
     2161 ccgcccctaa ctccgcccat cccgccccta actccgccca gttccgccca ttctccgccc
     2221 catggctgac taattttttt tatttatgca gaggccgagg ccgcctcggc ctctgagcta
     2281 ttccagaagt agtgaggagg cttttttgga ggcctaggga cgtacccaat tcgccctata
     2341 gtgagtcgta ttacgcgcgc tcactggccg tcgttttaca acgtcgtgac tgggaaaacc
     2401 ctggcgttac ccaacttaat cgccttgcag cacatccccc tttcgccagc tggcgtaata
     2461 gcgaagaggc ccgcaccgat cgcccttccc aacagttgcg cagcctgaat ggcgaatggg
     2521 acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg
     2581 ctacacttgc cagcgcccta gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca
     2641 cgttcgccgg ctttccccgt caagctctaa atcgggggct ccctttaggg ttccgattta
     2701 gtgctttacg gcacctcgac cccaaaaaac ttgattaggg tgatggttca cgtagtgggc
     2761 catcgccctg atagacggtt tttcgccctt tgacgttgga gtccacgttc tttaatagtg
     2821 gactcttgtt ccaaactgga acaacactca accctatctc ggtctattct tttgatttat
     2881 aagggatttt gccgatttcg gcctattggt taaaaaatga gctgatttaa caaaaattta
     2941 acgcgaattt taacaaaata ttaacgctta caatttaggt ggcacttttc ggggaaatgt
     3001 gcgcggaacc cctatttgtt tatttttcta aatacattca aatatgtatc cgctcatgag
     3061 acaataaccc tgataaatgc ttcaataata ttgaaaaagg aagagtatga gtattcaaca
     3121 tttccgtgtc gcccttattc ccttttttgc ggcattttgc cttcctgttt ttgctcaccc
     3181 agaaacgctg gtgaaagtaa aagatgctga agatcagttg ggtgcacgag tgggttacat
     3241 cgaactggat ctcaacagcg gtaagatcct tgagagtttt cgccccgaag aacgttttcc
     3301 aatgatgagc acttttaaag ttctgctatg tggcgcggta ttatcccgta ttgacgccgg
     3361 gcaagagcaa ctcggtcgcc gcatacacta ttctcagaat gacttggttg agtactcacc
     3421 agtcacagaa aagcatctta cggatggcat gacagtaaga gaattatgca gtgctgccat
     3481 aaccatgagt gataacactg cggccaactt acttctgaca acgatcggag gaccgaagga
     3541 gctaaccgct tttttgcaca acatggggga tcatgtaact cgccttgatc gttgggaacc
     3601 ggagctgaat gaagccatac caaacgacga gcgtgacacc acgatgcctg tagcaatggc
     3661 aacaacgttg cgcaaactat taactggcga actacttact ctagcttccc ggcaacaatt
     3721 aatagactgg atggaggcgg ataaagttgc aggaccactt ctgcgctcgg cccttccggc
     3781 tggctggttt attgctgata aatctggagc cggtgagcgt gggtctcgcg gtatcattgc
     3841 agcactgggg ccagatggta agccctcccg tatcgtagtt atctacacga cggggagtca
     3901 ggcaactatg gatgaacgaa atagacagat cgctgagata ggtgcctcac tgattaagca
     3961 ttggtaactg tcagaccaag tttactcata tatactttag attgatttaa aacttcattt
     4021 ttaatttaaa aggatctagg tgaagatcct ttttgataat ctcatgacca aaatccctta
     4081 acgtgagttt tcgttccact gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg
     4141 agatcctttt tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac cgctaccagc
     4201 ggtggtttgt ttgccggatc aagagctacc aactcttttt ccgaaggtaa ctggcttcag
     4261 cagagcgcag ataccaaata ctgttcttct agtgtagccg tagttaggcc accacttcaa
     4321 gaactctgta gcaccgccta catacctcgc tctgctaatc ctgttaccag tggctgctgc
     4381 cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac cggataaggc
     4441 gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc agcttggagc gaacgaccta
     4501 caccgaactg agatacctac agcgtgagct atgagaaagc gccacgcttc ccgaagggag
     4561 aaaggcggac aggtatccgg taagcggcag ggtcggaaca ggagagcgca cgagggagct
     4621 tccaggggga aacgcctggt atctttatag tcctgtcggg tttcgccacc tctgacttga
     4681 gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc
     4741 ggccttttta cggttcctgg ccttttgctg gccttttgct cacatgttct ttcctgcgtt
     4801 atcccctgat tctgtggata accgtattac cgcctttgag tgagctgata ccgctcgccg
     4861 cagccgaacg accgagcgca gcgagtcagt gagcgaggaa gcggaagagc gcccaatacg
     4921 caaaccgcct ctccccgcgc gttggccgat tcattaatgc agctggcacg acaggtttcc
     4981 cgactggaaa gcgggcagtg agcgcaacgc aattaatgtg agttagctca ctcattaggc
     5041 accccaggct ttacacttta tgcttccggc tcgtatgttg tgtggaattg tgagcggata
     5101 acaatttcac acaggaaaca gctatgacca tgattacgcc aagcgcgcaa ttaaccctca
     5161 ctaaagggaa caaaagctgg agctgcaagc ttaatgtagt cttatgcaat actcttgtag
     5221 tcttgcaaca tggtaacgat gagttagcaa catgccttac aaggagagaa aaagcaccgt
     5281 gcatgccgat tggtggaagt aaggtggtac gatcgtgcct tattaggaag gcaacagacg
     5341 ggtctgacat ggattggacg aaccactgaa ttgccgcatt gcagagatat tgtatttaag
     5401 tgcctagctc gatacataaa cgggtctctc tggttagacc agatctgagc ctgggagctc
     5461 tctggctaac tagggaaccc actgcttaag cctcaataaa gcttgccttg agtgcttcaa
     5521 gtagtgtgtg cccgtctgtt gtgtgactct ggtaactaga gatccctcag acccttttag
     5581 tcagtgtgga aaatctctag cagtggcgcc cgaacaggga cttgaaagcg aaagggaaac
     5641 cagaggagct ctctcgacgc aggactcggc ttgctgaagc gcgcacggca agaggcgagg
     5701 ggcggcgact ggtgagtacg ccaaaaattt tgactagcgg aggctagaag gagagagatg
     5761 ggtgcgagag cgtcagtatt aagcggggga gaattagatc gcgatgggaa aaaattcggt
     5821 taaggccagg gggaaagaaa aaatataaat taaaacatat agtatgggca agcagggagc
     5881 tagaacgatt cgcagttaat cctggcctgt tagaaacatc agaaggctgt agacaaatac
     5941 tgggacagct acaaccatcc cttcagacag gatcagaaga acttagatca ttatataata
     6001 cagtagcaac cctctattgt gtgcatcaaa ggatagagat aaaagacacc aaggaagctt
     6061 tagacaagat agaggaagag caaaacaaaa gtaagaccac cgcacagcaa gcggccgctg
     6121 atcttcagac ctggaggagg agatatgagg gacaattgga gaagtgaatt atataaatat
     6181 aaagtagtaa aaattgaacc attaggagta gcacccacca aggcaaagag aagagtggtg
     6241 cagagagaaa aaagagcagt gggaatagga gctttgttcc ttgggttctt gggagcagca
     6301 ggaagcacta tgggcgcagc gtcaatgacg ctgacggtac aggccagaca attattgtct
     6361 ggtatagtgc agcagcagaa caatttgctg agggctattg aggcgcaaca gcatctgttg
     6421 caactcacag tctggggcat caagcagctc caggcaagaa tcctggctgt ggaaagatac
     6481 ctaaaggatc aacagctcct ggggatttgg ggttgctctg gaaaactcat ttgcaccact
     6541 gctgtgcctt ggaatgctag ttggagtaat aaatctctgg aacagatttg gaatcacacg
     6601 acctggatgg agtgggacag agaaattaac aattacacaa gcttaataca ctccttaatt
     6661 gaagaatcgc aaaaccagca agaaaagaat gaacaagaat tattggaatt agataaatgg
     6721 gcaagtttgt ggaattggtt taacataaca aattggctgt ggtatataaa attattcata
     6781 atgatagtag gaggcttggt aggtttaaga atagtttttg ctgtactttc tatagtgaat
     6841 agagttaggc agggatattc accattatcg tttcagaccc acctcccaac cccgagggga
     6901 cccgacaggc ccgaaggaat agaagaagaa ggtggagaga gagacagaga cagatccatt
     6961 cgattagtga acggatctcg acggtatcga tcacgagact agcctcgagc ggccgccccc
     7021 ttcaccgagg gcctatttcc catgattcct tcatatttgc atatacgata caaggctgtt
     7081 agagagataa ttggaattaa tttgactgta aacacaaaga tattagtaca aaatacgtga
     7141 cgtagaaagt aataatttct tgggtagttt gcagttttaa aattatgttt taaaatggac
     7201 tatcatatgc ttaccgtaac ttgaaagtat ttcgatttct tggctttata tatcttgtgg
     7261 aaaggacgaa acaccggtcc gcaggtatgc acgcgtgaat tctcgacctc gagacaaatg
     7321 gcagtattca tccacaattt taaaagaaaa ggggggattg gggggtacag tgcaggggaa
     7381 agaatagtag acataatagc aacagacata caaactaaag aattacaaaa acaaattaca
     7441 aaaattcaaa attttcgggt ttattacagg gacagcagag atccactttg gccgcggctc
     7501 gaggggg
//
