LOCUS       sequence-142916-        7850 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 7850)
  AUTHORS   Navarro F, Gutman D, Meire E, Caceres M, Rigoutsos I, Bentwich Z, 
            Lieberman J
  TITLE     miR-34a contributes to megakaryocytic differentiation of K562 cells 
            independently of p53.
  JOURNAL   Blood. 2009 Sep 3. 114(10):2181-92.
  PUBMED    19584398
REFERENCE   2  (bases 1 to 7850)
  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: "Blood. 2009
            Sep 3. 114(10):2181-92."
COMMENT     Sequence Label: pLL3.7_hsa-miR-34a
FEATURES             Location/Qualifiers
     source          1..7850
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        184..563
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        565..763
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     LTR             781..961
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    1008..1133
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1630..1863
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             2048..2092
                     /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    2390..2507
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        2581..2829
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     2581..2601
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     2752..2771
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     primer_bind     3098..3114
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    complement(3156..3189)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     enhancer        3305..3608
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        3609..3812
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     regulatory      3851..3860
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(3857..3859,3860..3862,3863..4576)
                     /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(3857..3859,3860..3862,3863..4573)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(3902..3923)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(4163..4182)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     4510..4531
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     protein_bind    complement(4596..4629)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     misc_feature    4685..5273
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(4738..4758)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(5156..5167)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(5276..5292)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     LTR             5802..5982
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     rep_origin      complement(6044..6632)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6124..6143)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6803..7594,7595..7663))
                     /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     7426..7445
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7664..7768)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(join(7843..7850,1..12))
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 gctctgatgc cgcatagtta agccagtatc tgctccctgc ttgtgtgttg gaggtcgctg
       61 agtagtgcgc gagcaaaatt taagctacaa caaggcaagg cttgaccgac aattgcatga
      121 agaatctgct tagggttagg cgttttgcgc tgcttcgcga tgtacgggcc agatatacgc
      181 gttgacattg attattgact agttattaat agtaatcaat tacggggtca ttagttcata
      241 gcccatatat ggagttccgc gttacataac ttacggtaaa tggcccgcct ggctgaccgc
      301 ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt tcccatagta acgccaatag
      361 ggactttcca ttgacgtcaa tgggtggagt atttacggta aactgcccac ttggcagtac
      421 atcaagtgta tcatatgcca agtacgcccc ctattgacgt caatgacggt aaatggcccg
      481 cctggcatta tgcccagtac atgaccttat gggactttcc tacttggcag tacatctacg
      541 tattagtcat cgctattacc atggtgatgc ggttttggca gtacatcaat gggcgtggat
      601 agcggtttga ctcacgggga tttccaagtc tccaccccat tgacgtcaat gggagtttgt
      661 tttggcacca aaatcaacgg gactttccaa aatgtcgtaa caactccgcc ccattgacgc
      721 aaatgggcgg taggcgtgta cggtgggagg tctatataag cagcgcgttt tgcctgtact
      781 gggtctctct ggttagacca gatctgagcc tgggagctct ctggctaact agggaaccca
      841 ctgcttaagc ctcaataaag cttgccttga gtgcttcaag tagtgtgtgc ccgtctgttg
      901 tgtgactctg gtaactagag atccctcaga cccttttagt cagtgtggaa aatctctagc
      961 agtggcgccc gaacagggac ttgaaagcga aagggaaacc agaggagctc tctcgacgca
     1021 ggactcggct tgctgaagcg cgcacggcaa gaggcgaggg gcggcgactg gtgagtacgc
     1081 caaaaatttt gactagcgga ggctagaagg agagagatgg gtgcgagagc gtcagtatta
     1141 agcgggggag aattagatcg cgatgggaaa aaattcggtt aaggccaggg ggaaagaaaa
     1201 aatataaatt aaaacatata gtatgggcaa gcagggagct agaacgattc gcagttaatc
     1261 ctggcctgtt agaaacatca gaaggctgta gacaaatact gggacagcta caaccatccc
     1321 ttcagacagg atcagaagaa cttagatcat tatataatac agtagcaacc ctctattgtg
     1381 tgcatcaaag gatagagata aaagacacca aggaagcttt agacaagata gaggaagagc
     1441 aaaacaaaag taagaccacc gcacagcaag cggccggccg ctgatcttca gacctggagg
     1501 aggagatatg agggacaatt ggagaagtga attatataaa tataaagtag taaaaattga
     1561 accattagga gtagcaccca ccaaggcaaa gagaagagtg gtgcagagag aaaaaagagc
     1621 agtgggaata ggagctttgt tccttgggtt cttgggagca gcaggaagca ctatgggcgc
     1681 agcgtcaatg acgctgacgg tacaggccag acaattattg tctggtatag tgcagcagca
     1741 gaacaatttg ctgagggcta ttgaggcgca acagcatctg ttgcaactca cagtctgggg
     1801 catcaagcag ctccaggcaa gaatcctggc tgtggaaaga tacctaaagg atcaacagct
     1861 cctggggatt tggggttgct ctggaaaact catttgcacc actgctgtgc cttggaatgc
     1921 tagttggagt aataaatctc tggaacagat ttggaatcac acgacctgga tggagtggga
     1981 cagagaaatt aacaattaca caagcttaat acactcctta attgaagaat cgcaaaacca
     2041 gcaagaaaag aatgaacaag aattattgga attagataaa tgggcaagtt tgtggaattg
     2101 gtttaacata acaaattggc tgtggtatat aaaattattc ataatgatag taggaggctt
     2161 ggtaggttta agaatagttt ttgctgtact ttctatagtg aatagagtta ggcagggata
     2221 ttcaccatta tcgtttcaga cccacctccc aaccccgagg ggacccgaca ggcccgaagg
     2281 aatagaagaa gaaggtggag agagagacag agacagatcc attcgattag tgaacggatc
     2341 ggcactgcgt gcgccaattc tgcagacaaa tggcagtatt catccacaat tttaaaagaa
     2401 aaggggggat tggggggtac agtgcagggg aaagaatagt agacataata gcaacagaca
     2461 tacaaactaa agaattacaa aaacaaatta caaaaattca aaattttcgg gtttattaca
     2521 gggacagcag agatccagtt tggttagtac cgggcccgct ctagaagatc tgggcaggaa
     2581 gagggcctat ttcccatgat tccttcatat ttgcatatac gatacaaggc tgttagagag
     2641 ataattagaa ttaatttgac tgtaaacaca aagatattag tacaaaatac gtgacgtaga
     2701 aagtaataat ttcttgggta gtttgcagtt ttaaaattat gttttaaaat ggactatcat
     2761 atgcttaccg taacttgaaa gtatttcgat ttcttggctt tatatatctt gtggaaagga
     2821 cgaggatccc ccacatttcc ttcttatcaa caggtgctgg ggagaggcag gacaggcctg
     2881 tcccccgagt cccctccgga tgccgtggac cggccagctg tgagtgtttc tttggcagtg
     2941 tcttagctgg ttgttgtgag caatagtaag gaagcaatca gcaagtatac tgccctagaa
     3001 gtgctgcacg ttgtggggcc caagagggaa gatgaagcga gagatgccca gaccagtggg
     3061 agacgccagg acttcggaag ctcttctgcg aattcctcga ggtcgacggt atcgataagc
     3121 tcgcttcacg agattccagc aggtcgaggg acctaataac ttcgtatagc atacattata
     3181 cgaagttata ttaagggttc caagcttaag cggccgcgtg gataaccgta ttaccgccat
     3241 gcattagtta ttaatagtaa tcaattacgg ggtcattagt tcatagccca tatatggagt
     3301 tccgcgttac ataacttacg gtaaatggcc cgcctggctg accgcccaac gacccccgcc
     3361 cattgacgtc aataatgacg tatgttccca tagtaacgcc aatagggact ttccattgac
     3421 gtcaatgggt ggagtattta cggtaaactg cccacttggc agtacatcaa gtgtatcata
     3481 tgccaagtac gccccctatt gacgtcaatg acggtaaatg gcccgcctgg cattatgccc
     3541 agtacatgac cttatgggac tttcctactt ggcagtacat ctacgtatta gtcatcgcta
     3601 ttaccatggt gatgcggttt tggcagtaca tcaatgggcg tggatagcgg tttgactcac
     3661 ggggatttcc aagtctccac cccattgacg tcaatgggag tttgttttgg caccaaaatc
     3721 aacgggactt tccaaaatgt cgtaacaact ccgccccatt gacgcaaatg ggcggtaggc
     3781 gtgtacggtg ggaggtctat ataagcagag ctggtttagt gaaccgtcag atccgctagc
     3841 gctaccggtc gccaccatgg tgagcaaggg cgaggagctg ttcaccgggg tggtgcccat
     3901 cctggtcgag ctggacggcg acgtaaacgg ccacaagttc agcgtgtccg gcgagggcga
     3961 gggcgatgcc acctacggca agctgaccct gaagttcatc tgcaccaccg gcaagctgcc
     4021 cgtgccctgg cccaccctcg tgaccaccct gacctacggc gtgcagtgct tcagccgcta
     4081 ccccgaccac atgaagcagc acgacttctt caagtccgcc atgcccgaag gctacgtcca
     4141 ggagcgcacc atcttcttca aggacgacgg caactacaag acccgcgccg aggtgaagtt
     4201 cgagggcgac accctggtga accgcatcga gctgaagggc atcgacttca aggaggacgg
     4261 caacatcctg gggcacaagc tggagtacaa ctacaacagc cacaacgtct atatcatggc
     4321 cgacaagcag aagaacggca tcaaggtgaa cttcaagatc cgccacaaca tcgaggacgg
     4381 cagcgtgcag ctcgccgacc actaccagca gaacaccccc atcggcgacg gccccgtgct
     4441 gctgcccgac aaccactacc tgagcaccca gtccgccctg agcaaagacc ccaacgagaa
     4501 gcgcgatcac atggtcctgc tggagttcgt gaccgccgcc gggatcactc tcggcatgga
     4561 cgagctgtac aagtaggaat tcgtcgaggg acctaataac ttcgtatagc atacattata
     4621 cgaagttata catgtttaag ggttccggtt ccactaggta caattcgata tcaagcttat
     4681 cgataatcaa cctctggatt acaaaatttg tgaaagattg actggtattc ttaactatgt
     4741 tgctcctttt acgctatgtg gatacgctgc tttaatgcct ttgtatcatg ctattgcttc
     4801 ccgtatggct ttcattttct cctccttgta taaatcctgg ttgctgtctc tttatgagga
     4861 gttgtggccc gttgtcaggc aacgtggcgt ggtgtgcact gtgtttgctg acgcaacccc
     4921 cactggttgg ggcattgcca ccacctgtca gctcctttcc gggactttcg ctttccccct
     4981 ccctattgcc acggcggaac tcatcgccgc ctgccttgcc cgctgctgga caggggctcg
     5041 gctgttgggc actgacaatt ccgtggtgtt gtcggggaaa tcatcgtcct ttccttggct
     5101 gctcgcctgt gttgccacct ggattctgcg cgggacgtcc ttctgctacg tcccttcggc
     5161 cctcaatcca gcggaccttc cttcccgcgg cctgctgccg gctctgcggc ctcttccgcg
     5221 tcttcgcctt cgccctcaga cgagtcggat ctccctttgg gccgcctccc cgcatcgata
     5281 ccgtcgacct cgatcgagac ctagaaaaac atggagcaat cacaagtagc aatacagcag
     5341 ctaccaatgc tgattgtgcc tggctagaag cacaagagga ggaggaggtg ggttttccag
     5401 tcacacctca ggtaccttta agaccaatga cttacaaggc agctgtagat cttagccact
     5461 ttttaaaaga aaagggggga ctggaagggc taattcactc ccaacgaaga caagatatcc
     5521 ttgatctgtg gatctaccac acacaaggct acttccctga ttggcagaac tacacaccag
     5581 ggccagggat cagatatcca ctgacctttg gatggtgcta caagctagta ccagttgagc
     5641 aagagaaggt agaagaagcc aatgaaggag agaacacccg cttgttacac cctgtgagcc
     5701 tgcatgggat ggatgacccg gagagagaag tattagagtg gaggtttgac agccgcctag
     5761 catttcatca catggcccga gagctgcatc cggactgtac tgggtctctc tggttagacc
     5821 agatctgagc ctgggagctc tctggctaac tagggaaccc actgcttaag cctcaataaa
     5881 gcttgccttg agtgcttcaa gtagtgtgtg cccgtctgtt gtgtgactct ggtaactaga
     5941 gatccctcag acccttttag tcagtgtgga aaatctctag cagcatgtga gcaaaaggcc
     6001 agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc
     6061 cccctgacga gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac
     6121 tataaagata ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc
     6181 tgccgcttac cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata
     6241 gctcacgctg taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc
     6301 acgaaccccc cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca
     6361 acccggtaag acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag
     6421 cgaggtatgt aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta
     6481 gaagaacagt atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg
     6541 gtagctcttg atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc
     6601 agcagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt
     6661 ctgacgctca gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa
     6721 ggatcttcac ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat
     6781 atgagtaaac ttggtctgac agttaccaat gcttaatcag tgaggcacct atctcagcga
     6841 tctgtctatt tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatac
     6901 gggagggctt accatctggc cccagtgctg caatgatacc gcgagaccca cgctcaccgg
     6961 ctccagattt atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg
     7021 caactttatc cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt
     7081 cgccagttaa tagtttgcgc aacgttgttg ccattgctac aggcatcgtg gtgtcacgct
     7141 cgtcgtttgg tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat
     7201 cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta
     7261 agttggccgc agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca
     7321 tgccatccgt aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat
     7381 agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac
     7441 atagcagaac tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa
     7501 ggatcttacc gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt
     7561 cagcatcttt tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg
     7621 caaaaaaggg aataagggcg acacggaaat gttgaatact catactcttc ctttttcaat
     7681 attattgaag catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt
     7741 agaaaaataa acaaataggg gttccgcgca catttccccg aaaagtgcca cctgacgtcg
     7801 acggatcggg agatctcccg atcccctatg gtgcactctc agtacaatct
//
