LOCUS       sequence-6818-ex        7722 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 7722)
  AUTHORS   Landthaler M, Yalcin A, Tuschl T
  TITLE     The human DiGeorge syndrome critical region gene 8 and Its D. 
            melanogaster homolog are required for miRNA biogenesis.
  JOURNAL   Curr Biol. 2004 Dec 14. 14(23):2162-7.
  PUBMED    15589161
REFERENCE   2  (bases 1 to 7722)
  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: "Curr Biol. 
            2004 Dec 14. 14(23):2162-7."
COMMENT     Sequence Label: pFLAG/HA-DGCR8
FEATURES             Location/Qualifiers
     source          1..7722
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     863..882
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      946..955
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             955..978
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             979..1005
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     protein_bind    1021..1045
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     protein_bind    complement(3417..3441)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             3494..3535
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     intron          3616..3845
                     /label=chimeric intron
                     /note="chimera between introns from adenovirus and 
                     immunoglobulin heavy chain genes"
     primer_bind     3828..3847
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     misc_feature    join(3903..4477,4478..4480,4481..4489)
                     /label=IRES2
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(4070..4087)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     4297..4316
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             4509..5108
                     /codon_start=1
                     /gene="pac from Streptomyces"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVECPKDRATWCMTRKPGA"
     primer_bind     complement(4509..4528)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     5005..5025
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    5342..5463
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5379..5398)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(5565..5581)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5565..5581)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5578..5600)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5589..5605
                     /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)."
     promoter        complement(join(5613..5619,5620..5637,5638..5643))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5658..5679
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5796..5813)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5967..6555)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6047..6066)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6726..7517,7518..7586))
                     /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     7349..7368
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7587..7691)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtcgactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggac tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gcttggtacc
      901 gagctcggat cgatccatcg atagtgtggt ggaattctga tcgccgccgc catggactac
      961 aaggacgacg atgacaagta cccttatgac gtgcccgatt acgctttgaa ccagatatca
     1021 acaagtttgt acaaaaaagc aggctccacc atgggaacca attcagtcga catggagaca
     1081 gatgagagcc cctctccgct cccgtgtggg cccgcaggag aagcggtgat ggagagccga
     1141 gctcgcccct tccaagcgct gccccgtgag cagtctccac cacctcccct gcaaacgtcc
     1201 agtggtgcag aggtaatgga cgttggctct ggtggtgatg gacagtccga actccctgct
     1261 gaggacccct tcaacttcta cggagcttct cttctctcca aaggatcctt ctctaagggc
     1321 cgcctcctca tagacccgaa ctgtagtggc cacagcccgc gcaccgcccg gcacgcacct
     1381 gcggtccgga agttctcccc tgaccttaag ttgcttaagg atgtaaagat tagcgtgagc
     1441 tttaccgaga gctgcaggag taaggacagg aaggtgctgt acacaggagc agagcgcgac
     1501 gtgcgggcgg agtgcggtct gctccttagc cctgtcagtg gggacgtgca tgcttgtccc
     1561 tttggcggga gtgttggtga cggggtaggc atagggggtg agagtgctga taagaaggat
     1621 gaggagaatg agctggatca ggaaaagaga gtggagtatg cagtgctcga tgagttagaa
     1681 gattttactg acaatttgga gctagatgaa gaaggagcag gcgggttcac ggctaaagca
     1741 atcgttcaga gagacagagt ggatgaagag gccttgaatt tcccctacga ggatgacttt
     1801 gacaacgatg tggatgctct gctggaagaa ggcctttgtg cccccaaaaa gaggcgaaca
     1861 gaggaaaaat atggcggaga cagcgaccat ccgtccgatg gagagacaag tgtgcagccg
     1921 atgatgacca agattaaaac agtgctcaaa agtcgtggcc gcccacctac agagccgctg
     1981 cccgacgggt ggatcatgac attccataac tctggagtcc cggtgtacct acacagagag
     2041 tctcgggtgg tcacctggtc caggccatac ttcttgggaa cgggaagcat acggaaacac
     2101 gaccctcctc tgagtagcat cccttgtctg cattataaga aaatgaagga caacgaggaa
     2161 cgggagcaaa gcagtgacct cacccctagt ggggatgtgt cccccgtcaa gcccctgagc
     2221 cgatctgcag agctggagtt tcccctggat gagcctgact ctatgggtgc tgacccgggg
     2281 cccccggacg agaaagaccc actaggggct gaggcagccc ctggggccct ggggcaggtg
     2341 aaggccaaag tcgaggtgtg caaagatgaa tccgttgatc tcgaggaatt tcgaagctac
     2401 ctggagaagc gttttgactt tgagcaagtt actgtgaaaa aattcaggac ttgggctgag
     2461 cggcggcaat tcaatcggga aatgaagcgg aagcaggcgg agtccgagag gcccatcttg
     2521 ccagccaatc agaagctcat tactttatca gtgcaagatg cacccacaaa gaaagagttt
     2581 gttattaacc ccaacgggaa atccgaggtc tgcatcctgc acgagtacat gcagcgtgtc
     2641 ctcaaggtcc gccctgtcta taatttcttt gaatgtgaga acccaagtga gccttttggt
     2701 gcctcggtga ccattgatgg tgtgacttac ggatctggaa ctgcaagcag caaaaaactt
     2761 gcgaagaata aagctgcccg agctacactg gaaatcctca tccctgactt tgttaaacag
     2821 acctctgaag agaagcccaa agacagtgaa gaactcgagt attttaacca catcagcatc
     2881 gaggactcgc gggtctacga gctgaccagc aaggctgggc tgttgtctcc atatcagatc
     2941 ctccacgagt gccttaaaag aaaccatggg atgggtgaca cgtctatcaa gtttgaagtg
     3001 gttcctggga aaaaccagaa gagtgaatac gtcatggcgt gtggcaagca cacagtgcgc
     3061 gggtggtgta agaacaagag agttggaaag cagttagcct cacagaagat ccttcagctg
     3121 ctgcacccac atgtcaagaa ctgggggtct ttactgcgca tgtatggccg tgagagcagc
     3181 aagatggtca agcaggagac atcggacaag agtgtgattg agctgcagca gtatgccaag
     3241 aagaacaagc ccaacctgca catcctcagc aagctccaag aggagatgaa gaggctagct
     3301 gaggaaaggg aggagactcg aaagaagccc aagatgtcca ttgtggcgtc cgcccagcct
     3361 ggcggtgagc ccctgtgcac cgtggacgtg tgagcggccg cactcgagat atctagaccc
     3421 agctttcttg tacaaagtgg ttgatatcca gcacagtggc ggccgctcaa gtctagaggg
     3481 cccgcggttc gaaggtaagc ctatccctaa ccctctcctc ggtctcgatt ctacgcgtac
     3541 ctgatccgcg gccgcataga taactgatcc agtgtgctgg aattaattcg ctgtctgcga
     3601 gggccagctg ttggggtgag tactccctct caaaagcggg catgacttct gcgctaagat
     3661 tgtcagtttc caaaaacgag gaggatttga tattcacctg gcccgcggtg atgcctttga
     3721 gggtggccgc gtccatctgg tcagaaaaga caatcttttt gttgtcaagc ttgaggtgtg
     3781 gcaggcttga gatctggcca tacacttgag tgacaatgac atccactttg cctttctctc
     3841 cacaggtgtc cactcccagg tccaactgca ggtcgagcat gcatctaggg cggccaattc
     3901 cgcccctctc cctccccccc ccctaacgtt actggccgaa gccgcttgga ataaggccgg
     3961 tgtgcgtttg tctatatgtg attttccacc atattgccgt cttttggcaa tgtgagggcc
     4021 cggaaacctg gccctgtctt cttgacgagc attcctaggg gtctttcccc tctcgccaaa
     4081 ggaatgcaag gtctgttgaa tgtcgtgaag gaagcagttc ctctggaagc ttcttgaaga
     4141 caaacaacgt ctgtagcgac cctttgcagg cagcggaacc ccccacctgg cgacaggtgc
     4201 ctctgcggcc aaaagccacg tgtataagat acacctgcaa aggcggcaca accccagtgc
     4261 cacgttgtga gttggatagt tgtggaaaga gtcaaatggc tctcctcaag cgtattcaac
     4321 aaggggctga aggatgccca gaaggtaccc cattgtatgg gatctgatct ggggcctcgg
     4381 tgcacatgct ttacatgtgt ttagtcgagg ttaaaaaaac gtctaggccc cccgaaccac
     4441 ggggacgtgg ttttcctttg aaaaacacga tgataagctt gccacaaccc acaaggagac
     4501 gaccttccat gaccgagtac aagcccacgg tgcgcctcgc cacccgcgac gacgtccccc
     4561 gggccgtacg caccctcgcc gccgcgttcg ccgactaccc cgccacgcgc cacaccgtcg
     4621 acccggaccg ccacatcgag cgggtcaccg agctgcaaga actcttcctc acgcgcgtcg
     4681 ggctcgacat cggcaaggtg tgggtcgcgg acgacggcgc cgcggtggcg gtctggacca
     4741 cgccggagag cgtcgaagcg ggggcggtgt tcgccgagat cggcccgcgc atggccgagt
     4801 tgagcggttc ccggctggcc gcgcagcaac agatggaagg cctcctggcg ccgcaccggc
     4861 ccaaggagcc cgcgtggttc ctggccaccg tcggcgtctc gcccgaccac cagggcaagg
     4921 gtctgggcag cgccgtcgtg ctccccggag tggaggcggc cgagcgcgcc ggggtgcccg
     4981 ccttcctgga gacctccgcg ccccgcaacc tccccttcta cgagcggctc ggcttcaccg
     5041 tcaccgccga cgtcgagtgc ccgaaggacc gcgcgacctg gtgcatgacc cgcaagcccg
     5101 gtgcctgacg cccgccccac gacccgcagc gcccgaccga aaggagcgca cgaccccatg
     5161 gctccgaccg aagccgaccc gggcggcccc gccgaccccg cacccgcccc cgaggcccac
     5221 cgactctaga taactgatca taatcagcca taccacattt gtagaggttt tacttgcttt
     5281 aaaaaacctc ccacacctcc ccctgaacct gaaacataaa atgaatgcaa ttgttgttgt
     5341 taacttgttt attgcagctt ataatggtta caaataaagc aatagcatca caaatttcac
     5401 aaataaagca tttttttcac tgcattctag ttgtggtttg tccaaactca tcaatgtatc
     5461 ttaacgcgtc gagtgcattc tagttgtggt ttgtccaaac tcatcaatgt atcttatcat
     5521 gtctgtatac cgtcgacctc tagctagagc ttggcgtaat catggtcata gctgtttcct
     5581 gtgtgaaatt gttatccgct cacaattcca cacaacatac gagccggaag cataaagtgt
     5641 aaagcctggg gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg ctcactgccc
     5701 gctttccagt cgggaaacct gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg
     5761 agaggcggtt tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg
     5821 gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg cggtaatacg gttatccaca
     5881 gaatcagggg ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac
     5941 cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac
     6001 aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag ataccaggcg
     6061 tttccccctg gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac
     6121 ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc aatgctcacg ctgtaggtat
     6181 ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag
     6241 cccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccggt aagacacgac
     6301 ttatcgccac tggcagcagc cactggtaac aggattagca gagcgaggta tgtaggcggt
     6361 gctacagagt tcttgaagtg gtggcctaac tacggctaca ctagaaggac agtatttggt
     6421 atctgcgctc tgctgaagcc agttaccttc ggaaaaagag ttggtagctc ttgatccggc
     6481 aaacaaacca ccgctggtag cggtggtttt tttgtttgca agcagcagat tacgcgcaga
     6541 aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc tcagtggaac
     6601 gaaaactcac gttaagggat tttggtcatg agattatcaa aaaggatctt cacctagatc
     6661 cttttaaatt aaaaatgaag ttttaaatca atctaaagta tatatgagta aacttggtct
     6721 gacagttacc aatgcttaat cagtgaggca cctatctcag cgatctgtct atttcgttca
     6781 tccatagttg cctgactccc cgtcgtgtag ataactacga tacgggaggg cttaccatct
     6841 ggccccagtg ctgcaatgat accgcgagac ccacgctcac cggctccaga tttatcagca
     6901 ataaaccagc cagccggaag ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc
     6961 atccagtcta ttaattgttg ccgggaagct agagtaagta gttcgccagt taatagtttg
     7021 cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct
     7081 tcattcagct ccggttccca acgatcaagg cgagttacat gatcccccat gttgtgcaaa
     7141 aaagcggtta gctccttcgg tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta
     7201 tcactcatgg ttatggcagc actgcataat tctcttactg tcatgccatc cgtaagatgc
     7261 ttttctgtga ctggtgagta ctcaaccaag tcattctgag aatagtgtat gcggcgaccg
     7321 agttgctctt gcccggcgtc aatacgggat aataccgcgc cacatagcag aactttaaaa
     7381 gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct caaggatctt accgctgttg
     7441 agatccagtt cgatgtaacc cactcgtgca cccaactgat cttcagcatc ttttactttc
     7501 accagcgttt ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg
     7561 gcgacacgga aatgttgaat actcatactc ttcctttttc aatattattg aagcatttat
     7621 cagggttatt gtctcatgag cggatacata tttgaatgta tttagaaaaa taaacaaata
     7681 ggggttccgc gcacatttcc ccgaaaagtg ccacctgacg tc
//
