LOCUS       sequence-47393-e        7066 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 7066)
  AUTHORS   Yue SB, Trujillo RD, Tang Y, O'Gorman WE, Chen CZ
  TITLE     Loop nucleotides control primary and mature miRNA function in target
            recognition and repression.
  JOURNAL   RNA Biol. 2011 Nov-Dec;8(6):1115-23.
  PUBMED    22142974
REFERENCE   2  (bases 1 to 7066)
  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: "RNA Biol. 
            2011 Nov-Dec"; volume: "8(6)"; pages: "1115-23"
COMMENT     Sequence Label: MXW-PGK-IRES-EGFP
FEATURES             Location/Qualifiers
     source          1..7066
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_feature    580..921
                     /label=MESV Psi
                     /note="packaging signal of murine embryonic stem cell 
                     virus"
     CDS             988..1404
                     /label=gag (truncated)
                     /note="truncated Moloney murine leukemia virus (MMLV) gag 
                     gene lacking the start codon"
     primer_bind     1334..1356
                     /label=pLXSN 5'
                     /note="Murine stem cell virus, forward primer. Also called 
                     MSCV"
     primer_bind     1372..1388
                     /label=pBABE 5'
                     /note="Psi packaging signal, forward primer for pBABE 
                     vectors"
     promoter        1422..1922
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     1855..1874
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     misc_feature    1967..2518
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(2111..2128)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     2338..2357
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             join(2530..2532,2533..2535,2536..3249)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(2575..2596)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2836..2855)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3183..3204
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     LTR             3341..3856
                     /label=3' LTR
                     /note="3' long terminal repeat from murine embryonic stem 
                     cell virus"
     primer_bind     complement(4025..4041)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4025..4041)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4038..4060)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4049..4065
                     /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(4073..4079,4080..4097,4098..4103))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4118..4139
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4256..4273)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4427..5015)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4507..4526)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5186..5977,5978..6046))
                     /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     5809..5828
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6047..6151)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6219..6237
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(6275..6297)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     6397..6416
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     6610..6632
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     6624..6641
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     6625..6641
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     6844..6863
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     LTR             join(7065..7066,1..516)
                     /label=5' LTR
                     /note="5' long terminal repeat from murine embryonic stem 
                     cell virus"
ORIGIN
        1 tgaaagaccc cacctgtagg tttggcaagc tagcttaagt aacgccattt tgcaaggcat
       61 ggaaaataca taactgagaa tagagaagtt cagatcaagg ttaggaacag agagacagca
      121 gaatatgggc caaacaggat atctgtggta agcagttcct gccccggctc agggccaaga
      181 acagatggtc cccagatgcg gtcccgccct cagcagtttc tagagaacca tcagatgttt
      241 ccagggtgcc ccaaggacct gaaaatgacc ctgtgcctta tttgaactaa ccaatcagtt
      301 cgcttctcgc ttctgttcgc gcgcttctgc tccccgagct caataaaaga gcccacaacc
      361 cctcactcgg cgcgccagtc ctccgataga ctgcgtcgcc cgggtacccg tattcccaat
      421 aaagcctctt gctgtttgca tccgaatcgt ggactcgctg atccttggga gggtctcctc
      481 agattgattg actgcccacc tcgggggtct ttcatttgga ggttccaccg agatttggag
      541 acccctgcct agggaccacc gacccccccg ccgggaggta agctggccag cggtcgtttc
      601 gtgtctgtct ctgtctttgt gcgtgtttgt gccggcatct aatgtttgcg cctgcgtctg
      661 tactagttag ctaactagct ctgtatctgg cggacccgtg gtggaactga cgagttctga
      721 acacccggcc gcaaccctgg gagacgtccc agggactttg ggggccgttt ttgtggcccg
      781 acctgaggaa gggagtcgat gtggaatccg accccgtcag gatatgtggt tctggtagga
      841 gacgagaacc taaaacagtt cccgcctccg tctgaatttt tgctttcggt ttggaaccga
      901 agccgcgcgt cttgtctgct gcagcgctgc agcatcgttc tgtgttgtct ctgtctgact
      961 gtgtttctgt atttgtctga aaattagggc cagactgtta ccactccctt aagtttgacc
     1021 ttaggtcact ggaaagatgt cgagcggatc gctcacaacc agtcggtaga tgtcaagaag
     1081 agacgttggg ttaccttctg ctctgcagaa tggccaacct ttaacgtcgg atggccgcga
     1141 gacggcacct ttaaccgaga cctcatcacc caggttaaga tcaaggtctt ttcacctggc
     1201 ccgcatggac acccagacca ggtcccctac atcgtgacct gggaagcctt ggcttttgac
     1261 ccccctccct gggtcaagcc ctttgtacac cctaagcctc cgcctcctct tcctccatcc
     1321 gccccgtctc tcccccttga acctcctcgt tcgaccccgc ctcgatcctc cctttatcca
     1381 gccctcactc cttctctagg cgccggaatt agatccctac cgggtagggg aggcgctttt
     1441 cccaaggcag tctggagcat gcgctttagc ggccccgctg ggcacttggc gctacacaag
     1501 tggcctctgg cctcgcacac attccacatc caccggtagg cgccaaccgg ctccgttctt
     1561 tggtggcccc ttcgcgccac cttctactcc tcccctagtc aggaagttcc cccccgcccc
     1621 gcagctcgcg tcgtgcagga cgtgacaaat ggaagtagca cgtctcacta gtctcgtgca
     1681 gatggacagc accgctgagc aatggaagcg ggtaggcctt tggggcagcg gccaatagca
     1741 gctttggctc cttcgctttc tgggctcaga ggctgggaag gggtgggtcc gggggcgggc
     1801 tcaggggcgg gctcaggggc ggggcgggcg cccgaaggtc ctccggaggc ccggcattct
     1861 gcacgcttca aaagcgcacg tctgccgcgc tgttctcctc ttcctcatct ccgggccttt
     1921 cgacctgcac tcgaggttaa cgaattccgc cccccccccc cccctaacgt tactggccga
     1981 agccgcttgg aataaggccg gtgtgcgttt gtctatatgt tattttccac catattgccg
     2041 tcttttggca atgtgagggc ccggaaacct ggccctgtct tcttgacgag cattcctagg
     2101 ggtctttccc ctctcgccaa aggaatgcaa ggtctgttga atgtcgtgaa ggaagcagtt
     2161 cctctggaag cttcttgaag acaaacaacg tctgtagcga ccctttgcag gcagcggaac
     2221 cccccacctg gcgacaggtg cctctgcggc caaaagccac gtgtataaga tacacctgca
     2281 aaggcggcac aaccccagtg ccacgttgtg agttggatag ttgtggaaag agtcaaatgg
     2341 ctctcctcaa gcgtattcaa caaggggctg aaggatgccc agaaggtacc ccattgtatg
     2401 ggatctgatc tggggcctcg gtgcacatgc tttacatgtg tttagtcgag gttaaaaaac
     2461 gtctaggccc cccgaaccac ggggacgtgg ttttcctttg aaaaacacga tgataatatg
     2521 gccacaacca tggtgagcaa gggcgaggag ctgttcaccg gggtggtgcc catcctggtc
     2581 gagctggacg gcgacgtaaa cggccacaag ttcagcgtgt ccggcgaggg cgagggcgat
     2641 gccacctacg gcaagctgac cctgaagttc atctgcacca ccggcaagct gcccgtgccc
     2701 tggcccaccc tcgtgaccac cttcacctac ggcgtgcagt gcttcagccg ctaccccgac
     2761 cacatgaagc agcacgactt cttcaagtcc gccatgcccg aaggctacgt ccaggagcgc
     2821 accatcttct tcaaggacga cggcaactac aagacccgcg ccgaggtgaa gttcgagggc
     2881 gacaccctgg tgaaccgcat cgagctgaag ggcatcgact tcaaggagga cggcaacatc
     2941 ctggggcaca agctggagta caactacaac agccacaacg tctatatcat ggccgacaag
     3001 cagaagaacg gcatcaaggt gaacttcaag atccgccaca acatcgagga cggcagcgtg
     3061 cagctcgccg accactacca gcagaacacc cccatcggcg acggccccgt gctgctgccc
     3121 gacaaccact acctgagcac ccagtccgcc ctgagcaaag accccaacga gaagcgcgat
     3181 cacatggtcc tgctggagtt cgtgaccgcc gccgggatca ctctcggcat ggacgagctg
     3241 tacaagtaat gaattaatta agaattgcgg ccgcgtcgac ctgcagccaa gcttatcgat
     3301 aaaataaaag attttattta gtctccagaa aaagggggga atgaaagacc ccacctgtag
     3361 gtttggcaag ctagcttaag taacgccatt ttgcaaggca tggaaaatac ataactgaga
     3421 atagagaagt tcagatcaag gttaggaaca gagagacagc agaatatggg ccaaacagga
     3481 tatctgtggt aagcagttcc tgccccggct cagggccaag aacagatggt ccccagatgc
     3541 ggtcccgccc tcagcagttt ctagagaacc atcagatgtt tccagggtgc cccaaggacc
     3601 tgaaaatgac cctgtgcctt atttgaacta accaatcagt tcgcttctcg cttctgttcg
     3661 cgcgcttctg ctccccgagc tcaataaaag agcccacaac ccctcactcg gcgcgccagt
     3721 cctccgatag actgcgtcgc ccgggtaccc gtgtatccaa taaaccctct tgcagttgca
     3781 tccgacttgt ggtctcgctg ttccttggga gggtctcctc tgagtgattg actacccgtc
     3841 agcgggggtc tttcatgggt aacagtttct tgaagttgga gaacaacatt ctgagggtag
     3901 gagtcgaata ttaagtaatc ctgactcaat tagccactgt tttgaatcca catactccaa
     3961 tactcctgaa atagttcatt atggacagcg cagaagagct ggggagaatt aattcgtaat
     4021 catggtcata gctgtttcct gtgtgaaatt gttatccgct cacaattcca cacaacatac
     4081 gagccggaag cataaagtgt aaagcctggg gtgcctaatg agtgagctaa ctcacattaa
     4141 ttgcgttgcg ctcactgccc gctttccagt cgggaaacct gtcgtgccag ctgcattaat
     4201 gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg gcgctcttcc gcttcctcgc
     4261 tcactgactc gctgcgctcg gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg
     4321 cggtaatacg gttatccaca gaatcagggg ataacgcagg aaagaacatg tgagcaaaag
     4381 gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc
     4441 gcccccctga cgagcatcac aaaaatcgac gctcaagtca gaggtggcga aacccgacag
     4501 gactataaag ataccaggcg tttccccctg gaagctccct cgtgcgctct cctgttccga
     4561 ccctgccgct taccggatac ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc
     4621 atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg
     4681 tgcacgaacc ccccgttcag cccgaccgct gcgccttatc cggtaactat cgtcttgagt
     4741 ccaacccggt aagacacgac ttatcgccac tggcagcagc cactggtaac aggattagca
     4801 gagcgaggta tgtaggcggt gctacagagt tcttgaagtg gtggcctaac tacggctaca
     4861 ctagaaggac agtatttggt atctgcgctc tgctgaagcc agttaccttc ggaaaaagag
     4921 ttggtagctc ttgatccggc aaacaaacca ccgctggtag cggtggtttt tttgtttgca
     4981 agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg
     5041 ggtctgacgc tcagtggaac gaaaactcac gttaagggat tttggtcatg agattatcaa
     5101 aaaggatctt cacctagatc cttttaaatt aaaaatgaag ttttaaatca atctaaagta
     5161 tatatgagta aacttggtct gacagttacc aatgcttaat cagtgaggca cctatctcag
     5221 cgatctgtct atttcgttca tccatagttg cctgactccc cgtcgtgtag ataactacga
     5281 tacgggaggg cttaccatct ggccccagtg ctgcaatgat accgcgagac ccacgctcac
     5341 cggctccaga tttatcagca ataaaccagc cagccggaag ggccgagcgc agaagtggtc
     5401 ctgcaacttt atccgcctcc atccagtcta ttaattgttg ccgggaagct agagtaagta
     5461 gttcgccagt taatagtttg cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac
     5521 gctcgtcgtt tggtatggct tcattcagct ccggttccca acgatcaagg cgagttacat
     5581 gatcccccat gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc gttgtcagaa
     5641 gtaagttggc cgcagtgtta tcactcatgg ttatggcagc actgcataat tctcttactg
     5701 tcatgccatc cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag tcattctgag
     5761 aatagtgtat gcggcgaccg agttgctctt gcccggcgtc aatacgggat aataccgcgc
     5821 cacatagcag aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct
     5881 caaggatctt accgctgttg agatccagtt cgatgtaacc cactcgtgca cccaactgat
     5941 cttcagcatc ttttactttc accagcgttt ctgggtgagc aaaaacagga aggcaaaatg
     6001 ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat actcatactc ttcctttttc
     6061 aatattattg aagcatttat cagggttatt gtctcatgag cggatacata tttgaatgta
     6121 tttagaaaaa taaacaaata ggggttccgc gcacatttcc ccgaaaagtg ccacctgacg
     6181 tctaagaaac cattattatc atgacattaa cctataaaaa taggcgtatc acgaggccct
     6241 ttcgtctcgc gcgtttcggt gatgacggtg aaaacctctg acacatgcag ctcccggaga
     6301 cggtcacagc ttgtctgtaa gcggatgccg ggagcagaca agcccgtcag ggcgcgtcag
     6361 cgggtgttgg cgggtgtcgg ggctggctta actatgcggc atcagagcag attgtactga
     6421 gagtgcacca tatgcggtgt gaaataccgc acagatgcgt aaggagaaaa taccgcatca
     6481 ggcgccattc gccattcagg ctgcgcaact gttgggaagg gcgatcggtg cgggcctctt
     6541 cgctattacg ccagctggcg aaagggggat gtgctgcaag gcgattaagt tgggtaacgc
     6601 cagggttttc ccagtcacga cgttgtaaaa cgacggccag tgccaagctc gctctccctt
     6661 atgcgactcc tgcattagga agcagcccag tagtaggttg aggccgttga gcaccgccgc
     6721 cgcaaggaat ggtgcatgca aggagatggc gcccaacagt cccccggcca cggggcctgc
     6781 caccataccc acgccgaaac aagcgctcat gagcccgaag tggcgagccc gatcttcccc
     6841 atcggtgatg tcggcgatat aggcgccagc aaccgcacct gtggcgccgg tgatgccggc
     6901 cacgatgcgt ccggcgtaga ggatcgatta gtccaatttg ttaaagacag gatatcagtg
     6961 gtccaggctc tagttttgac tcaacaatat caccagctga agcctataga gtacgagcca
     7021 tagataaaat aaaagatttt atttagtctc cagaaaaagg ggggaa
//
