LOCUS       sequence-41583-e        7538 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 7538)
  AUTHORS   Premsrirut PK, Dow LE, Kim SY, Camiolo M, Malone CD, Miething C, 
            Scuoppo C, Zuber J, Dickins RA, Kogan SC, Shroyer KR, Sordella R, 
            Hannon GJ, Lowe SW
  TITLE     A rapid and scalable system for studying gene function in mice using
            conditional RNA interference.
  JOURNAL   Cell. 2011 Apr 1;145(1):145-58.
  PUBMED    21458673
REFERENCE   2  (bases 1 to 7538)
  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: "Cell."; 
            date: "2011-04-1"; volume: "145(1)"; pages: "145-58"
COMMENT     Sequence Label: pTGMP
FEATURES             Location/Qualifiers
     source          1..7538
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_feature    579..920
                     /label=MESV Psi
                     /note="packaging signal of murine embryonic stem cell 
                     virus"
     CDS             987..1403
                     /label=gag (truncated)
                     /note="truncated Moloney murine leukemia virus (MMLV) gag 
                     gene lacking the start codon"
     primer_bind     1333..1355
                     /label=pLXSN 5'
                     /note="Murine stem cell virus, forward primer. Also called 
                     MSCV"
     primer_bind     1371..1387
                     /label=pBABE 5'
                     /note="Psi packaging signal, forward primer for pBABE 
                     vectors"
     protein_bind    join(1451..1469,1470..1492,1493..1511,1512..1534,1535..1553,
                     1554..1576,1577..1595,1596..1618,1619..1637,1638..1660,
                     1661..1679,1680..1702,1703..1721)
                     /label=tetracycline response element
                     /bound_moiety="tetracycline repressor TetR"
                     /note="contains seven copies of the tetracycline operator 
                     tetO"
     promoter        1754..1791
                     /label=minimal CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     1788..1812
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             join(1885..1887,1888..1890,1891..2604)
                     /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(1885..1887,1888..1890,1891..2601)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1930..1951)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2191..2210)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2538..2559
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     ncRNA           2639..2732
                     /label=5' miR-30a
                     /note="sequence upstream of the 71-nt precursor of the 
                     human miR-30a microRNA (Zeng et al., 2002)"
                     /ncRNA_class="miRNA"
     ncRNA           2746..2858
                     /label=3' miR-30a
                     /note="sequence downstream of the 71-nt precursor of the 
                     human miR-30a microRNA (Zeng et al., 2002)"
                     /ncRNA_class="miRNA"
     promoter        2872..3371
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     complement(2907..2930)
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     primer_bind     3304..3323
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     CDS             3392..3991
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(3392..3411)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     3888..3908
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     primer_bind     complement(4583..4599)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4583..4599)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4596..4618)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4607..4623
                     /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(4631..4637,4638..4655,4656..4661))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4676..4697
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4814..4831)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4985..5573)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5065..5084)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5744..6535,6536..6604))
                     /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     6367..6386
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6605..6709)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6777..6795
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(6833..6855)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     6955..6974
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     7168..7190
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     7318..7337
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     LTR             join(7537..7538,1..515)
                     /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 gaaatgaccc tgtgccttat ttgaactaac caatcagttc
      301 gcttctcgct tctgttcgcg cgcttctgct ccccgagctc aataaaagag cccacaaccc
      361 ctcactcggc gcgccagtcc tccgatagac tgcgtcgccc gggtacccgt attcccaata
      421 aagcctcttg ctgtttgcat ccgaatcgtg gactcgctga tccttgggag ggtctcctca
      481 gattgattga ctgcccacct cgggggtctt tcatttggag gttccaccga gatttggaga
      541 cccctgccca gggaccaccg acccccccgc cgggaggtaa gctggccagc ggtcgtttcg
      601 tgtctgtctc tgtctttgtg cgtgtttgtg ccggcatcta atgtttgcgc ctgcgtctgt
      661 actagttagc taactagctc tgtatctggc ggacccgtgg tggaactgac gagttctgaa
      721 cacccggccg caaccctggg agacgtccca gggactttgg gggccgtttt tgtggcccga
      781 cctgaggaag ggagtcgatg tggaatccga ccccgtcagg atatgtggtt ctggtaggag
      841 acgagaacct aaaacagttc ccgcctccgt ctgaattttt gctttcggtt tggaaccgaa
      901 gccgcgcgtc ttgtctgctg cagcgctgca gcatcgttct gtgttgtctc tgtctgactg
      961 tgtttctgta tttgtctgaa aattagggcc agactgttac cactccctta agtttgacct
     1021 taggtcactg gaaagatgtc gagcggatcg ctcacaacca gtcggtagat gtcaagaaga
     1081 gacgttgggt taccttctgc tctgcagaat ggccaacctt taacgtcgga tggccgcgag
     1141 acggcacctt taaccgagac ctcatcaccc aggttaagat caaggtcttt tcacctggcc
     1201 cgcatggaca cccagaccag gtcccctaca tcgtgacctg ggaagccttg gcttttgacc
     1261 cccctccctg ggtcaagccc tttgtacacc ctaagcctcc gcctcctctt cctccatccg
     1321 ccccgtctct cccccttgaa cctcctcgtt cgaccccgcc tcgatcctcc ctttatccag
     1381 ccctcactcc ttctctaggc gccggaatta gatctggggg atcgatcctc tagagcatcg
     1441 agtttaccac tccctatcag tgatagagaa aagtgaaagt cgagtttacc actccctatc
     1501 agtgatagag aaaagtgaaa gtcgagttta ccactcccta tcagtgatag agaaaagtga
     1561 aagtcgagtt taccactccc tatcagtgat agagaaaagt gaaagtcgag tttaccactc
     1621 cctatcagtg atagagaaaa gtgaaagtcg agtttaccac tccctatcag tgatagagaa
     1681 aagtgaaagt cgagtttacc actccctatc agtgatagag aaaagtgaaa gtcgagctcg
     1741 gtacccgggt cgagtaggcg tgtacggtgg gaggcctata taagcagagc tcgtttagtg
     1801 aaccgtcaga tcgcctggag acgccatcca cgctgttttg acctccatag aagacaccgg
     1861 gaccgatcca gcctctcgac caccatggtg agcaagggcg aggagctgtt caccggggtg
     1921 gtgcccatcc tggtcgagct ggacggcgac gtaaacggcc acaagttcag cgtgtccggc
     1981 gagggcgagg gcgatgccac ctacggcaag ctgaccctga agttcatctg caccaccggc
     2041 aagctgcccg tgccctggcc caccctcgtg accaccctga cctacggcgt gcagtgcttc
     2101 agccgctacc ccgaccacat gaagcagcac gacttcttca agtccgccat gcccgaaggc
     2161 tacgtccagg agcgcaccat cttcttcaag gacgacggca actacaagac ccgcgccgag
     2221 gtgaagttcg agggcgacac cctggtgaac cgcatcgagc tgaagggcat cgacttcaag
     2281 gaggacggca acatcctggg gcacaagctg gagtacaact acaacagcca caacgtctat
     2341 atcatggccg acaagcagaa gaacggcatc aaggtgaact tcaagatccg ccacaacatc
     2401 gaggacggca gcgtgcagct cgccgaccac taccagcaga acacccccat cggcgacggc
     2461 cccgtgctgc tgcccgacaa ccactacctg agcacccagt ccgccctgag caaagacccc
     2521 aacgagaagc gcgatcacat ggtcctgctg gagttcgtga ccgccgccgg gatcactctc
     2581 ggcatggacg agctgtacaa gtaatgaacc ggtcgactag ggataacagg gtaattgttt
     2641 gaatgaggct tcagtacttt acagaatcgt tgcctgcaca tcttggaaac acttgctggg
     2701 attacttctt caggttaacc caacagaagg ctcgagcaac cagaattcaa ggggctactt
     2761 taggagcaat tatcttgttt actaaaactg aataccttgc tatctctttg atacattttt
     2821 acaaagctga attaaaatgg tataaattaa atcacttttt tcaattctac cgggtagggg
     2881 aggcgctttt cccaaggcag tctggagcat gcgctttagc agccccgctg ggcacttggc
     2941 gctacacaag tggcctctgg cctcgcacac attccacatc caccggtagg cgccaaccgg
     3001 ctccgttctt tggtggcccc ttcgcgccac cttctactcc tcccctagtc aggaagttcc
     3061 cccccgcccc gcagctcgcg tcgtgcagga cgtgacaaat ggaagtagca cgtctcacta
     3121 gtctcgtgca gatggacagc accgctgagc aatggaagcg ggtaggcctt tggggcagcg
     3181 gccaatagca gctttgctcc ttcgctttct gggctcagag gctgggaagg ggtgggtccg
     3241 ggggcgggct caggggcggg ctcaggggcg gggcgggcgc ccgaaggtcc tccggaggcc
     3301 cggcattctg cacgcttcaa aagcgcacgt ctgccgcgct gttctcctct tcctcatctc
     3361 cgggcctttc gacctgcagc ccaagcttac catgaccgag tacaagccca cggtgcgcct
     3421 cgccacccgc gacgacgtcc ccagggccgt acgcaccctc gccgccgcgt tcgccgacta
     3481 ccccgccacg cgccacaccg tcgatccgga ccgccacatc gagcgggtca ccgagctgca
     3541 agaactcttc ctcacgcgcg tcgggctcga catcggcaag gtgtgggtcg cggacgacgg
     3601 cgccgcggtg gcggtctgga ccacgccgga gagcgtcgaa gcgggggcgg tgttcgccga
     3661 gatcggcccg cgcatggccg agttgagcgg ttcccggctg gccgcgcagc aacagatgga
     3721 aggcctcctg gcgccgcacc ggcccaagga gcccgcgtgg ttcctggcca ccgtcggcgt
     3781 ctcgcccgac caccagggca agggtctggg cagcgccgtc gtgctccccg gagtggaggc
     3841 ggccgagcgc gccggggtgc ccgccttcct ggagacctcc gcgccccgca acctcccctt
     3901 ctacgagcgg ctcggcttca ccgtcaccgc cgacgtcgag gtgcccgaag gaccgcgcac
     3961 ctggtgcatg acccgcaagc ccggtgcctg acgcccgccc cacgacccgc agcgcccgac
     4021 cgaaaggagc gcacgacccc atgcatcgat aaaataaaag attttattta gtctccagaa
     4081 aaagggggga atgaaagacc ccacctgtag gtttggcaac tagagaacca tcagatgttt
     4141 ccagggtgcc ccaaggacct gaaatgaccc tgtgccttat ttgaactaac caatcagttc
     4201 gcttctcgct tctgttcgcg cgcttctgct ccccgagctc aataaaagag cccacaaccc
     4261 ctcactcggc gcgccagtcc tccgatagac tgcgtcgccc gggtacccgt gtatccaata
     4321 aaccctcttg cagttgcatc cgacttgtgg tctcgctgtt ccttgggagg gtctcctctg
     4381 agtgattgac tacccgtcag cgggggtctt tcatgggtaa cagtttcttg aagttggaga
     4441 acaacattct gagggtagga gtcgaatatt aagtaatcct gactcaatta gccactgttt
     4501 tgaatccaca tactccaata ctcctgaaat agttcattat ggacagcgca gaagagctgg
     4561 ggagaattaa ttcgtaatca tggtcatagc tgtttcctgt gtgaaattgt tatccgctca
     4621 caattccaca caacatacga gccggaagca taaagtgtaa agcctggggt gcctaatgag
     4681 tgagctaact cacattaatt gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt
     4741 cgtgccagct gcattaatga atcggccaac gcgcggggag aggcggtttg cgtattgggc
     4801 gctcttccgc ttcctcgctc actgactcgc tgcgctcggt cgttcggctg cggcgagcgg
     4861 tatcagctca ctcaaaggcg gtaatacggt tatccacaga atcaggggat aacgcaggaa
     4921 agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg
     4981 cgtttttcca taggctccgc ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga
     5041 ggtggcgaaa cccgacagga ctataaagat accaggcgtt tccccctgga agctccctcg
     5101 tgcgctctcc tgttccgacc ctgccgctta ccggatacct gtccgccttt ctcccttcgg
     5161 gaagcgtggc gctttctcat agctcacgct gtaggtatct cagttcggtg taggtcgttc
     5221 gctccaagct gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc gccttatccg
     5281 gtaactatcg tcttgagtcc aacccggtaa gacacgactt atcgccactg gcagcagcca
     5341 ctggtaacag gattagcaga gcgaggtatg taggcggtgc tacagagttc ttgaagtggt
     5401 ggcctaacta cggctacact agaaggacag tatttggtat ctgcgctctg ctgaagccag
     5461 ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa acaaaccacc gctggtagcg
     5521 gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct caagaagatc
     5581 ctttgatctt ttctacgggg tctgacgctc agtggaacga aaactcacgt taagggattt
     5641 tggtcatgag attatcaaaa aggatcttca cctagatcct tttaaattaa aaatgaagtt
     5701 ttaaatcaat ctaaagtata tatgagtaaa cttggtctga cagttaccaa tgcttaatca
     5761 gtgaggcacc tatctcagcg atctgtctat ttcgttcatc catagttgcc tgactccccg
     5821 tcgtgtagat aactacgata cgggagggct taccatctgg ccccagtgct gcaatgatac
     5881 cgcgagaccc acgctcaccg gctccagatt tatcagcaat aaaccagcca gccggaaggg
     5941 ccgagcgcag aagtggtcct gcaactttat ccgcctccat ccagtctatt aattgttgcc
     6001 gggaagctag agtaagtagt tcgccagtta atagtttgcg caacgttgtt gccattgcta
     6061 caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc ggttcccaac
     6121 gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc
     6181 ctccgatcgt tgtcagaagt aagttggccg cagtgttatc actcatggtt atggcagcac
     6241 tgcataattc tcttactgtc atgccatccg taagatgctt ttctgtgact ggtgagtact
     6301 caaccaagtc attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa
     6361 tacgggataa taccgcgcca catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt
     6421 cttcggggcg aaaactctca aggatcttac cgctgttgag atccagttcg atgtaaccca
     6481 ctcgtgcacc caactgatct tcagcatctt ttactttcac cagcgtttct gggtgagcaa
     6541 aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa tgttgaatac
     6601 tcatactctt cctttttcaa tattattgaa gcatttatca gggttattgt ctcatgagcg
     6661 gatacatatt tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc acatttcccc
     6721 gaaaagtgcc acctgacgtc taagaaacca ttattatcat gacattaacc tataaaaata
     6781 ggcgtatcac gaggcccttt cgtctcgcgc gtttcggtga tgacggtgaa aacctctgac
     6841 acatgcagct cccggagacg gtcacagctt gtctgtaagc ggatgccggg agcagacaag
     6901 cccgtcaggg cgcgtcagcg ggtgttggcg ggtgtcgggg ctggcttaac tatgcggcat
     6961 cagagcagat tgtactgaga gtgcaccata tgcggtgtga aataccgcac agatgcgtaa
     7021 ggagaaaata ccgcatcagg cgccattcgc cattcaggct gcgcaactgt tgggaagggc
     7081 gatcggtgcg ggcctcttcg ctattacgcc agctggcgaa agggggatgt gctgcaaggc
     7141 gattaagttg ggtaacgcca gggttttccc agtcacgacg ttgtaaaacg acggcgcaag
     7201 gaatggtgca tgcaaggaga tggcgcccaa cagtcccccg gccacggggc ctgccaccat
     7261 acccacgccg aaacaagcgc tcatgagccc gaagtggcga gcccgatctt ccccatcggt
     7321 gatgtcggcg atataggcgc cagcaaccgc acctgtggcg ccggtgatgc cggccacgat
     7381 gcgtccggcg tagaggcgat tagtccaatt tgttaaagac aggatatcag tggtccaggc
     7441 tctagttttg actcaacaat atcaccagct gaagcctata gagtacgagc catagataaa
     7501 ataaaagatt ttatttagtc tccagaaaaa ggggggaa
//
