LOCUS Exported 7934 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION Expresses miR-22 in mammalian cells. ACCESSION . VERSION . KEYWORDS pMSCV-PIG-miR-22 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 7934) AUTHORS Chang TC, Yu D, Lee YS, Wentzel EA, Arking DE, West KM, Dang CV, Thomas-Tikhonenko A, Mendell JT TITLE Widespread microRNA repression by Myc contributes to tumorigenesis. JOURNAL Nat Genet. 2008 Jan;40(1):43-50. Epub 2007 Dec 9. PUBMED 18066065 REFERENCE 2 (bases 1 to 7934) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..7934 /organism="synthetic DNA construct" /mol_type="other DNA" 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" promoter 1715..2214 /label=PGK promoter /note="mouse phosphoglycerate kinase 1 promoter" primer_bind complement(1750..1773) /label=MSCV-rev /note="Murine stem cell virus, reverse primer" primer_bind 2147..2166 /label=mPGK-F /note="Mouse PGK promoter, forward primer" CDS 2235..2834 /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(2235..2254) /label=Puro-R /note="Puromycin resistance gene, reverse primer. Also called puro-variant-R" primer_bind 2731..2751 /label=Puro-F /note="Puromycin resistance gene, forward primer" misc_feature 2912..3463 /label=IRES /note="internal ribosome entry site (IRES) of the encephalomyocarditis virus (EMCV)" primer_bind complement(3056..3073) /label=IRES reverse /note="IRES internal ribosome entry site, reverse primer. Also called pCDH-rev" primer_bind 3283..3302 /label=IRES-F /note="IRES internal ribosome entry site, forward primer" CDS 3475..4194 /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 3475..4191 /codon_start=1 /product="enhanced GFP" /label=EGFP /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL EFVTAAGITLGMDELYK" primer_bind complement(3520..3541) /label=EGFP-N /note="EGFP, reverse primer" primer_bind complement(3781..3800) /label=EXFP-R /note="For distinguishing EGFP variants, reverse primer" primer_bind 4128..4149 /label=EGFP-C /note="EGFP, forward primer" LTR 4296..4810 /label=3' LTR /note="3' long terminal repeat from murine embryonic stem cell virus" primer_bind complement(4979..4995) /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind complement(4979..4995) /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" primer_bind complement(4992..5014) /label=M13/pUC Reverse /note="In lacZ gene" protein_bind 5003..5019 /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(5027..5057) /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 5072..5093 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." primer_bind complement(5210..5227) /label=L4440 /note="L4440 vector, forward primer" rep_origin complement(5381..5969) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(5461..5480) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS complement(6140..7000) /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 6763..6782 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(7001..7105) /gene="bla" /label=AmpR promoter primer_bind 7173..7191 /label=pBRforEco /note="pBR322 vectors, upsteam of EcoRI site, forward primer" primer_bind complement(7229..7251) /label=pGEX 3' /note="pGEX vectors, reverse primer" primer_bind 7351..7370 /label=pRS-marker /note="pRS vectors, use to sequence yeast selectable marker" primer_bind 7564..7586 /label=M13/pUC Forward /note="In lacZ gene" primer_bind 7714..7733 /label=pBRrevBam /note="pBR322 vectors, tet region, downstream of BamHI, reverse primer" LTR 7933..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 gatctctcga gcatgccctg ctcagatctt 1441 tcccattttc cctccctttc ccttaggagc ctgttcctct cacgccctca cctggctgag 1501 ccgcagtagt tcttcagtgg caagctttat gtcctgaccc agctaaagct gccagttgaa 1561 gaactgttgc cctctgcccc tggcttcgag gaggaggagg agctgctttc cccatcatct 1621 ggaaggtgac agaaatgggc tgggaaggtc cgaacagcag ggtggatgat acgttttggg 1681 caagttggag agctcgaggt taacgaattc taccgggtag gggaggcgct tttcccaagg 1741 cagtctggag catgcgcttt agcagccccg ctgggcactt ggcgctacac aagtggcctc 1801 tggcctcgca cacattccac atccaccggt aggcgccaac cggctccgtt ctttggtggc 1861 cccttcgcgc caccttctac tcctccccta gtcaggaagt tcccccccgc cccgcagctc 1921 gcgtcgtgca ggacgtgaca aatggaagta gcacgtctca ctagtctcgt gcagatggac 1981 agcaccgctg agcaatggaa gcgggtaggc ctttggggca gcggccaata gcagctttgc 2041 tccttcgctt tctgggctca gaggctggga aggggtgggt ccgggggcgg gctcaggggc 2101 gggctcaggg gcggggcggg cgcccgaagg tcctccggag gcccggcatt ctgcacgctt 2161 caaaagcgca cgtctgccgc gctgttctcc tcttcctcat ctccgggcct ttcgacctgc 2221 agcccaagct taccatgacc gagtacaagc ccacggtgcg cctcgccacc cgcgacgacg 2281 tccccagggc cgtacgcacc ctcgccgccg cgttcgccga ctaccccgcc acgcgccaca 2341 ccgtcgatcc ggaccgccac atcgagcggg tcaccgagct gcaagaactc ttcctcacgc 2401 gcgtcgggct cgacatcggc aaggtgtggg tcgcggacga cggcgccgcg gtggcggtct 2461 ggaccacgcc ggagagcgtc gaagcggggg cggtgttcgc cgagatcggc ccgcgcatgg 2521 ccgagttgag cggttcccgg ctggccgcgc agcaacagat ggaaggcctc ctggcgccgc 2581 accggcccaa ggagcccgcg tggttcctgg ccaccgtcgg cgtctcgccc gaccaccagg 2641 gcaagggtct gggcagcgcc gtcgtgctcc ccggagtgga ggcggccgag cgcgccgggg 2701 tgcccgcctt cctggagacc tccgcgcccc gcaacctccc cttctacgag cggctcggct 2761 tcaccgtcac cgccgacgtc gaggtgcccg aaggaccgcg cacctggtgc atgacccgca 2821 agcccggtgc ctgacgcccg ccccacgacc cgcagcgccc gaccgaaagg agcgcacgac 2881 cccatgcatc gaattccgcc ccccccccct aacgttactg gccgaagccg cttggaataa 2941 ggccggtgtg cgtttgtcta tatgttattt tccaccatat tgccgtcttt tggcaatgtg 3001 agggcccgga aacctggccc tgtcttcttg acgagcattc ctaggggtct ttcccctctc 3061 gccaaaggaa tgcaaggtct gttgaatgtc gtgaaggaag cagttcctct ggaagcttct 3121 tgaagacaaa caacgtctgt agcgaccctt tgcaggcagc ggaacccccc acctggcgac 3181 aggtgcctct gcggccaaaa gccacgtgta taagatacac ctgcaaaggc ggcacaaccc 3241 cagtgccacg ttgtgagttg gatagttgtg gaaagagtca aatggctctc ctcaagcgta 3301 ttcaacaagg ggctgaagga tgcccagaag gtaccccatt gtatgggatc tgatctgggg 3361 cctcggtgca catgctttac atgtgtttag tcgaggttaa aaaacgtcta ggccccccga 3421 accacgggga cgtggttttc ctttgaaaaa cacgatgata atatggccac aaccatggtg 3481 agcaagggcg aggagctgtt caccggggtg gtgcccatcc tggtcgagct ggacggcgac 3541 gtaaacggcc acaagttcag cgtgtccggc gagggcgagg gcgatgccac ctacggcaag 3601 ctgaccctga agttcatctg caccaccggc aagctgcccg tgccctggcc caccctcgtg 3661 accaccctga cctacggcgt gcagtgcttc agccgctacc ccgaccacat gaagcagcac 3721 gacttcttca agtccgccat gcccgaaggc tacgtccagg agcgcaccat cttcttcaag 3781 gacgacggca actacaagac ccgcgccgag gtgaagttcg agggcgacac cctggtgaac 3841 cgcatcgagc tgaagggcat cgacttcaag gaggacggca acatcctggg gcacaagctg 3901 gagtacaact acaacagcca caacgtctat atcatggccg acaagcagaa gaacggcatc 3961 aaggtgaact tcaagatccg ccacaacatc gaggacggca gcgtgcagct cgccgaccac 4021 taccagcaga acacccccat cggcgacggc cccgtgctgc tgcccgacaa ccactacctg 4081 agcacccagt ccgccctgag caaagacccc aacgagaagc gcgatcacat ggtcctgctg 4141 gagttcgtga ccgccgccgg gatcactctc ggcatggacg agctgtacaa gtaatgaatt 4201 aattaagaat tatcaagctt atcgataccg tcgacctgca gccaagctta tcgataaaat 4261 aaaagatttt atttagtctc cagaaaaagg ggggaatgaa agaccccacc tgtaggtttg 4321 gcaagctagc ttaagtaacg ccattttgca aggcatggaa aatacataac tgagaataga 4381 gaagttcaga tcaaggttag gaacagagag acagcagaat atgggccaaa caggatatct 4441 gtggtaagca gttcctgccc cggctcaggg ccaagaacag atggtcccca gatgcggtcc 4501 cgccctcagc agtttctaga gaaccatcag atgtttccag ggtgccccaa ggacctgaaa 4561 tgaccctgtg ccttatttga actaaccaat cagttcgctt ctcgcttctg ttcgcgcgct 4621 tctgctcccc gagctcaata aaagagccca caacccctca ctcggcgcgc cagtcctccg 4681 atagactgcg tcgcccgggt acccgtgtat ccaataaacc ctcttgcagt tgcatccgac 4741 ttgtggtctc gctgttcctt gggagggtct cctctgagtg attgactacc cgtcagcggg 4801 ggtctttcat gggtaacagt ttcttgaagt tggagaacaa cattctgagg gtaggagtcg 4861 aatattaagt aatcctgact caattagcca ctgttttgaa tccacatact ccaatactcc 4921 tgaaatagtt cattatggac agcgcagaag agctggggag aattaattcg taatcatggt 4981 catagctgtt tcctgtgtga aattgttatc cgctcacaat tccacacaac atacgagccg 5041 gaagcataaa gtgtaaagcc tggggtgcct aatgagtgag ctaactcaca ttaattgcgt 5101 tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat taatgaatcg 5161 gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc tcgctcactg 5221 actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc agctcactca aaggcggtaa 5281 tacggttatc cacagaatca ggggataacg caggaaagaa catgtgagca aaaggccagc 5341 aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg ctccgccccc 5401 ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg acaggactat 5461 aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc 5521 cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt tctcatagct 5581 cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg 5641 aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc 5701 cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt agcagagcga 5761 ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc tacactagaa 5821 ggacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa agagttggta 5881 gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt tgcaagcagc 5941 agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct acggggtctg 6001 acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgagatta tcaaaaagga 6061 tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa agtatatatg 6121 agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc tcagcgatct 6181 gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact acgatacggg 6241 agggcttacc atctggcccc agtgctgcaa tgataccgcg agacccacgc tcaccggctc 6301 cagatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt ggtcctgcaa 6361 ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta agtagttcgc 6421 cagttaatag tttgcgcaac gttgttgcca ttgctacagg catcgtggtg tcacgctcgt 6481 cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt acatgatccc 6541 ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc agaagtaagt 6601 tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt actgtcatgc 6661 catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc tgagaatagt 6721 gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg ggataatacc gcgccacata 6781 gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa ctctcaagga 6841 tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac tgatcttcag 6901 catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa 6961 aaaagggaat aagggcgaca cggaaatgtt gaatactcat actcttcctt tttcaatatt 7021 attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa tgtatttaga 7081 aaaataaaca aataggggtt ccgcgcacat ttccccgaaa agtgccacct gacgtctaag 7141 aaaccattat tatcatgaca ttaacctata aaaataggcg tatcacgagg ccctttcgtc 7201 tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat gcagctcccg gagacggtca 7261 cagcttgtct gtaagcggat gccgggagca gacaagcccg tcagggcgcg tcagcgggtg 7321 ttggcgggtg tcggggctgg cttaactatg cggcatcaga gcagattgta ctgagagtgc 7381 accatatgcg gtgtgaaata ccgcacagat gcgtaaggag aaaataccgc atcaggcgcc 7441 attcgccatt caggctgcgc aactgttggg aagggcgatc ggtgcgggcc tcttcgctat 7501 tacgccagct ggcgaaaggg ggatgtgctg caaggcgatt aagttgggta acgccagggt 7561 tttcccagtc acgacgttgt aaaacgacgg cgcaaggaat ggtgcatgca aggagatggc 7621 gcccaacagt cccccggcca cggggcctgc caccataccc acgccgaaac aagcgctcat 7681 gagcccgaag tggcgagccc gatcttcccc atcggtgatg tcggcgatat aggcgccagc 7741 aaccgcacct gtggcgccgg tgatgccggc cacgatgcgt ccggcgtaga ggcgattagt 7801 ccaatttgtt aaagacagga tatcagtggt ccaggctcta gttttgactc aacaatatca 7861 ccagctgaag cctatagagt acgagccata gataaaataa aagattttat ttagtctcca 7921 gaaaaagggg ggaa //