LOCUS Exported 6963 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION Retroviral construct for miRNA expression. ACCESSION . VERSION . KEYWORDS MDH1-PGK-GFP_2.0 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 6963) AUTHORS Chen CZ, Li L, Lodish HF, Bartel DP TITLE MicroRNAs modulate hematopoietic lineage differentiation. JOURNAL Science. 2004 Jan 2. 303(5654):83-6. PUBMED 14657504 REFERENCE 2 (bases 1 to 6963) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..6963 /organism="synthetic DNA construct" /mol_type="other DNA" 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" primer_bind 1716..1732 /label=pBMN 5' /note="MMLV sequence, for inserts in pBMN retroviral vector" promoter 1796..2296 /label=PGK promoter /note="mouse phosphoglycerate kinase 1 promoter" primer_bind 2229..2248 /label=mPGK-F /note="Mouse PGK promoter, forward primer" CDS 2306..3025 /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(2351..2372) /label=EGFP-N /note="EGFP, reverse primer" primer_bind complement(2612..2631) /label=EXFP-R /note="For distinguishing EGFP variants, reverse primer" primer_bind 2959..2980 /label=EGFP-C /note="EGFP, forward primer" promoter complement(3168..3258) /label=H1 promoter /note="human H1 RNA promoter" primer_bind complement(3228..3247) /label=H1 /note="Human H1 promoter, forward primer" primer_bind complement(3922..3938) /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind complement(3922..3938) /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" primer_bind complement(3935..3957) /label=M13/pUC Reverse /note="In lacZ gene" protein_bind 3946..3962 /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(3970..4000) /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 4015..4036 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." primer_bind complement(4153..4170) /label=L4440 /note="L4440 vector, forward primer" rep_origin complement(4324..4912) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(4404..4423) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS complement(5083..5943) /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 5706..5725 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(5944..6048) /gene="bla" /label=AmpR promoter primer_bind 6116..6134 /label=pBRforEco /note="pBR322 vectors, upsteam of EcoRI site, forward primer" primer_bind complement(6172..6194) /label=pGEX 3' /note="pGEX vectors, reverse primer" primer_bind 6294..6313 /label=pRS-marker /note="pRS vectors, use to sequence yeast selectable marker" primer_bind 6507..6529 /label=M13/pUC Forward /note="In lacZ gene" primer_bind 6521..6538 /label=M13 Forward /note="In lacZ gene. Also called M13-F20 or M13 (-21) Forward" primer_bind 6522..6538 /label=M13 fwd /note="common sequencing primer, one of multiple similar variants" primer_bind 6741..6760 /label=pBRrevBam /note="pBR322 vectors, tet region, downstream of BamHI, reverse primer" LTR 6962..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 agatcttata tggggcaccc ccgccccttg 1441 taaacttccc tgaccctgac atgacaagag ttactaacag cccctctctc caagctcact 1501 tacaggctct ctacttagtc cagcacgaag tctggagacc tctggcggca gcctaccaag 1561 aacaactgga ccgaccggtg gtacctcacc cttaccgagt cggcgacaca gtgtgggtcc 1621 gccgacacca gactaagaac ctagaacctc gctggaaagg accttacaca gtcctgctga 1681 ccacccccac cgccctcaaa gtagacggca tcgcagcttg gatacacgcc gcccacgtga 1741 aggctgccga ccccgggggt ggaccatcct ctagactgcc ggatcaattc ctaccgggta 1801 ggggaggcgc ttttcccaag gcagtctgga gcatgcgctt tagcggcccc gctgggcact 1861 tggcgctaca caagtggcct ctggcctcgc acacattcca catccaccgg taggcgccaa 1921 ccggctccgt tctttggtgg ccccttcgcg ccaccttcta ctcctcccct agtcaggaag 1981 ttcccccccg ccccgcagct cgcgtcgtgc aggacgtgac aaatggaagt agcacgtctc 2041 actagtctcg tgcagatgga cagcaccgct gagcaatgga agcgggtagg cctttggggc 2101 agcggccaat agcagctttg gctccttcgc tttctgggct cagaggctgg gaaggggtgg 2161 gtccgggggc gggctcaggg gcgggctcag gggcggggcg ggcgcccgaa ggtcctccgg 2221 aggcccggca ttctgcacgc ttcaaaagcg cacgtctgcc gcgctgttct cctcttcctc 2281 atctccgggc ctttcgacct gcaccatggt gagcaagggc gaggagctgt tcaccggggt 2341 ggtgcccatc ctggtcgagc tggacggcga cgtaaacggc cacaagttca gcgtgtccgg 2401 cgagggcgag ggcgatgcca cctacggcaa gctgaccctg aagttcatct gcaccaccgg 2461 caagctgccc gtgccctggc ccaccctcgt gaccaccttc acctacggcg tgcagtgctt 2521 cagccgctac cccgaccaca tgaagcagca cgacttcttc aagtccgcca tgcccgaagg 2581 ctacgtccag gagcgcacca tcttcttcaa ggacgacggc aactacaaga cccgcgccga 2641 ggtgaagttc gagggcgaca ccctggtgaa ccgcatcgag ctgaagggca tcgacttcaa 2701 ggaggacggc aacatcctgg ggcacaagct ggagtacaac tacaacagcc acaacgtcta 2761 tatcatggcc gacaagcaga agaacggcat caaggtgaac ttcaagatcc gccacaacat 2821 cgaggacggc agcgtgcagc tcgccgacca ctaccagcag aacaccccca tcggcgacgg 2881 ccccgtgctg ctgcccgaca accactacct gagcacccag tccgccctga gcaaagaccc 2941 caacgagaag cgcgatcaca tggtcctgct ggagttcgtg accgccgccg ggatcactct 3001 cggcatggac gagctgtaca agtaatgaat taattaagaa ttgcggccgc gtcgacctgc 3061 agccaagctt atcgataaaa taaaagattt tatttagtct ccagaaaaag gggggaatga 3121 aagaccccac ctgtaggttt ggcaagaatt cgtttaaacg ggctcgagtg gtctcataca 3181 gaacttataa gattcccaaa tccaaagaca tttcacgttt atggtgattt cccagaacac 3241 atagcgacat gcaaatattg ggatccgcta gcttaagtaa cgccattttg caaggcatgg 3301 aaaatacata actgagaata gagaagttca gatcaaggtt aggaacagag agacagcaga 3361 atatgggcca aacaggatat ctgtggtaag cagttcctgc cccggctcag ggccaagaac 3421 agatggtccc cagatgcggt cccgccctca gcagtttcta gagaaccatc agatgtttcc 3481 agggtgcccc aaggacctga aaatgaccct gtgccttatt tgaactaacc aatcagttcg 3541 cttctcgctt ctgttcgcgc gcttctgctc cccgagctca ataaaagagc ccacaacccc 3601 tcactcggcg cgccagtcct ccgatagact gcgtcgcccg ggtacccgtg tatccaataa 3661 accctcttgc agttgcatcc gacttgtggt ctcgctgttc cttgggaggg tctcctctga 3721 gtgattgact acccgtcagc gggggtcttt catgggtaac agtttcttga agttggagaa 3781 caacattctg agggtaggag tcgaatatta agtaatcctg actcaattag ccactgtttt 3841 gaatccacat actccaatac tcctgaaata gttcattatg gacagcgcag aagagctggg 3901 gagaattaat tcgtaatcat ggtcatagct gtttcctgtg tgaaattgtt atccgctcac 3961 aattccacac aacatacgag ccggaagcat aaagtgtaaa gcctggggtg cctaatgagt 4021 gagctaactc acattaattg cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc 4081 gtgccagctg cattaatgaa tcggccaacg cgcggggaga ggcggtttgc gtattgggcg 4141 ctcttccgct tcctcgctca ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt 4201 atcagctcac tcaaaggcgg taatacggtt atccacagaa tcaggggata acgcaggaaa 4261 gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc 4321 gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct caagtcagag 4381 gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa gctccctcgt 4441 gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc tcccttcggg 4501 aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt aggtcgttcg 4561 ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg ccttatccgg 4621 taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg cagcagccac 4681 tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg 4741 gcctaactac ggctacacta gaaggacagt atttggtatc tgcgctctgc tgaagccagt 4801 taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg 4861 tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc aagaagatcc 4921 tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt aagggatttt 4981 ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa aatgaagttt 5041 taaatcaatc taaagtatat atgagtaaac ttggtctgac agttaccaat gcttaatcag 5101 tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcct gactccccgt 5161 cgtgtagata actacgatac gggagggctt accatctggc cccagtgctg caatgatacc 5221 gcgagaccca cgctcaccgg ctccagattt atcagcaata aaccagccag ccggaagggc 5281 cgagcgcaga agtggtcctg caactttatc cgcctccatc cagtctatta attgttgccg 5341 ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg ccattgctac 5401 aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg gttcccaacg 5461 atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc 5521 tccgatcgtt gtcagaagta agttggccgc agtgttatca ctcatggtta tggcagcact 5581 gcataattct cttactgtca tgccatccgt aagatgcttt tctgtgactg gtgagtactc 5641 aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat 5701 acgggataat accgcgccac atagcagaac tttaaaagtg ctcatcattg gaaaacgttc 5761 ttcggggcga aaactctcaa ggatcttacc gctgttgaga tccagttcga tgtaacccac 5821 tcgtgcaccc aactgatctt cagcatcttt tactttcacc agcgtttctg ggtgagcaaa 5881 aacaggaagg caaaatgccg caaaaaaggg aataagggcg acacggaaat gttgaatact 5941 catactcttc ctttttcaat attattgaag catttatcag ggttattgtc tcatgagcgg 6001 atacatattt gaatgtattt agaaaaataa acaaataggg gttccgcgca catttccccg 6061 aaaagtgcca cctgacgtct aagaaaccat tattatcatg acattaacct ataaaaatag 6121 gcgtatcacg aggccctttc gtctcgcgcg tttcggtgat gacggtgaaa acctctgaca 6181 catgcagctc ccggagacgg tcacagcttg tctgtaagcg gatgccggga gcagacaagc 6241 ccgtcagggc gcgtcagcgg gtgttggcgg gtgtcggggc tggcttaact atgcggcatc 6301 agagcagatt gtactgagag tgcaccatat gcggtgtgaa ataccgcaca gatgcgtaag 6361 gagaaaatac cgcatcaggc gccattcgcc attcaggctg cgcaactgtt gggaagggcg 6421 atcggtgcgg gcctcttcgc tattacgcca gctggcgaaa gggggatgtg ctgcaaggcg 6481 attaagttgg gtaacgccag ggttttccca gtcacgacgt tgtaaaacga cggccagtgc 6541 caagctcgct ctcccttatg cgactcctgc attaggaagc agcccagtag taggttgagg 6601 ccgttgagca ccgccgccgc aaggaatggt gcatgcaagg agatggcgcc caacagtccc 6661 ccggccacgg ggcctgccac catacccacg ccgaaacaag cgctcatgag cccgaagtgg 6721 cgagcccgat cttccccatc ggtgatgtcg gcgatatagg cgccagcaac cgcacctgtg 6781 gcgccggtga tgccggccac gatgcgtccg gcgtagagga tcgattagtc caatttgtta 6841 aagacaggat atcagtggtc caggctctag ttttgactca acaatatcac cagctgaagc 6901 ctatagagta cgagccatag ataaaataaa agattttatt tagtctccag aaaaaggggg 6961 gaa //