LOCUS Exported 5406 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION Homology arms and mEGFP-linker sequence for N-terminus tagging of human RAB5A. ACCESSION . VERSION . KEYWORDS AICSDP-39:RAB5A-mEGFP SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 5406) AUTHORS Allen Institute for Cell Science TITLE Allen Institute for Cell Science Plasmids JOURNAL Unpublished REFERENCE 2 (bases 1 to 5406) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..5406 /organism="synthetic DNA construct" /mol_type="other DNA" primer_bind 173..195 /label=M13/pUC Forward /note="In lacZ gene" primer_bind 187..204 /label=M13 Forward /note="In lacZ gene. Also called M13-F20 or M13 (-21) Forward" primer_bind 188..204 /label=M13 fwd /note="common sequencing primer, one of multiple similar variants" CDS 1222..1938 /codon_start=1 /product="enhanced GFP with monomerizing A206K mutation (Zacharias et al., 2002)" /label=mEGFP /note="mammalian codon-optimized" /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLL EFVTAAGITLGMDELYK" primer_bind complement(1267..1288) /label=EGFP-N /note="EGFP, reverse primer" primer_bind complement(1528..1547) /label=EXFP-R /note="For distinguishing EGFP variants, reverse primer" primer_bind 1875..1896 /label=EGFP-C /note="EGFP, forward primer" primer_bind complement(2994..3010) /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind complement(2994..3010) /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" primer_bind complement(3007..3029) /label=M13/pUC Reverse /note="In lacZ gene" protein_bind 3018..3034 /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(3042..3072) /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 3087..3108 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." primer_bind complement(3225..3242) /label=L4440 /note="L4440 vector, forward primer" rep_origin complement(3396..3984) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(3476..3495) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS complement(4155..5015) /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 4778..4797 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(5016..5120) /gene="bla" /label=AmpR promoter primer_bind 5188..5206 /label=pBRforEco /note="pBR322 vectors, upsteam of EcoRI site, forward primer" primer_bind complement(5244..5266) /label=pGEX 3' /note="pGEX vectors, reverse primer" primer_bind 5366..5385 /label=pRS-marker /note="pRS vectors, use to sequence yeast selectable marker" ORIGIN 1 tgtgaaatac cgcacagatg cgtaaggaga aaataccgca tcaggcgcca ttcgccattc 61 aggctgcgca actgttggga agggcgatcg gtgcgggcct cttcgctatt acgccagctg 121 gcgaaagggg gatgtgctgc aaggcgatta agttgggtaa cgccagggtt ttcccagtca 181 cgacgttgta aaacgacggc cagtgaattg acgcgtattg ggatgttctg agcgcctgta 241 gtctcagcta ctcaggaggt tgaggcagga ggattgcttg agcccaggag gtctaggttg 301 cagtgagctg tgaccacgcc actgcactcc agcctgggtg agagagcaaa accctttctc 361 caaaaaaatt tttttaaaaa tggcatgtat attgttcgtc taaagactac aaaaaagtta 421 caattattgt aagtaaataa gtcagtaata atcaccacat ggagaacaac tgttagcatt 481 tctgtgtatg gtgtgcttga gcagatgagg aaacagtttt aagtctgccg ttaggcacac 541 aaacctggga gactgaaaat atttgttgtc caaaattacc attaattaaa tgaaaaatgc 601 atcgtttaac agaaaacttc aaataatact aaattttagg atctgttcta aaagtttccc 661 atttattaat aacataattt ttctcaaaac aatgttaatc caagtagcct ggctacagaa 721 cttgaactcg agtcatctct caggttgcct gggtttgaaa ctcggctctg tcacttctgt 781 gacaataact gtgtgacctc aattaagctt cttaacatct ctgtgcttgt ttcctcattt 841 ataaggattg taataatatc tcattttata ggattgttga gaagttaagt acttaaagca 901 ctgcctggca caaacttgaa gctttgaggt tagcctttaa aaaaaaaagt aattttggac 961 tcacatccta tacctttgaa tttcctttaa aagatagtga cataatctaa agggcattga 1021 aggtctagta gagtttaaga taatcatgct ataggcaata tttaaagtga ttaatagtaa 1081 atttgcttta ctgatttgta tatttaattc ataattgttt ctttacaggt ttctttacct 1141 ccagaaagaa gaatattggc cccttgaatt ctggaagttc attgaagagt ctgaaattag 1201 ggacttattt caaatttgga catggtgagc aagggcgagg agctgttcac cggggtggtg 1261 cccatcctgg tcgagctgga cggcgacgta aacggccaca agttcagcgt gtccggcgag 1321 ggcgagggcg atgccaccta cggcaagctg accctgaagt tcatctgcac caccggcaag 1381 ctgcccgtgc cctggcccac cctcgtgacc accctgacct acggcgtgca gtgcttcagc 1441 cgctaccccg accacatgaa gcagcacgac ttcttcaagt ccgccatgcc cgaaggctac 1501 gtccaggagc gcaccatctt cttcaaggac gacggcaact acaagacccg cgccgaggtg 1561 aagttcgagg gcgacaccct ggtgaaccgc atcgagctga agggcatcga cttcaaggag 1621 gacggcaaca tcctggggca caagctggag tacaactaca acagccacaa cgtctatatc 1681 atggccgaca agcagaagaa cggcatcaag gtgaacttca agatccgcca caacatcgag 1741 gacggcagcg tgcagctcgc cgaccactac cagcagaaca cccccatcgg cgacggcccc 1801 gtgctgctgc ccgacaacca ctacctgagc acccagtcca agctgagcaa agaccccaac 1861 gagaagcgcg atcacatggt cctgctggag ttcgtgaccg ccgccgggat cactctcggc 1921 atggacgagc tgtacaagag cggcctgaga agcagggccg ctagtcgagg cgccacacgg 1981 ccaaacgggc caaatacggg aaataaaata tgccagttca aactagtact tctgggagag 2041 tccgctgttg gcaaatcaag cctagtgctt cgttttgtga aaggccaatt tcatgaattt 2101 caagagagta ccattggggg tgagattttc ttttttccct gcttcattta atcgaatcat 2161 acattgacag aaaatatttt gatgttccaa ttgtgtgatt atgaatagct aaataagtca 2221 tagagtgaag aaaagagctg ttcagcttac cttagcttta aaacttgcct ttatctttgt 2281 taaaatgatg tgtttttata tttgattgtt tggttaatcc cttgtggtta tattgtagct 2341 aaaatacaga taaaaaacat gagtagaatc agtgtctcaa gtaataacat ttgacctgat 2401 acctatttct agctaacaac tagaagcttc ttcaaattaa tacagtacat gtttgtgctc 2461 aattatattg gtttaaaatg aatagtaaaa aaggtaacct taaattctga tctatataac 2521 tgaagtattt gcaaataatt atacttactt gtaaaattaa atgtaattaa atactgctag 2581 atgtactttt ttttggtatt taagagatgg gatcttgctg tcgcctaggc tggagtgcag 2641 tgtccggatc atagctcact gcagactgca gcctgaaact ctaggctcaa atgatcttct 2701 tgccttagcc tcccaccaag tagttggacc taagggtgca tgccaccatg ccaggcaagt 2761 tttttttttt tttttttaag agtcagagtc tcgctgtgtt tccaggttgg tcttgaactc 2821 ctggcctcaa gtgagcctcc agcctcaacc tcccaggtag ctgtgattac aggtgctccc 2881 tgctgagatg catatgtaag aatgctccta agctgaggga agagaaaaag tgattaatgg 2941 aaagggacat aacaggaaaa tcccaatggc gcgccgagct tggcgtaatc atggtcatag 3001 ctgtttcctg tgtgaaattg ttatccgctc acaattccac acaacatacg agccggaagc 3061 ataaagtgta aagcctgggg tgcctaatga gtgagctaac tcacattaat tgcgttgcgc 3121 tcactgcccg ctttccagtc gggaaacctg tcgtgccagc tgcattaatg aatcggccaa 3181 cgcgcgggga gaggcggttt gcgtattggg cgctcttccg cttcctcgct cactgactcg 3241 ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg 3301 ttatccacag aatcagggga taacgcagga aagaacatgt gagcaaaagg ccagcaaaag 3361 gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac 3421 gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga 3481 taccaggcgt ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt 3541 accggatacc tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc 3601 tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc 3661 cccgttcagc ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta 3721 agacacgact tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat 3781 gtaggcggtg ctacagagtt cttgaagtgg tggcctaact acggctacac tagaagaaca 3841 gtatttggta tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct 3901 tgatccggca aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt 3961 acgcgcagaa aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct 4021 cagtggaacg aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc 4081 acctagatcc ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa 4141 acttggtctg acagttacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta 4201 tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga taactacgat acgggagggc 4261 ttaccatctg gccccagtgc tgcaatgata ccgcgagacc cacgctcacc ggctccagat 4321 ttatcagcaa taaaccagcc agccggaagg gccgagcgca gaagtggtcc tgcaacttta 4381 tccgcctcca tccagtctat taattgttgc cgggaagcta gagtaagtag ttcgccagtt 4441 aatagtttgc gcaacgttgt tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt 4501 ggtatggctt cattcagctc cggttcccaa cgatcaaggc gagttacatg atcccccatg 4561 ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg ttgtcagaag taagttggcc 4621 gcagtgttat cactcatggt tatggcagca ctgcataatt ctcttactgt catgccatcc 4681 gtaagatgct tttctgtgac tggtgagtac tcaaccaagt cattctgaga atagtgtatg 4741 cggcgaccga gttgctcttg cccggcgtca atacgggata ataccgcgcc acatagcaga 4801 actttaaaag tgctcatcat tggaaaacgt tcttcggggc gaaaactctc aaggatctta 4861 ccgctgttga gatccagttc gatgtaaccc actcgtgcac ccaactgatc ttcagcatct 4921 tttactttca ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag 4981 ggaataaggg cgacacggaa atgttgaata ctcatactct tcctttttca atattattga 5041 agcatttatc agggttattg tctcatgagc ggatacatat ttgaatgtat ttagaaaaat 5101 aaacaaatag gggttccgcg cacatttccc cgaaaagtgc cacctgacgt ctaagaaacc 5161 attattatca tgacattaac ctataaaaat aggcgtatca cgaggccctt tcgtctcgcg 5221 cgtttcggtg atgacggtga aaacctctga cacatgcagc tcccggagac ggtcacagct 5281 tgtctgtaag cggatgccgg gagcagacaa gcccgtcagg gcgcgtcagc gggtgttggc 5341 gggtgtcggg gctggcttaa ctatgcggca tcagagcaga ttgtactgag agtgcaccat 5401 atgcgg //