LOCUS Exported 4097 bp ds-DNA circular SYN 13-MAY-2021 DEFINITION Low level expression of GFP fused human RhoA wild type . ACCESSION . VERSION . KEYWORDS pCMVd1-EGFP-hRhoA-wt SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 4097) TITLE In press, Current Biology: RhoA Controls Axon Extension Independent of Specification in the Developing Brain REFERENCE 2 (bases 1 to 4097) AUTHORS . TITLE Direct Submission JOURNAL Exported May 13, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..4097 /organism="synthetic DNA construct" /mol_type="other DNA" promoter 54..153 /label=CMVd1 promoter /note="truncated human cytomegalovirus (CMV) immediate early promoter that yields slightly reduced expression relative to the full-length promoter" primer_bind 104..124 /label=CMV-F /note="Human CMV immediate early promoter, forward primer" regulatory 186..195 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" CDS 192..908 /codon_start=1 /product="enhanced GFP" /label=EGFP /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL EFVTAAGITLGMDELYK" CDS 192..908 /codon_start=1 /product="the original enhanced GFP (Yang et al., 1996)" /label=EGFP /note="mammalian codon-optimized" /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL EFVTAAGITLGMDELYK" primer_bind complement(237..258) /label=EGFP-N /note="EGFP, reverse primer" primer_bind complement(498..517) /label=EXFP-R /note="For distinguishing EGFP variants, reverse primer" primer_bind 845..866 /label=EGFP-C /note="EGFP, forward primer" CDS 915..1496 /codon_start=1 /product="human Rho family small GTPase RhoA" /label=RhoA /translation="MAAIRKKLVIVGDGACGKTCLLIVFSKDQFPEVYVPTVFENYVAD IEVDGKQVELALWDTAGQEDYDRLRPLSYPDTDVILMCFSIDSPDSLENIPEKWTPEVK HFCPNVPIILVGNKKDLRNDEHTRRELAKMKQEPVKPEEGRDMANRIGAFGYMECSAKT KDGVREVFEMATRAALQARRGKKKSGCLVL" primer_bind complement(1569..1587) /label=T7 Term /note="T7 terminator, reverse primer" polyA_signal 1624..1745 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(1635..1654) /label=EBV-rev /note="SV40 polyA terminator, reverse primer" primer_bind 1689..1708 /label=SV40pA-R /note="SV40 polyA, reverse primer" CDS 2144..2938 /codon_start=1 /gene="aph(3')-II (or nptII)" /product="aminoglycoside phosphotransferase from Tn5" /label=NeoR/KanR /note="confers resistance to neomycin, kanamycin, and G418 (Geneticin(R))" /translation="MLEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF" primer_bind complement(2198..2217) /label=Neo-R /note="Neomycin resistance gene, reverse primer" primer_bind 2808..2827 /label=Neo-F /note="Neomycin resistance gene, forward primer" rep_origin complement(3110..3698) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(3190..3209) /label=pBR322ori-F /note="pBR322 origin, forward primer" misc_feature 3814..4097 /label=cer region /note="ColE1-derived recombination site that helps to maintain plasmids as monomers" ORIGIN 1 gataaaccgc agtactccgg ccacgatgcg tccggcgtag aggatcgaga tctccaaaat 61 caacgggact ttccaaaatg tcgtaataac cccgccccgt tgacgcaaat gggcggtagg 121 cgtgtacggt gggaggtcta tataagcaga gctggtttag tgaaccgtca gatcactaga 181 agcttgccac catggtgagc aagggcgagg agctgttcac cggggtggtg cccatcctgg 241 tcgagctgga cggcgacgta aacggccaca agttcagcgt gtccggcgag ggcgagggcg 301 atgccaccta cggcaagctg accctgaagt tcatctgcac caccggcaag ctgcccgtgc 361 cctggcccac cctcgtgacc accctgacct acggcgtgca gtgcttcagc cgctaccccg 421 accacatgaa gcagcacgac ttcttcaagt ccgccatgcc cgaaggctac gtccaggagc 481 gcaccatctt cttcaaggac gacggcaact acaagacccg cgccgaggtg aagttcgagg 541 gcgacaccct ggtgaaccgc atcgagctga agggcatcga cttcaaggag gacggcaaca 601 tcctggggca caagctggag tacaactaca acagccacaa cgtctatatc atggccgaca 661 agcagaagaa cggcatcaag gtgaacttca agatccgcca caacatcgag gacggcagcg 721 tgcagctcgc cgaccactac cagcagaaca cccccatcgg cgacggcccc gtgctgctgc 781 ccgacaacca ctacctgagc acccagtccg ccctgagcaa agaccccaac gagaagcgcg 841 atcacatggt cctgctggag ttcgtgaccg ccgccgggat cactctcggc atggacgagc 901 tgtacaagga attcatggct gccatccgga agaaactggt gattgttggt gatggagcct 961 gtggaaagac atgcttgctc atagtcttca gcaaggacca gttcccagag gtgtatgtgc 1021 ccacagtgtt tgagaactat gtggcagata tcgaggtgga tggaaagcag gtagagttgg 1081 ctttgtggga cacagctggg caggaagatt atgatcgcct gaggcccctc tcctacccag 1141 ataccgatgt tatactgatg tgtttttcca tcgacagccc tgatagttta gaaaacatcc 1201 cagaaaagtg gaccccagaa gtcaagcatt tctgtcccaa cgtgcccatc atcctggttg 1261 ggaataagaa ggatcttcgg aatgatgagc acacaaggcg ggagctagcc aagatgaagc 1321 aggagccggt gaaacctgaa gaaggcagag atatggcaaa caggattggc gcttttgggt 1381 acatggagtg ttcagcaaag accaaagatg gagtgagaga ggtttttgaa atggctacga 1441 gagctgctct gcaagctaga cgtgggaaga aaaaatctgg ttgccttgtc ttgtgactcg 1501 acctgcaggc atgcaagctg atccggctgc taacaaagcc cgaaaggaag ctgagttggc 1561 tgctgccacc gctgagcaat aactagcata accccttggg gcggccgctt cgagcagaca 1621 tgataagata cattgatgag tttggacaaa ccacaactag aatgcagtga aaaaaatgct 1681 ttatttgtga aatttgtgat gctattgctt tatttgtaac cattataagc tgcaataaac 1741 aagttaacaa caacaattgc attcatttta tgtttcaggt tcagggggag atgtgggagg 1801 tttttttaag caagtaaaac ctctacaaat gtggtaaaat cgaattctaa tggatcctct 1861 ttgcgcttgc gttttccctt gtccagatag cccagtagct gacattcatc cggggtcagc 1921 accgtttctg cggactggct ttctacgtgt tccgcttcct ttagcagccc ttgcgccctg 1981 agtgcttgcg gcagcgtgag cttcaaaaga attgccagct ggggcgccct ctggtaaggt 2041 tgggaagccc tgcaaagtaa actggatggc tttcttgccg ccaaggatct gatggcgcag 2101 gggatcaaga tctgatcaag agacaggatg acggtcgttt cgcatgcttg aacaagatgg 2161 attgcacgca ggttctccgg ccgcttgggt ggagaggcta ttcggctatg actgggcaca 2221 acagacaatc ggctgctctg atgccgccgt gttccggctg tcagcgcagg ggcgcccggt 2281 tctttttgtc aagaccgacc tgtccggtgc cctgaatgaa ctgcaggacg aggcagcgcg 2341 gctatcgtgg ctggccacga cgggcgttcc ttgcgcagct gtgctcgacg ttgtcactga 2401 agcgggaagg gactggctgc tattgggcga agtgccgggg caggatctcc tgtcatctca 2461 ccttgctcct gccgagaaag tatccatcat ggctgatgca atgcggcggc tgcatacgct 2521 tgatccggct acctgcccat tcgaccacca agcgaaacat cgcatcgagc gagcacgcac 2581 tcggatggaa gccggtcttg tcgatcagga tgatctggac gaagagcatc aggggctcgc 2641 gccagccgaa ctgttcgcca ggctcaaggc gcgtatgccg gatggtgagg atctcgtcgt 2701 gactcatggc gatgcctgct tgccgaatat catggtggaa aatggccgct tttctggatt 2761 catcgactgt ggccggctgg gtgtggcgga ccgctatcag gacatagcgt tggctacccg 2821 tgatattgct gaagagcttg gcggcgaatg ggctgaccgc ttcctcgtgc tttacggtat 2881 cgccgctccc gattcgcagc gcatcgcctt ctatcgcctt cttgacgagt tcttctgagc 2941 gggactctgg ggttcgaaat gaccgaccaa gcgacgccca accggtatca gctcactcaa 3001 aggcggtaat acggttatcc acagaatcag gggataacgc aggaaagaac atgtgagcaa 3061 aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc 3121 tccgcccccc tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga 3181 caggactata aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc 3241 cgaccctgcc gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt 3301 ctcatagctc acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct 3361 gtgtgcacga accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg 3421 agtccaaccc ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta 3481 gcagagcgag gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct 3541 acactagaag gacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa 3601 gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt 3661 gcaagcagca gattacgcgc agaaaaaaag gatttcaaga agatcctttg atcttttcta 3721 cggggtctga cgctcagtgg aacgaaaact cacgttaagg gattttggtc atgagattat 3781 caaaaaggat cttcacctag atccttttat agtccggaaa tacaggaacg cacgctggat 3841 ggcccttcgc tgggatggtg aaaccatgaa aaatggcagc ttcagtggat taagtggggg 3901 taatgtggcc tgtaccctct ggttgcatag gtattcatac ggttaaaatt tatcaggcgc 3961 gattgcggca gtttttcggg tggtttgttg ccatttttac ctgtctgctg ccgtgatcgc 4021 gctgaacgcg ttttagcggt gcgtacaatt aagggattat ggtaaatcca cttactgtct 4081 gccctcgtag ccatcga //