LOCUS Exported 5481 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION Expresses mCherry-tagged PH(K14A) domain of human Akt1. ACCESSION . VERSION . KEYWORDS pmCherry-PH1(K14A) SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 5481) AUTHORS Ebner M, Lucic I, Leonard TA, Yudushkin I TITLE PI(3,4,5)P3 Engagement Restricts Akt Activity to Cellular Membranes. JOURNAL Mol Cell. 2017 Feb 2;65(3):416-431.e6. doi: 10.1016/j.molcel.2016.12.028. PUBMED 28157504 REFERENCE 2 (bases 1 to 5481) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..5481 /organism="synthetic DNA construct" /mol_type="other DNA" rep_origin 5..593 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 494..513 /label=pBR322ori-F /note="pBR322 origin, forward primer" enhancer 768..1071 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" promoter 1072..1275 /label=CMV promoter /note="human cytomegalovirus (CMV) immediate early promoter" primer_bind 1226..1246 /label=CMV-F /note="Human CMV immediate early promoter, forward primer" regulatory 1314..1323 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" CDS 1320..2027 /codon_start=1 /product="monomeric derivative of DsRed fluorescent protein (Shaner et al., 2004)" /label=mCherry /note="mammalian codon-optimized" /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA EGRHSTGGMDELYK" primer_bind complement(1465..1483) /label=mCherry-R /note="mCherry, reverse primer" primer_bind 1733..1752 /label=mCherry-F /note="mCherry, forward primer" polyA_signal 2976..3097 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(3013..3032) /label=SV40pA-R /note="SV40 polyA, reverse primer" primer_bind 3067..3086 /label=EBV-rev /note="SV40 polyA terminator, reverse primer" rep_origin complement(3104..3559) /direction=LEFT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(3241..3262) /label=F1ori-F /note="F1 origin, forward primer" primer_bind 3453..3472 /label=F1ori-R /note="F1 origin, reverse primer" promoter 3586..3690 /gene="bla" /label=AmpR promoter promoter 3692..4049 /label=SV40 promoter /note="SV40 enhancer and early promoter" primer_bind complement(3715..3735) /label=pBABE 3' /note="SV40 enhancer, reverse primer for pBABE vectors" rep_origin 3900..4035 /label=SV40 ori /note="SV40 origin of replication" primer_bind 3962..3981 /label=SV40pro-F /note="SV40 promoter/origin, forward primer" CDS 4084..4878 /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="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF" primer_bind complement(4138..4157) /label=Neo-R /note="Neomycin resistance gene, reverse primer" primer_bind 4748..4767 /label=Neo-F /note="Neomycin resistance gene, forward primer" primer_bind complement(5066..5085) /label=TK-pA-R /note="Thymidine kinase polyA, reverse primer" polyA_signal 5110..5157 /label=HSV TK poly(A) signal /note="herpes simplex virus thymidine kinase polyadenylation signal (Cole and Stacy, 1985)" ORIGIN 1 cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc 61 taccagcggt ggtttgtttg ccggatcaag agctaccaac tctttttccg aaggtaactg 121 gcttcagcag agcgcagata ccaaatactg ttcttctagt gtagccgtag ttaggccacc 181 acttcaagaa ctctgtagca ccgcctacat acctcgctct gctaatcctg ttaccagtgg 241 ctgctgccag tggcgataag tcgtgtctta ccgggttgga ctcaagacga tagttaccgg 301 ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa 361 cgacctacac cgaactgaga tacctacagc gtgagctatg agaaagcgcc acgcttcccg 421 aagggagaaa ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga 481 gggagcttcc agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct 541 gacttgagcg tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaaaacgcca 601 gcaacgcggc ctttttacgg ttcctggcct tttgctggcc ttttgctcac atgttctttc 661 ctgcgttatc ccctgattct gtggataacc gtattaccgc catgcattag ttattaatag 721 taatcaatta cggggtcatt agttcatagc ccatatatgg agttccgcgt tacataactt 781 acggtaaatg gcccgcctgg ctgaccgccc aacgaccccc gcccattgac gtcaataatg 841 acgtatgttc ccatagtaac gccaataggg actttccatt gacgtcaatg ggtggagtat 901 ttacggtaaa ctgcccactt ggcagtacat caagtgtatc atatgccaag tacgccccct 961 attgacgtca atgacggtaa atggcccgcc tggcattatg cccagtacat gaccttatgg 1021 gactttccta cttggcagta catctacgta ttagtcatcg ctattaccat ggtgatgcgg 1081 ttttggcagt acatcaatgg gcgtggatag cggtttgact cacggggatt tccaagtctc 1141 caccccattg acgtcaatgg gagtttgttt tggcaccaaa atcaacggga ctttccaaaa 1201 tgtcgtaaca actccgcccc attgacgcaa atgggcggta ggcgtgtacg gtgggaggtc 1261 tatataagca gagctggttt agtgaaccgt cagatccgct agcgctaccg gtcgccacca 1321 tggtgagcaa gggcgaggag gataacatgg ccatcatcaa ggagttcatg cgcttcaagg 1381 tgcacatgga gggctccgtg aacggccacg agttcgagat cgagggcgag ggcgagggcc 1441 gcccctacga gggcacccag accgccaagc tgaaggtgac caagggtggc cccctgccct 1501 tcgcctggga catcctgtcc cctcagttca tgtacggctc caaggcctac gtgaagcacc 1561 ccgccgacat ccccgactac ttgaagctgt ccttccccga gggcttcaag tgggagcgcg 1621 tgatgaactt cgaggacggc ggcgtggtga ccgtgaccca ggactcctcc ctgcaggacg 1681 gcgagttcat ctacaaggtg aagctgcgcg gcaccaactt cccctccgac ggccccgtaa 1741 tgcagaagaa gaccatgggc tgggaggcct cctccgagcg gatgtacccc gaggacggcg 1801 ccctgaaggg cgagatcaag cagaggctga agctgaagga cggcggccac tacgacgctg 1861 aggtcaagac cacctacaag gccaagaagc ccgtgcagct gcccggcgcc tacaacgtca 1921 acatcaagtt ggacatcacc tcccacaacg aggactacac catcgtggaa cagtacgaac 1981 gcgccgaggg ccgccactcc accggcggca tggacgagct gtacaagtcc ggactcagat 2041 ctcgagctat gagcgacgtg gctattgtga aggagggttg gctgcacgca cgaggggagt 2101 acatcaagac ctggcggcca cgctacttcc tcctcaagaa tgatggcacc ttcattggct 2161 acaaggagcg gccgcaggat gtggaccaac gtgaggctcc cctcaacaac ttctctgtgg 2221 cgcagtgcca gctgatgaag acggagcggc cccggcccaa caccttcatc atccgctgcc 2281 tgcagtggac cactgtcatc gaacgcacct tccatgtgga gactcctgag gagcgggagg 2341 agtggacaac cgccatccag actgtggctg acggcctcaa gaagttggac atcacctccc 2401 acaacgagga ctacaccatc gtggaacagt acgaacgcgc cgagggccgc cactccaccg 2461 gcggcatgga cgagctgtac aagtccggac tcagatctcg agctatgagc gacgtggcta 2521 ttgtgaagga gggttggctg cacgcacgag gggagtacat caagacctgg cggccacgct 2581 acttcctcct caagaatgat ggcaccttca ttggctacaa ggagcggccg caggatgtgg 2641 accaacgtga ggctcccctc aacaacttct ctgtggcgca gtgccagctg atgaagacgg 2701 agcggccccg gcccaacacc ttcatcatcc gctgcctgca gtggaccact gtcatcgaac 2761 gcaccttcca tgtggagact cctgaggagc gggaggagtg gacaaccgcc atccagactg 2821 tggctgacgg cctcaagaag caggaggagg aggagtaatc tagataactg atcataatca 2881 gccataccac atttgtagag gttttacttg ctttaaaaaa cctcccacac ctccccctga 2941 acctgaaaca taaaatgaat gcaattgttg ttgttaactt gtttattgca gcttataatg 3001 gttacaaata aagcaatagc atcacaaatt tcacaaataa agcatttttt tcactgcatt 3061 ctagttgtgg tttgtccaaa ctcatcaatg tatcttaacg cgtaaattgt aagcgttaat 3121 attttgttaa aattcgcgtt aaatttttgt taaatcagct cattttttaa ccaataggcc 3181 gaaatcggca aaatccctta taaatcaaaa gaatagaccg agatagggtt gagtgttgtt 3241 ccagtttgga acaagagtcc actattaaag aacgtggact ccaacgtcaa agggcgaaaa 3301 accgtctatc agggcgatgg cccactacgt gaaccatcac cctaatcaag ttttttgggg 3361 tcgaggtgcc gtaaagcact aaatcggaac cctaaaggga gcccccgatt tagagcttga 3421 cggggaaagc cggcgaacgt ggcgagaaag gaagggaaga aagcgaaagg agcgggcgct 3481 agggcgctgg caagtgtagc ggtcacgctg cgcgtaacca ccacacccgc cgcgcttaat 3541 gcgccgctac agggcgcgtc aggtggcact tttcggggaa atgtgcgcgg aacccctatt 3601 tgtttatttt tctaaataca ttcaaatatg tatccgctca tgagacaata accctgataa 3661 atgcttcaat aatattgaaa aaggaagagt cctgaggcgg aaagaaccag ctgtggaatg 3721 tgtgtcagtt agggtgtgga aagtccccag gctccccagc aggcagaagt atgcaaagca 3781 tgcatctcaa ttagtcagca accaggtgtg gaaagtcccc aggctcccca gcaggcagaa 3841 gtatgcaaag catgcatctc aattagtcag caaccatagt cccgccccta actccgccca 3901 tcccgcccct aactccgccc agttccgccc attctccgcc ccatggctga ctaatttttt 3961 ttatttatgc agaggccgag gccgcctcgg cctctgagct attccagaag tagtgaggag 4021 gcttttttgg aggcctaggc ttttgcaaag atcgatcaag agacaggatg aggatcgttt 4081 cgcatgattg aacaagatgg attgcacgca ggttctccgg ccgcttgggt ggagaggcta 4141 ttcggctatg actgggcaca acagacaatc ggctgctctg atgccgccgt gttccggctg 4201 tcagcgcagg ggcgcccggt tctttttgtc aagaccgacc tgtccggtgc cctgaatgaa 4261 ctgcaagacg aggcagcgcg gctatcgtgg ctggccacga cgggcgttcc ttgcgcagct 4321 gtgctcgacg ttgtcactga agcgggaagg gactggctgc tattgggcga agtgccgggg 4381 caggatctcc tgtcatctca ccttgctcct gccgagaaag tatccatcat ggctgatgca 4441 atgcggcggc tgcatacgct tgatccggct acctgcccat tcgaccacca agcgaaacat 4501 cgcatcgagc gagcacgtac tcggatggaa gccggtcttg tcgatcagga tgatctggac 4561 gaagagcatc aggggctcgc gccagccgaa ctgttcgcca ggctcaaggc gagcatgccc 4621 gacggcgagg atctcgtcgt gacccatggc gatgcctgct tgccgaatat catggtggaa 4681 aatggccgct tttctggatt catcgactgt ggccggctgg gtgtggcgga ccgctatcag 4741 gacatagcgt tggctacccg tgatattgct gaagagcttg gcggcgaatg ggctgaccgc 4801 ttcctcgtgc tttacggtat cgccgctccc gattcgcagc gcatcgcctt ctatcgcctt 4861 cttgacgagt tcttctgagc gggactctgg ggttcgaaat gaccgaccaa gcgacgccca 4921 acctgccatc acgagatttc gattccaccg ccgccttcta tgaaaggttg ggcttcggaa 4981 tcgttttccg ggacgccggc tggatgatcc tccagcgcgg ggatctcatg ctggagttct 5041 tcgcccaccc tagggggagg ctaactgaaa cacggaagga gacaataccg gaaggaaccc 5101 gcgctatgac ggcaataaaa agacagaata aaacgcacgg tgttgggtcg tttgttcata 5161 aacgcggggt tcggtcccag ggctggcact ctgtcgatac cccaccgaga ccccattggg 5221 gccaatacgc ccgcgtttct tccttttccc caccccaccc cccaagttcg ggtgaaggcc 5281 cagggctcgc agccaacgtc ggggcggcag gccctgccat agcctcaggt tactcatata 5341 tactttagat tgatttaaaa cttcattttt aatttaaaag gatctaggtg aagatccttt 5401 ttgataatct catgaccaaa atcccttaac gtgagttttc gttccactga gcgtcagacc 5461 ccgtagaaaa gatcaaagga t //