LOCUS Exported 6038 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION synchronize trafficking of mCherry-GPI from the ER (RUSH system). ACCESSION . VERSION . KEYWORDS Str-KDEL_SBP-mCherry-GPI SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 6038) AUTHORS Boncompain G, Divoux S, Gareil N, de Forges H, Lescure A, Latreche L, Mercanti V, Jollivet F, Raposo G, Perez F TITLE Synchronization of secretory protein traffic in populations of cells. JOURNAL Nat Methods. 2012 Mar 11;9(5):493-8. doi: 10.1038/nmeth.1928. PUBMED 22406856 REFERENCE 2 (bases 1 to 6038) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..6038 /organism="synthetic DNA construct" /mol_type="other DNA" primer_bind complement(44..63) /label=pRS-marker /note="pRS vectors, use to sequence yeast selectable marker" enhancer 235..614 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" promoter 615..818 /label=CMV promoter /note="human cytomegalovirus (CMV) immediate early promoter" primer_bind 769..789 /label=CMV-F /note="Human CMV immediate early promoter, forward primer" primer_bind 863..882 /label=T7 /note="T7 promoter, forward primer" promoter 863..881 /label=T7 promoter /note="promoter for bacteriophage T7 RNA polymerase" intron 1646..1875 /label=chimeric intron /note="chimera between introns from adenovirus and immunoglobulin heavy chain genes" primer_bind 1858..1877 /label=pMT2-F /note="Synthetic intron, forward primer" misc_feature 1933..2506 /label=IRES /note="internal ribosome entry site (IRES) of the encephalomyocarditis virus (EMCV)" primer_bind complement(2100..2117) /label=IRES reverse /note="IRES internal ribosome entry site, reverse primer. Also called pCDH-rev" primer_bind 2327..2346 /label=IRES-F /note="IRES internal ribosome entry site, forward primer" sig_peptide 2533..2592 /product="human interleukin-2 signal sequence" /label=IL-2 signal sequence CDS 2593..2703 /codon_start=1 /product="streptavidin-binding peptide" /label=SBP /note="selected from a peptide library; binds streptavidin with nanomolar affinity (Keefe et al., 2001)" /translation="DEKTTGWRGGHVVEGLAGELEQLRARLEHHPQGQREP" CDS 2713..3417 /codon_start=1 /product="monomeric derivative of DsRed fluorescent protein (Shaner et al., 2004)" /label=mCherry /note="mammalian codon-optimized" /translation="VSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEGT QTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFE DGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALKG EIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERAE GRHSTGGMDELYK" primer_bind complement(2855..2873) /label=mCherry-R /note="mCherry, reverse primer" primer_bind 3123..3142 /label=mCherry-F /note="mCherry, forward primer" polyA_signal 3658..3779 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(3695..3714) /label=SV40pA-R /note="SV40 polyA, reverse primer" primer_bind complement(3881..3897) /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind complement(3881..3897) /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" primer_bind complement(3894..3916) /label=M13/pUC Reverse /note="In lacZ gene" protein_bind 3905..3921 /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(3929..3959) /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 3974..3995 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." primer_bind complement(4112..4129) /label=L4440 /note="L4440 vector, forward primer" rep_origin complement(4283..4871) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(4363..4382) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS complement(5042..5902) /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 5665..5684 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(5903..6007) /gene="bla" /label=AmpR promoter ORIGIN 1 gacggatcgg gagatctccc gatcccctat ggtcgactct cagtacaatc tgctctgatg 61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg 121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc 181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt 241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata 301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc 361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc 421 attgacgtca atgggtggac tatttacggt aaactgccca cttggcagta catcaagtgt 481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt 541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca 601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg 661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc 721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg 781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca 841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gcttggtacc 901 gagctcggat cgatatctgc ggcctagcta gcgcttaagg cctgttaacc ggtatggatg 961 tatgcgtccg tcttgccctg tggctcctct ggggactcct cctgcaccag ggccagagcc 1021 tcagccatag tcacagtgag aaggcgacag gaaccagctc gggcgccggt ttaaacgacc 1081 ctagcaaaga ctcaaaagct caggtgtccg ctgccgaggc tggcattact ggaacatggt 1141 acaatcagct cgggagcacc tttattgtga ctgctggagc cgatggagcc ctcaccggaa 1201 catacgaatc tgctgtggga aacgccgaat cacggtacgt cctcactggc cgatacgata 1261 gtgcccctgc caccgacgga tctgggactg ccctgggatg gactgtcgct tggaaaaaca 1321 actaccggaa tgctcattct gccacaacat ggagtggaca gtacgtggga ggcgctgagg 1381 ctagaatcaa tacacagtgg ctgctcacat ctggcacaac cgaggcaaat gcttggaaat 1441 ccaccctggt gggacatgac acattcacca aagtgaaacc ctccgccgct tcaatcgatg 1501 ccgccaaaaa agccggagtc aacaacggca atcctctgga tgccgtccag cagctcggac 1561 cgaaagatga actgtgagcg gccgcataga taactgatcc agtgtgctgg aattaattcg 1621 ctgtctgcga gggccagctg ttggggtgag tactccctct caaaagcggg catgacttct 1681 gcgctaagat tgtcagtttc caaaaacgag gaggatttga tattcacctg gcccgcggtg 1741 atgcctttga gggtggccgc gtccatctgg tcagaaaaga caatcttttt gttgtcaagc 1801 ttgaggtgtg gcaggcttga gatctggcca tacacttgag tgacaatgac atccactttg 1861 cctttctctc cacaggtgtc cactcccagg tccaactgca ggtcgagcat gcatctaggg 1921 cggccaattc cgcccctctc cctccccccc ccctaacgtt actggccgaa gccgcttgga 1981 ataaggccgg tgtgcgtttg tctatatgtt attttccacc atattgccgt cttttggcaa 2041 tgtgagggcc cggaaacctg gccctgtctt cttgacgagc attcctaggg gtctttcccc 2101 tctcgccaaa ggaatgcaag gtctgttgaa tgtcgtgaag gaagcagttc ctctggaagc 2161 ttcttgaaga caaacaacgt ctgtagcgac cctttgcagg cagcggaacc ccccacctgg 2221 cgacaggtgc ctctgcggcc aaaagccacg tgtataagat acacctgcaa aggcggcaca 2281 accccagtgc cacgttgtga gttggatagt tgtggaaaga gtcaaatggc tctcctcaag 2341 cgtattcaac aaggggctga aggatgccca gaaggtaccc cattgtatgg gatctgatct 2401 ggggcctcgg tgcacatgct ttacatgtgt ttagtcgagg ttaaaaaaac gtctaggccc 2461 cccgaaccac ggggacgtgg ttttcctttg aaaaacacga tgataagctt gccacaaccc 2521 gggaggcgcg ccatgtacag gatgcaactc ctgtcttgca ttgcactaag tcttgcactt 2581 gtcacgaatt ccgacgagaa gaccactggt tggcgaggtg gacacgttgt tgaaggactg 2641 gctggggaac ttgaacaact tcgtgcacga ctggagcatc acccacaagg tcaacgtgaa 2701 ccacctgcag gtgtgagcaa gggcgaggag gataacatgg ccatcatcaa ggagttcatg 2761 cgcttcaagg tgcacatgga gggctccgtg aacggccacg agttcgagat cgagggcgag 2821 ggcgagggcc gcccctacga gggcacccag accgccaagc tgaaggtgac caagggtggc 2881 cccctgccct tcgcctggga catcctgtcc cctcagttca tgtacggctc caaggcctac 2941 gtgaagcacc ccgccgacat ccccgactac ttgaagctgt ccttccccga gggcttcaag 3001 tgggagcgcg tgatgaactt cgaggacggc ggcgtggtga ccgtgaccca ggactcctcc 3061 ctacaggacg gcgagttcat ctacaaggtg aagctgcgcg gcaccaactt cccctccgac 3121 ggccccgtaa tgcagaagaa gaccatgggc tgggaggcct cctccgagcg gatgtacccc 3181 gaggacggcg ccctgaaggg cgagatcaag cagaggctga agctgaagga cggcggccac 3241 tacgacgctg aggtcaagac cacctacaag gccaagaagc ccgtgcagct gcccggcgcc 3301 tacaacgtca acatcaagtt ggacatcacc tcccacaacg aggactacac catcgtggaa 3361 cagtacgaac gcgccgaggg ccgccactcc accggcggca tggacgagct gtacaagggc 3421 cggccactgg aaaatggcgg aacctctctg tctgaaaaaa ctgtgctgct gctggtgact 3481 ccctttctgg ccgctgcttg gtccctccac ccatgaggcc tcgagggcca attaattaac 3541 tctagataac tgatcataat cagccatacc acatttgtag aggttttact tgctttaaaa 3601 aacctcccac acctccccct gaacctgaaa cataaaatga atgcaattgt tgttgttaac 3661 ttgtttattg cagcttataa tggttacaaa taaagcaata gcatcacaaa tttcacaaat 3721 aaagcatttt tttcactgca ttctagttgt ggtttgtcca aactcatcaa tgtatcttaa 3781 cgcgtcgagt gcattctagt tgtggtttgt ccaaactcat caatgtatct tatcatgtct 3841 gtataccgtc gacctctagc tagagcttgg cgtaatcatg gtcatagctg tttcctgtgt 3901 gaaattgtta tccgctcaca attccacaca acatacgagc cggaagcata aagtgtaaag 3961 cctggggtgc ctaatgagtg agctaactca cattaattgc gttgcgctca ctgcccgctt 4021 tccagtcggg aaacctgtcg tgccagctgc attaatgaat cggccaacgc gcggggagag 4081 gcggtttgcg tattgggcgc tcttccgctt cctcgctcac tgactcgctg cgctcggtcg 4141 ttcggctgcg gcgagcggta tcagctcact caaaggcggt aatacggtta tccacagaat 4201 caggggataa cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta 4261 aaaaggccgc gttgctggcg tttttccata ggctccgccc ccctgacgag catcacaaaa 4321 atcgacgctc aagtcagagg tggcgaaacc cgacaggact ataaagatac caggcgtttc 4381 cccctggaag ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt 4441 ccgcctttct cccttcggga agcgtggcgc tttctcaatg ctcacgctgt aggtatctca 4501 gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc gttcagcccg 4561 accgctgcgc cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat 4621 cgccactggc agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta 4681 cagagttctt gaagtggtgg cctaactacg gctacactag aaggacagta tttggtatct 4741 gcgctctgct gaagccagtt accttcggaa aaagagttgg tagctcttga tccggcaaac 4801 aaaccaccgc tggtagcggt ggtttttttg tttgcaagca gcagattacg cgcagaaaaa 4861 aaggatctca agaagatcct ttgatctttt ctacggggtc tgacgctcag tggaacgaaa 4921 actcacgtta agggattttg gtcatgagat tatcaaaaag gatcttcacc tagatccttt 4981 taaattaaaa atgaagtttt aaatcaatct aaagtatata tgagtaaact tggtctgaca 5041 gttaccaatg cttaatcagt gaggcaccta tctcagcgat ctgtctattt cgttcatcca 5101 tagttgcctg actccccgtc gtgtagataa ctacgatacg ggagggctta ccatctggcc 5161 ccagtgctgc aatgataccg cgagacccac gctcaccggc tccagattta tcagcaataa 5221 accagccagc cggaagggcc gagcgcagaa gtggtcctgc aactttatcc gcctccatcc 5281 agtctattaa ttgttgccgg gaagctagag taagtagttc gccagttaat agtttgcgca 5341 acgttgttgc cattgctaca ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat 5401 tcagctccgg ttcccaacga tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag 5461 cggttagctc cttcggtcct ccgatcgttg tcagaagtaa gttggccgca gtgttatcac 5521 tcatggttat ggcagcactg cataattctc ttactgtcat gccatccgta agatgctttt 5581 ctgtgactgg tgagtactca accaagtcat tctgagaata gtgtatgcgg cgaccgagtt 5641 gctcttgccc ggcgtcaata cgggataata ccgcgccaca tagcagaact ttaaaagtgc 5701 tcatcattgg aaaacgttct tcggggcgaa aactctcaag gatcttaccg ctgttgagat 5761 ccagttcgat gtaacccact cgtgcaccca actgatcttc agcatctttt actttcacca 5821 gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga ataagggcga 5881 cacggaaatg ttgaatactc atactcttcc tttttcaata ttattgaagc atttatcagg 5941 gttattgtct catgagcgga tacatatttg aatgtattta gaaaaataaa caaatagggg 6001 ttccgcgcac atttccccga aaagtgccac ctgacgtc //