LOCUS       sequence-120582-        6281 bp    DNA     circular SYN 14-APR-2025
DEFINITION  synchronize trafficking of ManII from the ER (RUSH system).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6281)
  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 6281)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; doi: "10.1038/nmeth"; 
            journalName: "Nat Methods"; date: "2012-03-11- 11"; volume: "9"; 
            issue: "5"; pages: "493-8"
COMMENT     Sequence Label: Str-KDEL_ManII-SBP-EGFP
FEATURES             Location/Qualifiers
     source          1..6281
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     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    join(1933..2507,2508..2510,2511..2519)
                     /label=IRES2
                     /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"
     CDS             2908..3021
                     /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="MDEKTTGWRGGHVVEGLAGELEQLRARLEHHPQGQREP"
     CDS             join(3031..3033,3034..3036,3037..3747)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(3031..3033,3034..3036,3037..3747)
                     /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(3076..3097)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(3337..3356)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3684..3705
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    3901..4022
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3938..3957)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(4124..4140)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4124..4140)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4137..4159)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4148..4164
                     /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(join(4172..4178,4179..4196,4197..4202))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4217..4238
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4355..4372)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4526..5114)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4606..4625)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5285..6076,6077..6145))
                     /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     5908..5927
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6146..6250)
                     /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 ccatgaagtt aagtcgccag ttcaccgtgt ttggcagcgc gatcttctgc
     2581 gtcgtaatct tctcactcta cctgatgctg gacaggggtc acttggacta ccctcggggc
     2641 ccgcgccagg agggctcctt tccgcagggc cagctttcaa tattgcaaga aaagattgac
     2701 catttggagc gtttgctcgc tgagaacaac gagattatct caaatatcag agactcagtc
     2761 atcaacctga gcgagtctgt ggaggacggc ccgcgggggt caccaggcaa cgccagccaa
     2821 ggctccatcc acctccactc gccacagttg gccctgcagg ctgaccccag agactgtttg
     2881 gatcccaccg gtcgccaccg gaattccatg gacgagaaga ccactggttg gcgaggtgga
     2941 cacgttgttg aaggactggc tggggaactt gaacaacttc gtgcacgact ggagcatcac
     3001 ccacaaggtc aacgtgaacc acctgcaggt atggtgagca aaggcgagga gctgttcacc
     3061 ggggtggtgc ccatcctggt cgagctggac ggcgacgtaa acggccacaa gttcagcgtg
     3121 tccggcgagg gcgagggcga tgccacctac ggcaagctga ccctgaagtt catctgcacc
     3181 accggcaagc tgcccgtgcc ctggcccacc ctcgtgacca ccctgaccta cggcgtgcag
     3241 tgcttcagcc gctaccccga ccacatgaag cagcacgact tcttcaagtc cgccatgccc
     3301 gaaggctacg tccaggagcg caccatcttc ttcaaggacg acggcaacta caagacccgc
     3361 gccgaggtga agttcgaggg cgacaccctg gtgaaccgca tcgagctgaa gggcatcgac
     3421 ttcaaggagg acggcaacat cctggggcac aagctggagt acaactacaa cagccacaac
     3481 gtctatatca tggccgacaa gcagaagaac ggcatcaagg tgaacttcaa gatccgccac
     3541 aacatcgagg acggcagcgt gcagctcgcc gaccactacc agcagaacac ccccatcggc
     3601 gacggccccg tgctgctgcc cgacaaccac tacctgagca cccagtccgc cctgagcaaa
     3661 gaccccaacg agaagcgcga tcacatggtc ctgctggagt tcgtgaccgc cgccgggatc
     3721 actctcggca tggacgagct gtacaagggc cggccttaag gcctcgaggg ccaattaatt
     3781 aactctagat aactgatcat aatcagccat accacatttg tagaggtttt acttgcttta
     3841 aaaaacctcc cacacctccc cctgaacctg aaacataaaa tgaatgcaat tgttgttgtt
     3901 aacttgttta ttgcagctta taatggttac aaataaagca atagcatcac aaatttcaca
     3961 aataaagcat ttttttcact gcattctagt tgtggtttgt ccaaactcat caatgtatct
     4021 taacgcgtcg agtgcattct agttgtggtt tgtccaaact catcaatgta tcttatcatg
     4081 tctgtatacc gtcgacctct agctagagct tggcgtaatc atggtcatag ctgtttcctg
     4141 tgtgaaattg ttatccgctc acaattccac acaacatacg agccggaagc ataaagtgta
     4201 aagcctgggg tgcctaatga gtgagctaac tcacattaat tgcgttgcgc tcactgcccg
     4261 ctttccagtc gggaaacctg tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga
     4321 gaggcggttt gcgtattggg cgctcttccg cttcctcgct cactgactcg ctgcgctcgg
     4381 tcgttcggct gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg ttatccacag
     4441 aatcagggga taacgcagga aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc
     4501 gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac gagcatcaca
     4561 aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga taccaggcgt
     4621 ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt accggatacc
     4681 tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca atgctcacgc tgtaggtatc
     4741 tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc
     4801 ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta agacacgact
     4861 tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat gtaggcggtg
     4921 ctacagagtt cttgaagtgg tggcctaact acggctacac tagaaggaca gtatttggta
     4981 tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct tgatccggca
     5041 aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa
     5101 aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct cagtggaacg
     5161 aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc acctagatcc
     5221 ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa acttggtctg
     5281 acagttacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta tttcgttcat
     5341 ccatagttgc ctgactcccc gtcgtgtaga taactacgat acgggagggc ttaccatctg
     5401 gccccagtgc tgcaatgata ccgcgagacc cacgctcacc ggctccagat ttatcagcaa
     5461 taaaccagcc agccggaagg gccgagcgca gaagtggtcc tgcaacttta tccgcctcca
     5521 tccagtctat taattgttgc cgggaagcta gagtaagtag ttcgccagtt aatagtttgc
     5581 gcaacgttgt tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt
     5641 cattcagctc cggttcccaa cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa
     5701 aagcggttag ctccttcggt cctccgatcg ttgtcagaag taagttggcc gcagtgttat
     5761 cactcatggt tatggcagca ctgcataatt ctcttactgt catgccatcc gtaagatgct
     5821 tttctgtgac tggtgagtac tcaaccaagt cattctgaga atagtgtatg cggcgaccga
     5881 gttgctcttg cccggcgtca atacgggata ataccgcgcc acatagcaga actttaaaag
     5941 tgctcatcat tggaaaacgt tcttcggggc gaaaactctc aaggatctta ccgctgttga
     6001 gatccagttc gatgtaaccc actcgtgcac ccaactgatc ttcagcatct tttactttca
     6061 ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg
     6121 cgacacggaa atgttgaata ctcatactct tcctttttca atattattga agcatttatc
     6181 agggttattg tctcatgagc ggatacatat ttgaatgtat ttagaaaaat aaacaaatag
     6241 gggttccgcg cacatttccc cgaaaagtgc cacctgacgt c
//
