LOCUS       sequence-328222-        6235 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  RUSH cargo: Str-KDEL_IRES_SBP-Halo-GPI.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6235)
  AUTHORS   Weigel AV, Chang CL, Shtengel G, Xu CS, Hoffman DP, Freeman M, Iyer 
            N, Aaron J, Khuon S, Bogovic J, Qiu W, Hess HF, Lippincott-Schwartz 
            J
  TITLE     ER-to-Golgi protein delivery through an interwoven, tubular network 
            extending from ER.
  JOURNAL   Cell. 2021 Apr 29;184(9):2412-2429.e16. doi: 
            10.1016/j.cell.2021.03.035. Epub 2021 Apr 13.
  PUBMED    33852913
REFERENCE   2  (bases 1 to 6235)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; doi: 
            "10.1016/j.cell.2021.03.035"; journalName: "Cell"; date: 
            "2021-04-29- 29"; volume: "184"; issue: "9"; pages: "2412-2429.e16"
COMMENT     Sequence Label: Halo-GPI-RUSH
FEATURES             Location/Qualifiers
     source          1..6235
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        117..496
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        497..700
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     651..671
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     745..764
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        745..763
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     intron          1528..1757
                     /label=chimeric intron
                     /note="chimera between introns from adenovirus and 
                     immunoglobulin heavy chain genes"
     primer_bind     1740..1759
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     misc_feature    join(1829..2403,2404..2406,2407..2415)
                     /label=IRES2
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(1996..2013)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     2223..2242
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     sig_peptide     2429..2488
                     /codon_start=1
                     /product="human interleukin-2 signal sequence"
                     /label=IL-2 signal sequence
                     /translation="MYRMQLLSCIALSLALVTNS"
     CDS             2489..2599
                     /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             2615..3499
                     /codon_start=1
                     /product="modified bacterial dehalogenase that forms 
                     covalent bonds with chloroalkane derivatives"
                     /label=HaloTag(R)
                     /translation="EIGTGFPFDPHYVEVLGERMHYVDVGPRDGTPVLFLHGNPTSSYV
                     WRNIIPHVAPTHRCIAPDLIGMGKSDKPDLGYFFDDHVRFMDAFIEALGLEEVVLVIHD
                     WGSALGFHWAKRNPERVKGIAFMEFIRPIPTWDEWPEFARETFQAFRTTDVGRKLIIDQ
                     NVFIEGTLPMGVVRPLTEVEMDHYREPFLNPVDREPLWRFPNELPIAGEPANIVALVEE
                     YMDWLHQSPVPKLLFWGTPGVLIPPAEAARLAKSLPNCKAVDIGPGLNLLQEDNPDLIG
                     SEIARWLSTLEISG"
     polyA_signal    3737..3858
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3774..3793)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(3960..3976)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3960..3976)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3973..3995)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3984..4000
                     /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(4008..4014,4015..4032,4033..4038))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4053..4074
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4191..4208)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4362..4950)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4442..4461)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5121..5912,5913..5981))
                     /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     5744..5763
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5982..6086)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(6161..6180)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 cgcgagcaaa atttaagcta caacaaggca aggcttgacc gacaattgca tgaagaatct
       61 gcttagggtt aggcgttttg cgctgcttcg cgatgtacgg gccagatata cgcgttgaca
      121 ttgattattg actagttatt aatagtaatc aattacgggg tcattagttc atagcccata
      181 tatggagttc cgcgttacat aacttacggt aaatggcccg cctggctgac cgcccaacga
      241 cccccgccca ttgacgtcaa taatgacgta tgttcccata gtaacgccaa tagggacttt
      301 ccattgacgt caatgggtgg agtatttacg gtaaactgcc cacttggcag tacatcaagt
      361 gtatcatatg ccaagtacgc cccctattga cgtcaatgac ggtaaatggc ccgcctggca
      421 ttatgcccag tacatgacct tatgggactt tcctacttgg cagtacatct acgtattagt
      481 catcgctatt accatggtga tgcggttttg gcagtacatc aatgggcgtg gatagcggtt
      541 tgactcacgg ggatttccaa gtctccaccc cattgacgtc aatgggagtt tgttttggca
      601 ccaaaatcaa cgggactttc caaaatgtcg taacaactcc gccccattga cgcaaatggg
      661 cggtaggcgt gtacggtggg aggtctatat aagcagagct ctctggctaa ctagagaacc
      721 cactgcttac tggcttatcg aaattaatac gactcactat agggagaccc aagcttggta
      781 ccgagctcgg atcgatatct gcggcctagc tagcgcttaa ggcctgttaa ccggtatgga
      841 tgtatgcgtc cgtcttgccc tgtggctcct ctggggactc ctcctgcacc agggccagag
      901 cctcagccat agtcacagtg agaaggcgac aggaaccagc tcgggcgccg gtttaaacga
      961 ccctagcaaa gactcaaaag ctcaggtgtc cgctgccgag gctggcatta ctggaacatg
     1021 gtacaatcag ctcgggagca cctttattgt gactgctgga gccgatggag ccctcaccgg
     1081 aacatacgaa tctgctgtgg gaaacgccga atcacggtac gtcctcactg gccgatacga
     1141 tagtgcccct gccaccgacg gatctgggac tgccctggga tggactgtcg cttggaaaaa
     1201 caactaccgg aatgctcatt ctgccacaac atggagtgga cagtacgtgg gaggcgctga
     1261 ggctagaatc aatacacagt ggctgctcac atctggcaca accgaggcaa atgcttggaa
     1321 atccaccctg gtgggacatg acacattcac caaagtgaaa ccctccgccg cttcaatcga
     1381 tgccgccaaa aaagccggag tcaacaacgg caatcctctg gatgccgtcc agcagctcgg
     1441 accgaaagat gaactgtgag cggccgcata gataactgat ccagtgtgct ggaattaatt
     1501 cgctgtctgc gagggccagc tgttggggtg agtactccct ctcaaaagcg ggcatgactt
     1561 ctgcgctaag attgtcagtt tccaaaaacg aggaggattt gatattcacc tggcccgcgg
     1621 tgatgccttt gagggtggcc gcgtccatct ggtcagaaaa gacaatcttt ttgttgtcaa
     1681 gcttgaggtg tggcaggctt gagatctggc catacacttg agtgacaatg acatccactt
     1741 tgcctttctc tccacaggtg tccactccca ggtccaactg caggtcgagc atgcatctag
     1801 ggcggccaat tccgcccctc tccccccccc cctctccctc ccccccccct aacgttactg
     1861 gccgaagccg cttggaataa ggccggtgtg cgtttgtcta tatgttattt tccaccatat
     1921 tgccgtcttt tggcaatgtg agggcccgga aacctggccc tgtcttcttg acgagcattc
     1981 ctaggggtct ttcccctctc gccaaaggaa tgcaaggtct gttgaatgtc gtgaaggaag
     2041 cagttcctct ggaagcttct tgaagacaaa caacgtctgt agcgaccctt tgcaggcagc
     2101 ggaacccccc acctggcgac aggtgcctct gcggccaaaa gccacgtgta taagatacac
     2161 ctgcaaaggc ggcacaaccc cagtgccacg ttgtgagttg gatagttgtg gaaagagtca
     2221 aatggctctc ctcaagcgta ttcaacaagg ggctgaagga tgcccagaag gtaccccatt
     2281 gtatgggatc tgatctgggg cctcggtgca catgctttac atgtgtttag tcgaggttaa
     2341 aaaaacgtct aggccccccg aaccacgggg acgtggtttt cctttgaaaa acacgatgat
     2401 aagcttgcca caacccggga ggcgcgccat gtacaggatg caactcctgt cttgcattgc
     2461 actaagtctt gcacttgtca cgaattccga cgagaagacc actggttggc gaggtggaca
     2521 cgttgttgaa ggactggctg gggaacttga acaacttcgt gcacgactgg agcatcaccc
     2581 acaaggtcaa cgtgaaccac ctgcaggtgg atccgaaatc ggtactggct ttccattcga
     2641 cccccattat gtggaagtcc tgggcgagcg catgcactac gtcgatgttg gtccgcgcga
     2701 tggcacccct gtgctgttcc tgcacggtaa cccgacctcc tcctacgtgt ggcgcaacat
     2761 catcccgcat gttgcaccga cccatcgctg cattgctcca gacctgatcg gtatgggcaa
     2821 atccgacaaa ccagacctgg gttatttctt cgacgaccac gtccgcttca tggatgcctt
     2881 catcgaagcc ctgggtctgg aagaggtcgt cctggtcatt cacgactggg gctccgctct
     2941 gggtttccac tgggccaagc gcaatccaga gcgcgtcaaa ggtattgcat ttatggagtt
     3001 catccgccct atcccgacct gggacgaatg gccagaattt gcccgcgaga ccttccaggc
     3061 cttccgcacc accgacgtcg gccgcaagct gatcatcgat cagaacgttt ttatcgaggg
     3121 tacgctgccg atgggtgtcg tccgcccgct gactgaagtc gagatggacc attaccgcga
     3181 gccgttcctg aatcctgttg accgcgagcc actgtggcgc ttcccaaacg agctgccaat
     3241 cgccggtgag ccagcgaaca tcgtcgcgct ggtcgaagaa tacatggact ggctgcacca
     3301 gtcccctgtc ccgaagctgc tgttctgggg caccccaggc gttctgatcc caccggccga
     3361 agccgctcgc ctggccaaaa gcctgcctaa ctgcaaggct gtggacatcg gcccgggtct
     3421 gaatctgctg caagaagaca acccggacct gatcggcagc gagatcgcgc gctggctgtc
     3481 tactctggag atttccggcc ggccactgga aaatggcgga acctctctgt ctgaaaaaac
     3541 tgtgctgctg ctggtgactc cctttctggc cgctgcttgg tccctccacc catgaggcct
     3601 cgagggccaa ttaattaact ctagataact gatcataatc agccatacca catttgtaga
     3661 ggttttactt gctttaaaaa acctcccaca cctccccctg aacctgaaac ataaaatgaa
     3721 tgcaattgtt gttgttaact tgtttattgc agcttataat ggttacaaat aaagcaatag
     3781 catcacaaat ttcacaaata aagcattttt ttcactgcat tctagttgtg gtttgtccaa
     3841 actcatcaat gtatcttaac gcgtcgagtg cattctagtt gtggtttgtc caaactcatc
     3901 aatgtatctt atcatgtctg tataccgtcg acctctagct agagcttggc gtaatcatgg
     3961 tcatagctgt ttcctgtgtg aaattgttat ccgctcacaa ttccacacaa catacgagcc
     4021 ggaagcataa agtgtaaagc ctggggtgcc taatgagtga gctaactcac attaattgcg
     4081 ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt gccagctgca ttaatgaatc
     4141 ggccaacgcg cggggagagg cggtttgcgt attgggcgct cttccgcttc ctcgctcact
     4201 gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat cagctcactc aaaggcggta
     4261 atacggttat ccacagaatc aggggataac gcaggaaaga acatgtgagc aaaaggccag
     4321 caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag gctccgcccc
     4381 cctgacgagc atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc gacaggacta
     4441 taaagatacc aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt tccgaccctg
     4501 ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc
     4561 tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac
     4621 gaaccccccg ttcagcccga ccgctgcgcc ttatccggta actatcgtct tgagtccaac
     4681 ccggtaagac acgacttatc gccactggca gcagccactg gtaacaggat tagcagagcg
     4741 aggtatgtag gcggtgctac agagttcttg aagtggtggc ctaactacgg ctacactaga
     4801 agaacagtat ttggtatctg cgctctgctg aagccagtta ccttcggaaa aagagttggt
     4861 agctcttgat ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag
     4921 cagattacgc gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc tacggggtct
     4981 gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt atcaaaaagg
     5041 atcttcacct agatcctttt aaattaaaaa tgaagtttta aatcaatcta aagtatatat
     5101 gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc
     5161 tgtctatttc gttcatccat agttgcctga ctccccgtcg tgtagataac tacgatacgg
     5221 gagggcttac catctggccc cagtgctgca atgataccgc gagacccacg ctcaccggct
     5281 ccagatttat cagcaataaa ccagccagcc ggaagggccg agcgcagaag tggtcctgca
     5341 actttatccg cctccatcca gtctattaat tgttgccggg aagctagagt aagtagttcg
     5401 ccagttaata gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt gtcacgctcg
     5461 tcgtttggta tggcttcatt cagctccggt tcccaacgat caaggcgagt tacatgatcc
     5521 cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag
     5581 ttggccgcag tgttatcact catggttatg gcagcactgc ataattctct tactgtcatg
     5641 ccatccgtaa gatgcttttc tgtgactggt gagtactcaa ccaagtcatt ctgagaatag
     5701 tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac gggataatac cgcgccacat
     5761 agcagaactt taaaagtgct catcattgga aaacgttctt cggggcgaaa actctcaagg
     5821 atcttaccgc tgttgagatc cagttcgatg taacccactc gtgcacccaa ctgatcttca
     5881 gcatctttta ctttcaccag cgtttctggg tgagcaaaaa caggaaggca aaatgccgca
     5941 aaaaagggaa taagggcgac acggaaatgt tgaatactca tactcttcct ttttcaatat
     6001 tattgaagca tttatcaggg ttattgtctc atgagcggat acatatttga atgtatttag
     6061 aaaaataaac aaataggggt tccgcgcaca tttccccgaa aagtgccacc tgacgtcgac
     6121 ggatcgggag atctcccgat cccctatggt gcactctcag tacaatctgc tctgatgccg
     6181 catagttaag ccagtatctg ctccctgctt gtgtgttgga ggtcgctgag tagtg
//
