LOCUS       sequence-328224-        8380 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  RUSH cargo: Str-KDEL_IRES_Transferrin Receptor (TfR)-SBP-Halo.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8380)
  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 8380)
  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-TfR-RUSH
FEATURES             Location/Qualifiers
     source          1..8380
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(92..109)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     319..338
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             complement(1464..1475)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     CDS             2814..2927
                     /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             2943..3827
                     /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    3978..4099
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4015..4034)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(4201..4217)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4201..4217)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4214..4236)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4225..4241
                     /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(4249..4255,4256..4273,4274..4279))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4294..4315
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4432..4449)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4603..5191)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4683..4702)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5362..6153,6154..6222))
                     /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     5985..6004
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6223..6327)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(6402..6421)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        6593..6972
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        6973..7176
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     7127..7147
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     7221..7240
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        7221..7239
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     intron          8004..8233
                     /label=chimeric intron
                     /note="chimera between introns from adenovirus and 
                     immunoglobulin heavy chain genes"
     primer_bind     8216..8235
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     misc_feature    join(8305..8380,1..499,500..502,503..511)
                     /label=IRES2
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
ORIGIN
        1 ttattttcca ccatattgcc gtcttttggc aatgtgaggg cccggaaacc tggccctgtc
       61 ttcttgacga gcattcctag gggtctttcc cctctcgcca aaggaatgca aggtctgttg
      121 aatgtcgtga aggaagcagt tcctctggaa gcttcttgaa gacaaacaac gtctgtagcg
      181 accctttgca ggcagcggaa ccccccacct ggcgacaggt gcctctgcgg ccaaaagcca
      241 cgtgtataag atacacctgc aaaggcggca caaccccagt gccacgttgt gagttggata
      301 gttgtggaaa gagtcaaatg gctctcctca agcgtattca acaaggggct gaaggatgcc
      361 cagaaggtac cccattgtat gggatctgat ctggggcctc ggtgcacatg ctttacatgt
      421 gtttagtcga ggttaaaaaa acgtctaggc cccccgaacc acggggacgt ggttttcctt
      481 tgaaaaacac gatgataagc ttgccacaac ccgggaggcg cgccatgatg gatcaagcta
      541 gatcagcatt ctctaacttg tttggtggag aaccattgtc atatacccgg ttcagcctgg
      601 ctcggcaagt agatggcgat aacagtcatg tggagatgaa acttgctgta gatgaagaag
      661 aaaatgctga caataacaca aaggccaatg tcacaaaacc aaaaaggtgt agtggaagta
      721 tctgctatgg gactattgct gtgatcgtct ttttcttgat tggatttatg attggctact
      781 tgggctattg taaaggggta gaaccaaaaa ctgagtgtga gagactggca ggaaccgagt
      841 ctccagtgag ggaggagcca ggagaggact tccctgcagc acgtcgctta tattgggatg
      901 acctgaagag aaagttgtcg gagaaactgg acagcacaga cttcaccagc accatcaagc
      961 tgctgaatga aaattcatat gtccctcgtg aggctggatc tcaaaaagat gaaaatcttg
     1021 cgttgtatgt tgaaaatcaa tttcgtgaat ttaaactcag caaagtctgg cgtgatcaac
     1081 attttgttaa gattcaggtc aaagacagcg ctcaaaactc ggtgatcata gttgataaga
     1141 acggtagact tgtttacctg gtggagaatc ctgggggtta tgtggcgtat agtaaggctg
     1201 caacagttac tggtaaactg gtccatgcta attttggtac taaaaaagat tttgaggatt
     1261 tatacactcc tgtgaatgga tctatagtga ttgtcagagc agggaaaatc acctttgcag
     1321 aaaaggttgc aaatgctgaa agcttaaatg caattggtgt gttgatatac atggaccaga
     1381 ctaaatttcc cattgttaac gcagaacttt cattctttgg acatgctcat ctggggacag
     1441 gtgaccctta cacacctgga ttcccttcct tcaatcacac tcagtttcca ccatctcggt
     1501 catcaggatt gcctaatata cctgtccaga caatctccag agctgctgca gaaaagctgt
     1561 ttgggaatat ggaaggagac tgtccctctg actggaaaac agactctaca tgtaggatgg
     1621 taacctcaga aagcaagaat gtgaagctca ctgtgagcaa tgtgctgaaa gagataaaaa
     1681 ttcttaacat ctttggagtt attaaaggct ttgtagaacc agatcactat gttgtagttg
     1741 gggcccagag agatgcatgg ggccctggag ctgcaaaatc cggtgtaggc acagctctcc
     1801 tattgaaact tgcccagatg ttctcagata tggtcttaaa agatgggttt cagcccagca
     1861 gaagcattat ctttgccagt tggagtgctg gagactttgg atcggttggt gccactgaat
     1921 ggctagaggg atacctttcg tccctgcatt taaaggcttt cacttatatt aatctggata
     1981 aagcggttct tggtaccagc aacttcaagg tttctgccag cccactgttg tatacgctta
     2041 ttgagaaaac aatgcaaaat gtgaagcatc cggttactgg gcaatttcta tatcaggaca
     2101 gcaactgggc cagcaaagtt gagaaactca ctttagacaa tgctgctttc cctttccttg
     2161 catattctgg aatcccagca gtttctttct gtttttgcga ggacacagat tatccttatt
     2221 tgggtaccac catggacacc tataaggaac tgattgagag gattcctgag ttgaacaaag
     2281 tggcacgagc agctgcagag gtcgctggtc agttcgtgat taaactaacc catgatgttg
     2341 aattgaacct ggactatgag aggtacaaca gccaactgct ttcatttgtg agggatctga
     2401 accaatacag agcagacata aaggaaatgg gcctgagttt acagtggctg tattctgctc
     2461 gtggagactt cttccgtgct acttccagac taacaacaga tttcgggaat gctgagaaaa
     2521 cagacagatt tgtcatgaag aaactcaatg atcgtgtcat gagagtggag tatcacttcc
     2581 tctctcccta cgtatctcca aaagagtctc ctttccgaca tgtcttctgg ggctccggct
     2641 ctcacacgct gccagcttta ctggagaact tgaaactgcg taaacaaaat aacggtgctt
     2701 ttaatgaaac gctgttcaga aaccagttgg ctctagctac ttggactatt cagggagctg
     2761 caaatgccct ctctggtgac gtttgggaca ttgacaatga gtttctgaat tccatggacg
     2821 agaagaccac tggttggcga ggtggacacg ttgttgaagg actggctggg gaacttgaac
     2881 aacttcgtgc acgactggag catcacccac aaggtcaacg tgaaccacct gcaggtggat
     2941 ccgaaatcgg tactggcttt ccattcgacc cccattatgt ggaagtcctg ggcgagcgca
     3001 tgcactacgt cgatgttggt ccgcgcgatg gcacccctgt gctgttcctg cacggtaacc
     3061 cgacctcctc ctacgtgtgg cgcaacatca tcccgcatgt tgcaccgacc catcgctgca
     3121 ttgctccaga cctgatcggt atgggcaaat ccgacaaacc agacctgggt tatttcttcg
     3181 acgaccacgt ccgcttcatg gatgccttca tcgaagccct gggtctggaa gaggtcgtcc
     3241 tggtcattca cgactggggc tccgctctgg gtttccactg ggccaagcgc aatccagagc
     3301 gcgtcaaagg tattgcattt atggagttca tccgccctat cccgacctgg gacgaatggc
     3361 cagaatttgc ccgcgagacc ttccaggcct tccgcaccac cgacgtcggc cgcaagctga
     3421 tcatcgatca gaacgttttt atcgagggta cgctgccgat gggtgtcgtc cgcccgctga
     3481 ctgaagtcga gatggaccat taccgcgagc cgttcctgaa tcctgttgac cgcgagccac
     3541 tgtggcgctt cccaaacgag ctgccaatcg ccggtgagcc agcgaacatc gtcgcgctgg
     3601 tcgaagaata catggactgg ctgcaccagt cccctgtccc gaagctgctg ttctggggca
     3661 ccccaggcgt tctgatccca ccggccgaag ccgctcgcct ggccaaaagc ctgcctaact
     3721 gcaaggctgt ggacatcggc ccgggtctga atctgctgca agaagacaac ccggacctga
     3781 tcggcagcga gatcgcgcgc tggctgtcta ctctggagat ttccggccgg ccttaaggcc
     3841 tcgagggcca attaattaac tctagataac tgatcataat cagccatacc acatttgtag
     3901 aggttttact tgctttaaaa aacctcccac acctccccct gaacctgaaa cataaaatga
     3961 atgcaattgt tgttgttaac ttgtttattg cagcttataa tggttacaaa taaagcaata
     4021 gcatcacaaa tttcacaaat aaagcatttt tttcactgca ttctagttgt ggtttgtcca
     4081 aactcatcaa tgtatcttaa cgcgtcgagt gcattctagt tgtggtttgt ccaaactcat
     4141 caatgtatct tatcatgtct gtataccgtc gacctctagc tagagcttgg cgtaatcatg
     4201 gtcatagctg tttcctgtgt gaaattgtta tccgctcaca attccacaca acatacgagc
     4261 cggaagcata aagtgtaaag cctggggtgc ctaatgagtg agctaactca cattaattgc
     4321 gttgcgctca ctgcccgctt tccagtcggg aaacctgtcg tgccagctgc attaatgaat
     4381 cggccaacgc gcggggagag gcggtttgcg tattgggcgc tcttccgctt cctcgctcac
     4441 tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta tcagctcact caaaggcggt
     4501 aatacggtta tccacagaat caggggataa cgcaggaaag aacatgtgag caaaaggcca
     4561 gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg tttttccata ggctccgccc
     4621 ccctgacgag catcacaaaa atcgacgctc aagtcagagg tggcgaaacc cgacaggact
     4681 ataaagatac caggcgtttc cccctggaag ctccctcgtg cgctctcctg ttccgaccct
     4741 gccgcttacc ggatacctgt ccgcctttct cccttcggga agcgtggcgc tttctcatag
     4801 ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca
     4861 cgaacccccc gttcagcccg accgctgcgc cttatccggt aactatcgtc ttgagtccaa
     4921 cccggtaaga cacgacttat cgccactggc agcagccact ggtaacagga ttagcagagc
     4981 gaggtatgta ggcggtgcta cagagttctt gaagtggtgg cctaactacg gctacactag
     5041 aagaacagta tttggtatct gcgctctgct gaagccagtt accttcggaa aaagagttgg
     5101 tagctcttga tccggcaaac aaaccaccgc tggtagcggt ggtttttttg tttgcaagca
     5161 gcagattacg cgcagaaaaa aaggatctca agaagatcct ttgatctttt ctacggggtc
     5221 tgacgctcag tggaacgaaa actcacgtta agggattttg gtcatgagat tatcaaaaag
     5281 gatcttcacc tagatccttt taaattaaaa atgaagtttt aaatcaatct aaagtatata
     5341 tgagtaaact tggtctgaca gttaccaatg cttaatcagt gaggcaccta tctcagcgat
     5401 ctgtctattt cgttcatcca tagttgcctg actccccgtc gtgtagataa ctacgatacg
     5461 ggagggctta ccatctggcc ccagtgctgc aatgataccg cgagacccac gctcaccggc
     5521 tccagattta tcagcaataa accagccagc cggaagggcc gagcgcagaa gtggtcctgc
     5581 aactttatcc gcctccatcc agtctattaa ttgttgccgg gaagctagag taagtagttc
     5641 gccagttaat agtttgcgca acgttgttgc cattgctaca ggcatcgtgg tgtcacgctc
     5701 gtcgtttggt atggcttcat tcagctccgg ttcccaacga tcaaggcgag ttacatgatc
     5761 ccccatgttg tgcaaaaaag cggttagctc cttcggtcct ccgatcgttg tcagaagtaa
     5821 gttggccgca gtgttatcac tcatggttat ggcagcactg cataattctc ttactgtcat
     5881 gccatccgta agatgctttt ctgtgactgg tgagtactca accaagtcat tctgagaata
     5941 gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata cgggataata ccgcgccaca
     6001 tagcagaact ttaaaagtgc tcatcattgg aaaacgttct tcggggcgaa aactctcaag
     6061 gatcttaccg ctgttgagat ccagttcgat gtaacccact cgtgcaccca actgatcttc
     6121 agcatctttt actttcacca gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc
     6181 aaaaaaggga ataagggcga cacggaaatg ttgaatactc atactcttcc tttttcaata
     6241 ttattgaagc atttatcagg gttattgtct catgagcgga tacatatttg aatgtattta
     6301 gaaaaataaa caaatagggg ttccgcgcac atttccccga aaagtgccac ctgacgtcga
     6361 cggatcggga gatctcccga tcccctatgg tgcactctca gtacaatctg ctctgatgcc
     6421 gcatagttaa gccagtatct gctccctgct tgtgtgttgg aggtcgctga gtagtgcgcg
     6481 agcaaaattt aagctacaac aaggcaaggc ttgaccgaca attgcatgaa gaatctgctt
     6541 agggttaggc gttttgcgct gcttcgcgat gtacgggcca gatatacgcg ttgacattga
     6601 ttattgacta gttattaata gtaatcaatt acggggtcat tagttcatag cccatatatg
     6661 gagttccgcg ttacataact tacggtaaat ggcccgcctg gctgaccgcc caacgacccc
     6721 cgcccattga cgtcaataat gacgtatgtt cccatagtaa cgccaatagg gactttccat
     6781 tgacgtcaat gggtggagta tttacggtaa actgcccact tggcagtaca tcaagtgtat
     6841 catatgccaa gtacgccccc tattgacgtc aatgacggta aatggcccgc ctggcattat
     6901 gcccagtaca tgaccttatg ggactttcct acttggcagt acatctacgt attagtcatc
     6961 gctattacca tggtgatgcg gttttggcag tacatcaatg ggcgtggata gcggtttgac
     7021 tcacggggat ttccaagtct ccaccccatt gacgtcaatg ggagtttgtt ttggcaccaa
     7081 aatcaacggg actttccaaa atgtcgtaac aactccgccc cattgacgca aatgggcggt
     7141 aggcgtgtac ggtgggaggt ctatataagc agagctctct ggctaactag agaacccact
     7201 gcttactggc ttatcgaaat taatacgact cactataggg agacccaagc ttggtaccga
     7261 gctcggatcg atatctgcgg cctagctagc gcttaaggcc tgttaaccgg tatggatgta
     7321 tgcgtccgtc ttgccctgtg gctcctctgg ggactcctcc tgcaccaggg ccagagcctc
     7381 agccatagtc acagtgagaa ggcgacagga accagctcgg gcgccggttt aaacgaccct
     7441 agcaaagact caaaagctca ggtgtccgct gccgaggctg gcattactgg aacatggtac
     7501 aatcagctcg ggagcacctt tattgtgact gctggagccg atggagccct caccggaaca
     7561 tacgaatctg ctgtgggaaa cgccgaatca cggtacgtcc tcactggccg atacgatagt
     7621 gcccctgcca ccgacggatc tgggactgcc ctgggatgga ctgtcgcttg gaaaaacaac
     7681 taccggaatg ctcattctgc cacaacatgg agtggacagt acgtgggagg cgctgaggct
     7741 agaatcaata cacagtggct gctcacatct ggcacaaccg aggcaaatgc ttggaaatcc
     7801 accctggtgg gacatgacac attcaccaaa gtgaaaccct ccgccgcttc aatcgatgcc
     7861 gccaaaaaag ccggagtcaa caacggcaat cctctggatg ccgtccagca gctcggaccg
     7921 aaagatgaac tgtgagcggc cgcatagata actgatccag tgtgctggaa ttaattcgct
     7981 gtctgcgagg gccagctgtt ggggtgagta ctccctctca aaagcgggca tgacttctgc
     8041 gctaagattg tcagtttcca aaaacgagga ggatttgata ttcacctggc ccgcggtgat
     8101 gcctttgagg gtggccgcgt ccatctggtc agaaaagaca atctttttgt tgtcaagctt
     8161 gaggtgtggc aggcttgaga tctggccata cacttgagtg acaatgacat ccactttgcc
     8221 tttctctcca caggtgtcca ctcccaggtc caactgcagg tcgagcatgc atctagggcg
     8281 gccaattccg cccctctccc cccccccctc tccctccccc ccccctaacg ttactggccg
     8341 aagccgcttg gaataaggcc ggtgtgcgtt tgtctatatg
//
