LOCUS       sequence-481950-        5469 bp    DNA     circular SYN 04-AUG-2025
DEFINITION  Express TCEB1-HaloTag Fusion Protein in Mammalian Cells under a CMV 
            promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5469)
  AUTHORS   Promega
  TITLE     Promega Plasmid Collection
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 5469)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Aug 4, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: 
            "Unpublished"
COMMENT     Sequence Label: TCEB1-HaloTag Fusion Vector
FEATURES             Location/Qualifiers
     source          1..5469
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        63..366
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        367..570
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     521..541
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     intron          706..838
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     821..840
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     primer_bind     882..901
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        882..900
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             1271..1291
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="EDLYFQS"
     CDS             1307..2191
                     /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    2278..2399
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2289..2308)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     protein_bind    complement(2514..2547)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     promoter        2557..2604
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             2636..3430
                     /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
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(2690..2709)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3300..3319
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     rep_origin      complement(3588..4176)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3668..3687)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     polyA_signal    4302..4350
                     /note="synthetic polyadenylation signal"
     CDS             complement(4374..4772)
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
                     VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
                     KAIVKDSDGQPTAVGIRELLPSGYVWEG"
     regulatory      4769..4778
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     rep_origin      4817..4951
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     enhancer        4948..5139
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     5096..5116
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     polyA_signal    5218..5299
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
ORIGIN
        1 actagttatt aatagtaatc aattacgggg tcattagttc atagcccata tatggagttc
       61 cgcgttacat aacttacggt aaatggcccg cctggctgac cgcccaacga cccccgccca
      121 ttgacgtcaa taatgacgta tgttcccata gtaacgccaa tagggacttt ccattgacgt
      181 caatgggtgg agtatttacg gtaaactgcc cacttggcag tacatcaagt gtatcatatg
      241 ccaagtccgc cccctattga cgtcaatgac ggtaaatggc ccgcctggca ttatgcccag
      301 tacatgacct tacgggactt tcctacttgg cagtacatct acgtattagt catcgctatt
      361 accatggtga tgcggttttg gcagtacacc aatgggcgtg gatagcggtt tgactcacgg
      421 ggatttccaa gtctccaccc cattgacgtc aatgggagtt tgttttggca ccaaaatcaa
      481 cgggactttc caaaatgtcg taataacccc gccccgttga cgcaaatggg cggtaggcgt
      541 gtacggtggg aggtctatat aagcagagct ggtttagtga accgtcagat cactagaagc
      601 tttattgcgg tagtttatca cagttaaatt gctaacgcag tcagtgcttc tgacacaaca
      661 gtctcgaact taagctgcag aagttggtcg tgaggcactg ggcaggtaag tatcaaggtt
      721 acaagacagg tttaaggaga ccaatagaaa ctgggcttgt cgagacagag aagactcttg
      781 cgtttctgat aggcacctat tggtcttact gacatccact ttgcctttct ctccacaggt
      841 gtccactccc agttcaatta cagctcttaa ggctagagta ttaatacgac tcactatagg
      901 gctagcgatc gccatggatg gagaggagaa aacctatggt ggctgtgaag gacctgatgc
      961 catgtatgtc aaattgatat catctgatgg ccatgaattt attgtaaaaa gagaacatgc
     1021 attaacatca ggcacgataa aagccatgtt gagtggccca ggtcagtttg ctgagaacga
     1081 aaccaatgag gtcaatttta gagagatacc ttcacatgtg ctatcgaaag tatgcatgta
     1141 ttttacgtac aaggttcgct acactaacag ctccaccgag attcctgaat tcccaattgc
     1201 acctgaaatt gcactggaac tgctgatggc tgcgaacttc ttagattgtg tttctctcga
     1261 gccaaccact gaggatctgt actttcagag cgataacgat ggatccgaaa tcggtactgg
     1321 ctttccattc gacccccatt atgtggaagt cctgggcgag cgcatgcact acgtcgatgt
     1381 tggtccgcgc gatggcaccc ctgtgctgtt cctgcacggt aacccgacct cctcctacgt
     1441 gtggcgcaac atcatcccgc atgttgcacc gacccatcgc tgcattgctc cagacctgat
     1501 cggtatgggc aaatccgaca aaccagacct gggttatttc ttcgacgacc acgtccgctt
     1561 catggatgcc ttcatcgaag ccctgggtct ggaagaggtc gtcctggtca ttcacgactg
     1621 gggctccgct ctgggtttcc actgggccaa gcgcaatcca gagcgcgtca aaggtattgc
     1681 atttatggag ttcatccgcc ctatcccgac ctgggacgaa tggccagaat ttgcccgcga
     1741 gaccttccag gccttccgca ccaccgacgt cggccgcaag ctgatcatcg atcagaacgt
     1801 ttttatcgag ggtacgctgc cgatgggtgt cgtccgcccg ctgactgaag tcgagatgga
     1861 ccattaccgc gagccgttcc tgaatcctgt tgaccgcgag ccactgtggc gcttcccaaa
     1921 cgagctgcca atcgccggtg agccagcgaa catcgtcgcg ctggtcgaag aatacatgga
     1981 ctggctgcac cagtcccctg tcccgaagct gctgttctgg ggcaccccag gcgttctgat
     2041 cccaccggcc gaagccgctc gcctggccaa aagcctgcct aactgcaagg ctgtggacat
     2101 cggcccgggt ctgaatctgc tgcaagaaga caacccggac ctgatcggca gcgagatcgc
     2161 gcgctggctg tctactctgg agatttccgg ttaatagaat tctagagtcg acctgcaggc
     2221 atgcaagctg atttggctgc tgccaccgct gagcaaggcc gcttcgagca gacatgataa
     2281 gatacattga tgagtttgga caaaccacaa ctagaatgca gtgaaaaaaa tgctttattt
     2341 gtgaaatttg tgatgctatt gctttatttg taaccattat aagctgcaat aaacaagtta
     2401 acaacaacaa ttgcattcat tttatgtttc aggttcaggg ggagatgtgg gaggtttttt
     2461 taagcaagta aaacctctac aaatgtggta aaatcttaat taagaattct aatataactt
     2521 cgtatagcat acattatacg aagttatgga tccactgttg acaattaatc atcggcatag
     2581 tatatcggca tagtataata cgacaaggtg aggaactaaa cccaggaggc agatcatgat
     2641 tgaacaagat ggattgcacg caggttctcc ggccgcttgg gtggagaggc tattcggcta
     2701 tgactgggca caacagacaa tcggctgctc tgatgccgcc gtgttccggc tgtcagcgca
     2761 ggggcgcccg gttctttttg tcaagaccga cctgtccggt gccctgaatg aactgcagga
     2821 cgaggcagcg cggctatcgt ggctggccac gacgggcgtt ccttgcgcag ctgtgctcga
     2881 cgttgtcact gaagcgggaa gggactggct gctattgggc gaagtgccgg ggcaggatct
     2941 cctgtcatct caccttgctc ctgccgagaa agtatccatc atggctgatg caatgcggcg
     3001 gctgcatacg cttgatccgg ctacctgccc attcgaccac caagcgaaac atcgcatcga
     3061 gcgagcacgt actcggatgg aagccggtct tgtcgatcag gatgatctgg acgaagagca
     3121 tcaggggctc gcgccagccg aactgttcgc caggctcaag gcgcgcatgc ccgacggcga
     3181 ggatctcgtc gtgacccatg gcgatgcctg cttgccgaat atcatggtgg aaaatggccg
     3241 cttttctgga ttcatcgact gtggccggct gggtgtggcg gaccgctatc aggacatagc
     3301 gttggctacc cgtgatattg ctgaagagct tggcggcgaa tgggctgacc gcttcctcgt
     3361 gctttacggt atcgccgctc ccgattcgca gcgcatcgcc ttctatcgcc ttcttgacga
     3421 gttcttctga ttcgaaatga ccgaccaagc gacgcccaac cggtatcagc tcactcaaag
     3481 gcggtaatac ggttatccac agaatcaggg gataacgcag gaaagaacat gtgagcaaaa
     3541 ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc
     3601 cgcccccctg acgagcatca caaaaatcga cgctcaagtc agaggtggcg aaacccgaca
     3661 ggactataaa gataccaggc gtttccccct ggaagctccc tcgtgcgctc tcctgttccg
     3721 accctgccgc ttaccggata cctgtccgcc tttctccctt cgggaagcgt ggcgctttct
     3781 catagctcac gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt
     3841 gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag
     3901 tccaacccgg taagacacga cttatcgcca ctggcagcag ccactggtaa caggattagc
     3961 agagcgaggt atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa ctacggctac
     4021 actagaagga cagtatttgg tatctgcgct ctgctgaagc cagttacctt cggaaaaaga
     4081 gttggtagct cttgatccgg caaacaaacc accgctggta gcggtggttt ttttgtttgc
     4141 aagcagcaga ttacgcgcag aaaaaaagga tttcaagaag atcctttgat cttttctacg
     4201 gggtctgacg ctcagtggaa cgaaaactca cgttaaggga ttttggtcat gagattatca
     4261 aaaaggatct tcacctagat ccttttatag tccggaatcg tcacacaaaa aaccaacaca
     4321 cagatgtaat gaaaataaag atattttatt gcggccatcg tgatggctag agtttagccc
     4381 tcccacacat aaccagaggg cagcaattca cgaatcccaa ctgccgtcgg ctgtccatca
     4441 ctgtccttca ctatggcttt gatcccagga tgcagatcga gaagcacctg tcggcaccgt
     4501 ccgcaggggc tcaagatgcc cctgttctca tttccgatcg cgacgataca agtcaggttg
     4561 ccagctgccg cagcagcagc agtgcccagc accacgagtt ctgcacaagg tcccccagta
     4621 aaatgatata cattgacacc agtgaagatg cggccgtcgc tagagagagc tgcgctggcg
     4681 acgctgtagt cttcagagat ggggatgctg ttgattgtag ccgttgctct ttcaatgagg
     4741 gtggattctt cttgagacaa aggcttggcc atggtggcgc tagtgtcaga agaatcgagc
     4801 tttttgcaaa agcctaggcc tccaaaaaag cctcctcact acttctggaa tagctcagag
     4861 gccgaggcgg cctcggcctc tgcataaata aaaaaaatta gtcagccatg gggcggagaa
     4921 tgggcggaac tgggcggagt taggggcggg actatggttg ctgactaatt gagatgcatg
     4981 ctttgcatac ttctgcctgc tggggagcct ggggactttc cacacctggt tgctgactaa
     5041 ttgagatgca tgctttgcat acttctgcct gctggggagc ctggggactt tccacaccct
     5101 aactgacaca cattccacag ctggttcttt ccgcctcaga aggtacctaa ccaagttcct
     5161 ctttcagagg ttatttcagg ccatggtgct gcgcaagacg cgttaattaa gacgtcggca
     5221 gtgaaaaaaa tgctttattt gtgaaatttg tgatgctatt gctttatttg taaccattat
     5281 aagctgcaat aaacaagtta acaacaacaa ttgcattcat tttatgtttc aggttcaggg
     5341 ggaggtgtgg gaggtttttt aaagcaagta aaacctctac aaatgtggta tggctgatta
     5401 tgatccgcgg ccgcaatacg tcgacgttat cagctgactt cgtacgagag cctaggatta
     5461 tggcgcgcc
//
