LOCUS       sequence-352594-        6893 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian URT vector -pKP496 SacII-ClaI-flag.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6893)
  AUTHORS   Wadas B, Piatkov KI, Brower CS, Varshavsky A
  TITLE     Analyzing N-terminal Arginylation through the Use of Peptide Arrays 
            and Degradation Assays.
  JOURNAL   J Biol Chem. 2016 Sep 30;291(40):20976-20992. doi: 
            10.1074/jbc.M116.747956. Epub 2016 Aug 10.
  PUBMED    27510035
REFERENCE   2  (bases 1 to 6893)
  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: 
            "10.1074/jbc.M116.747956"; journalName: "J Biol Chem"; date: 
            "2016-09-30- 30"; volume: "291"; issue: "40"; pages: "20976-20992"
COMMENT     Sequence Label: pBW359
FEATURES             Location/Qualifiers
     source          1..6893
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        132..511
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        512..715
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     666..686
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     760..779
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        760..778
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             795..818
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             828..1388
                     /codon_start=1
                     /gene="Mus musculus Dhfr"
                     /product="mouse dihydrofolate reductase"
                     /label=DHFR
                     /translation="MVRPLNCIVAVSQNMGIGKNGDLPWPPLRNEFKYFQRMTTTSSVE
                     GKQNLVIMGRKTWFSIPEKNRPLKDRINIVLSRELKEPPRGAHFLAKSLDDALRLIEQP
                     ELASKVDMVWIVGGSSVYQEAMNQPGHLRLFVTRIMQEFESDTFFPEIDLGKYKLLPEY
                     PGVLSEVQEEKGIKYKFEVYEKKD"
     CDS             1395..1421
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             2298..2321
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     promoter        complement(2343..2361)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(2344..2361)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(2381..2398)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2387..2611
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2657..3085
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2744..2763)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2954..2975
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3094..3114)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3099..3428
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3279..3414
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3341..3360
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3495..4289
                     /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(3549..3568)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4159..4178
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4463..4584
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4500..4519)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4554..4573
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4633..4649)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4633..4649)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4646..4668)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4657..4673
                     /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(4681..4687,4688..4705,4706..4711))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4726..4747
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4864..4881)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5035..5623)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5115..5134)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5794..6585,6586..6654))
                     /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     6417..6436
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6655..6759)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(6834..6853)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 ggtcgctgag tagtgcgcga gcaaaattta agctacaaca aggcaaggct tgaccgacaa
       61 ttgcatgaag aatctgctta gggttaggcg ttttgcgctg cttcgcgatg tacgggccag
      121 atatacgcgt tgacattgat tattgactag ttattaatag taatcaatta cggggtcatt
      181 agttcatagc ccatatatgg agttccgcgt tacataactt acggtaaatg gcccgcctgg
      241 ctgaccgccc aacgaccccc gcccattgac gtcaataatg acgtatgttc ccatagtaac
      301 gccaataggg actttccatt gacgtcaatg ggtggagtat ttacggtaaa ctgcccactt
      361 ggcagtacat caagtgtatc atatgccaag tacgccccct attgacgtca atgacggtaa
      421 atggcccgcc tggcattatg cccagtacat gaccttatgg gactttccta cttggcagta
      481 catctacgta ttagtcatcg ctattaccat ggtgatgcgg ttttggcagt acatcaatgg
      541 gcgtggatag cggtttgact cacggggatt tccaagtctc caccccattg acgtcaatgg
      601 gagtttgttt tggcaccaaa atcaacggga ctttccaaaa tgtcgtaaca actccgcccc
      661 attgacgcaa atgggcggta ggcgtgtacg gtgggaggtc tatataagca gagctctctg
      721 gctaactaga gaacccactg cttactggct tatcgaaatt aatacgactc actataggga
      781 gacccaagct tatggactac aaggacgacg atgacaaggg tggatccatg gttcgaccat
      841 tgaactgcat cgtcgccgtg tcccaaaata tggggattgg caagaacgga gacctaccct
      901 ggcctccgct caggaacgag ttcaagtact tccaaagaat gaccacaacc tcttcagtgg
      961 aaggtaaaca gaatctggtg attatgggta ggaaaacctg gttctccatt cctgagaaga
     1021 atcgaccttt aaaggacaga attaatatag ttctcagtag agaactcaaa gaaccaccac
     1081 gaggagctca ttttcttgcc aaaagtttgg atgatgcctt aagacttatt gaacaaccgg
     1141 aattggcaag taaagtagac atggtttgga tagtcggagg cagttctgtt taccaggaag
     1201 ccatgaatca accaggccac ctcagactct ttgtgacaag gatcatgcag gaatttgaaa
     1261 gtgacacgtt tttcccagaa attgatttgg ggaaatataa acttctccca gaatacccag
     1321 gcgtcctctc tgaggtccag gaggaaaaag gcatcaagta taagtttgaa gtctacgaga
     1381 agaaagacgg tacctaccca tacgatgtcc cagattacgc cgctttcctt gcgggtagtg
     1441 ggtcgaccat gcagattttc gtcaagactt tgaccggtaa aaccataaca ttggaagttg
     1501 aatcttccga taccatcgac aacgttaagt cgaaaattca agacaaggaa ggtatccctc
     1561 cagatcaaca aagattgatc tttgccggta ggcagctaga agacggtaga acgctgtctg
     1621 attacaacat tcagaaggag tccaccttac atcttgtgct aaggctccgc ggtggtgaca
     1681 gaggaagcgg cggtctgccg gcttcctgcc tgaggagtgc aaaggacatg aaacatcggc
     1741 tggggttcct gctgcagaag tcagattcct gcgaacacag ttcttcacac agcaagaagg
     1801 acaaagtagt aacttgccag agggtaagcc aagaagaagt caagaaatgg gctgaatcgt
     1861 tggaaaacct gattcaccat gaatgtggac tggcagcttt caaagctttc ctgaagtcgg
     1921 aatacagcga ggagaacatt gacttctgga tcagctgtga ggagtacaag aagatcaagt
     1981 caccttctaa actaagtccc aaagccaaga agatctacaa cgagttcatc tcagtgcaag
     2041 caacaaaaga ggtgaacctg gactcttgca ccagagagga gacaagccgg aacatgttac
     2101 agcccacaat aacctgtttt gatgaagccc agaagaagat tttcaacctg atggaaaagg
     2161 attcctaccg ccgcttcctc aagtctcgat tctatcttga cctgaccaat ccttccagct
     2221 gcggggcaga gaagcagaaa ggagccaaga gttctgcaga ctgcacttcc ctggtctccc
     2281 agtgtgccat cgatatcgac tacaaagacg atgacgacaa aggttaatct agagggccct
     2341 attctatagt gtcacctaaa tgctagagct cgctgatcag cctcgactgt gccttctagt
     2401 tgccagccat ctgttgtttg cccctccccc gtgccttcct tgaccctgga aggtgccact
     2461 cccactgtcc tttcctaata aaatgaggaa attgcatcgc attgtctgag taggtgtcat
     2521 tctattctgg ggggtggggt ggggcaggac agcaaggggg aggattggga agacaatagc
     2581 aggcatgctg gggatgcggt gggctctatg gcttctgagg cggaaagaac cagctggggc
     2641 tctagggggt atccccacgc gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt
     2701 acgcgcagcg tgaccgctac acttgccagc gccctagcgc ccgctccttt cgctttcttc
     2761 ccttcctttc tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg ggggctccct
     2821 ttagggttcc gatttagtgc tttacggcac ctcgacccca aaaaacttga ttagggtgat
     2881 ggttcacgta gtgggccatc gccctgatag acggtttttc gccctttgac gttggagtcc
     2941 acgttcttta atagtggact cttgttccaa actggaacaa cactcaaccc tatctcggtc
     3001 tattcttttg atttataagg gattttgccg atttcggcct attggttaaa aaatgagctg
     3061 atttaacaaa aatttaacgc gaattaattc tgtggaatgt gtgtcagtta gggtgtggaa
     3121 agtccccagg ctccccagca ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa
     3181 ccaggtgtgg aaagtcccca ggctccccag caggcagaag tatgcaaagc atgcatctca
     3241 attagtcagc aaccatagtc ccgcccctaa ctccgcccat cccgccccta actccgccca
     3301 gttccgccca ttctccgccc catggctgac taattttttt tatttatgca gaggccgagg
     3361 ccgcctctgc ctctgagcta ttccagaagt agtgaggagg cttttttgga ggcctaggct
     3421 tttgcaaaaa gctcccggga gcttgtatat ccattttcgg atctgatcaa gagacaggat
     3481 gaggatcgtt tcgcatgatt gaacaagatg gattgcacgc aggttctccg gccgcttggg
     3541 tggagaggct attcggctat gactgggcac aacagacaat cggctgctct gatgccgccg
     3601 tgttccggct gtcagcgcag gggcgcccgg ttctttttgt caagaccgac ctgtccggtg
     3661 ccctgaatga actgcaggac gaggcagcgc ggctatcgtg gctggccacg acgggcgttc
     3721 cttgcgcagc tgtgctcgac gttgtcactg aagcgggaag ggactggctg ctattgggcg
     3781 aagtgccggg gcaggatctc ctgtcatctc accttgctcc tgccgagaaa gtatccatca
     3841 tggctgatgc aatgcggcgg ctgcatacgc ttgatccggc tacctgccca ttcgaccacc
     3901 aagcgaaaca tcgcatcgag cgagcacgta ctcggatgga agccggtctt gtcgatcagg
     3961 atgatctgga cgaagagcat caggggctcg cgccagccga actgttcgcc aggctcaagg
     4021 cgcgcatgcc cgacggcgag gatctcgtcg tgacccatgg cgatgcctgc ttgccgaata
     4081 tcatggtgga aaatggccgc ttttctggat tcatcgactg tggccggctg ggtgtggcgg
     4141 accgctatca ggacatagcg ttggctaccc gtgatattgc tgaagagctt ggcggcgaat
     4201 gggctgaccg cttcctcgtg ctttacggta tcgccgctcc cgattcgcag cgcatcgcct
     4261 tctatcgcct tcttgacgag ttcttctgag cgggactctg gggttcgaaa tgaccgacca
     4321 agcgacgccc aacctgccat cacgagattt cgattccacc gccgccttct atgaaaggtt
     4381 gggcttcgga atcgttttcc gggacgccgg ctggatgatc ctccagcgcg gggatctcat
     4441 gctggagttc ttcgcccacc ccaacttgtt tattgcagct tataatggtt acaaataaag
     4501 caatagcatc acaaatttca caaataaagc atttttttca ctgcattcta gttgtggttt
     4561 gtccaaactc atcaatgtat cttatcatgt ctgtataccg tcgacctcta gctagagctt
     4621 ggcgtaatca tggtcatagc tgtttcctgt gtgaaattgt tatccgctca caattccaca
     4681 caacatacga gccggaagca taaagtgtaa agcctggggt gcctaatgag tgagctaact
     4741 cacattaatt gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt cgtgccagct
     4801 gcattaatga atcggccaac gcgcggggag aggcggtttg cgtattgggc gctcttccgc
     4861 ttcctcgctc actgactcgc tgcgctcggt cgttcggctg cggcgagcgg tatcagctca
     4921 ctcaaaggcg gtaatacggt tatccacaga atcaggggat aacgcaggaa agaacatgtg
     4981 agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca
     5041 taggctccgc ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa
     5101 cccgacagga ctataaagat accaggcgtt tccccctgga agctccctcg tgcgctctcc
     5161 tgttccgacc ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc
     5221 gctttctcat agctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct
     5281 gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg
     5341 tcttgagtcc aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag
     5401 gattagcaga gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta
     5461 cggctacact agaagaacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg
     5521 aaaaagagtt ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt
     5581 tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct caagaagatc ctttgatctt
     5641 ttctacgggg tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgag
     5701 attatcaaaa aggatcttca cctagatcct tttaaattaa aaatgaagtt ttaaatcaat
     5761 ctaaagtata tatgagtaaa cttggtctga cagttaccaa tgcttaatca gtgaggcacc
     5821 tatctcagcg atctgtctat ttcgttcatc catagttgcc tgactccccg tcgtgtagat
     5881 aactacgata cgggagggct taccatctgg ccccagtgct gcaatgatac cgcgagaccc
     5941 acgctcaccg gctccagatt tatcagcaat aaaccagcca gccggaaggg ccgagcgcag
     6001 aagtggtcct gcaactttat ccgcctccat ccagtctatt aattgttgcc gggaagctag
     6061 agtaagtagt tcgccagtta atagtttgcg caacgttgtt gccattgcta caggcatcgt
     6121 ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc ggttcccaac gatcaaggcg
     6181 agttacatga tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt
     6241 tgtcagaagt aagttggccg cagtgttatc actcatggtt atggcagcac tgcataattc
     6301 tcttactgtc atgccatccg taagatgctt ttctgtgact ggtgagtact caaccaagtc
     6361 attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa
     6421 taccgcgcca catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg
     6481 aaaactctca aggatcttac cgctgttgag atccagttcg atgtaaccca ctcgtgcacc
     6541 caactgatct tcagcatctt ttactttcac cagcgtttct gggtgagcaa aaacaggaag
     6601 gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa tgttgaatac tcatactctt
     6661 cctttttcaa tattattgaa gcatttatca gggttattgt ctcatgagcg gatacatatt
     6721 tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc
     6781 acctgacgtc gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc
     6841 tgctctgatg ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt gga
//
