LOCUS       sequence-478488-        5315 bp    DNA     circular SYN 07-AUG-2025
DEFINITION  Express HiBiT-GPR65 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 5315)
  AUTHORS   Promega
  TITLE     Promega Plasmid Collection
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 5315)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Aug 7, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: 
            "Unpublished"
COMMENT     Sequence Label: HiBiT-GPR65 Vector
FEATURES             Location/Qualifiers
     source          1..5315
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        138..517
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        518..721
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     672..692
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     intron          857..989
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     972..991
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     primer_bind     1033..1052
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1033..1051
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2258..2276)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     polyA_signal    2313..2434
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2324..2343)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     2378..2397
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     promoter        2679..3036
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2679..2870
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2702..2722)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      2887..3022
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     promoter        3051..3098
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             3130..3924
                     /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(3184..3203)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3794..3813
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    3988..4036
                     /note="synthetic polyadenylation signal"
     rep_origin      complement(4275..4863)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4355..4374)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     misc_feature    4979..5262
                     /label=cer region
                     /note="ColE1-derived recombination site that helps to 
                     maintain plasmids as monomers"
ORIGIN
        1 tcaatattgg ccattagcca tattattcat tggttatata gcataaatca atattggcta
       61 ttggccattg catacgttgt atctatatca taatatgtac atttatattg gctcatgtcc
      121 aatatgaccg ccatgttggc attgattatt gactagttat taatagtaat caattacggg
      181 gtcattagtt catagcccat atatggagtt ccgcgttaca taacttacgg taaatggccc
      241 gcctggctga ccgcccaacg acccccgccc attgacgtca ataatgacgt atgttcccat
      301 agtaacgcca atagggactt tccattgacg tcaatgggtg gagtatttac ggtaaactgc
      361 ccacttggca gtacatcaag tgtatcatat gccaagtccg ccccctattg acgtcaatga
      421 cggtaaatgg cccgcctggc attatgccca gtacatgacc ttacgggact ttcctacttg
      481 gcagtacatc tacgtattag tcatcgctat taccatggtg atgcggtttt ggcagtacac
      541 caatgggcgt ggatagcggt ttgactcacg gggatttcca agtctccacc ccattgacgt
      601 caatgggagt ttgttttggc accaaaatca acgggacttt ccaaaatgtc gtaataaccc
      661 cgccccgttg acgcaaatgg gcggtaggcg tgtacggtgg gaggtctata taagcagagc
      721 tggtttagtg aaccgtcaga tcactagaag ctttattgcg gtagtttatc acagttaaat
      781 tgctaacgca gtcagtgctt ctgacacaac agtctcgaac ttaagctgca gaagttggtc
      841 gtgaggcact gggcaggtaa gtatcaaggt tacaagacag gtttaaggag accaatagaa
      901 actgggcttg tcgagacaga gaagactctt gcgtttctga taggcaccta ttggtcttac
      961 tgacatccac tttgcctttc tctccacagg tgtccactcc cagttcaatt acagctctta
     1021 aggctagagt attaatacga ctcactatag ggctagcgct caccatggtg agcggctggc
     1081 ggctgttcaa gaagattagc gggagctccg gtggctcgag cggtgcgatc gccatgaaca
     1141 gcacatgtat tgaagaacag catgacctgg atcactattt gtttcccatt gtttacatct
     1201 ttgtgattat agtcagcatt ccagccaata ttggatctct gtgtgtgtct ttcctgcaag
     1261 caaagaagga aagtgaacta ggaatttacc tcttcagttt gtcactatca gatttactct
     1321 atgcattaac tctcccttta tggattgatt atacttggaa taaagacaac tggactttct
     1381 ctcctgcctt gtgcaaaggg agtgcttttc tcatgtacat gaatttttac agcagcacag
     1441 cattcctcac ctgcattgcc gttgatcggt atttggctgt tgtctaccct ttgaagtttt
     1501 ttttcctaag gacaagaaga tttgcactca tggtcagcct gtccatctgg atattggaaa
     1561 ccatcttcaa tgctgtcatg ttgtgggaag atgaaacagt tgttgaatat tgcgatgccg
     1621 aaaagtctaa ttttacttta tgctatgaca aatacccttt agagaaatgg caaatcaacc
     1681 tcaacttgtt caggacgtgt acaggctatg caataccttt ggtcaccatc ctgatctgca
     1741 accggaaagt ctaccaagct gtgcggcaca ataaagccac ggaaaacaag gaaaagaaga
     1801 gaatcataaa actacttgtc agcatcacag ttacttttgt cttatgcttt actccctttc
     1861 atgtgatgtt gctgattcgc tgcattttag agcatgctgt gaacttcgaa gaccacagca
     1921 attctgggaa gcgaacttac acaatgtata gaatcacggt tgcattaaca agtttaaatt
     1981 gtgttgctga tccaattctg tactgttttg taaccgaaac aggaagatat gatatgtgga
     2041 atatattaaa attctgcact gggaggtgta atacatcaca aagacaaaga aaacgcatac
     2101 tttctgtgtc tacaaaagat actatggaat tagaggtcct tgaggtttaa acgaattcgg
     2161 gctcggtacc cggggatcct ctagagtcga cctgcaggca tgcaagctga tccggctgct
     2221 aacaaagccc gaaaggaagc tgagttggct gctgccaccg ctgagcaata actagcataa
     2281 ccccttgggg cggccgcttc gagcagacat gataagatac attgatgagt ttggacaaac
     2341 cacaactaga atgcagtgaa aaaaatgctt tatttgtgaa atttgtgatg ctattgcttt
     2401 atttgtaacc attataagct gcaataaaca agttaacaac aacaattgca ttcattttat
     2461 gtttcaggtt cagggggaga tgtgggaggt ttttttaagc aagtaaaacc tctacaaatg
     2521 tggtaaaatc gaattttaac aaaatattaa cgcttacaat ttcctgatgc ggtattttct
     2581 ccttacgcat ctgtgcggta tttcacaccg catacgcgga tctgcgcagc accatggcct
     2641 gaaataacct ctgaaagagg aacttggtta ggtaccttct gaggcggaaa gaaccagctg
     2701 tggaatgtgt gtcagttagg gtgtggaaag tccccaggct ccccagcagg cagaagtatg
     2761 caaagcatgc atctcaatta gtcagcaacc aggtgtggaa agtccccagg ctccccagca
     2821 ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa ccatagtccc gcccctaact
     2881 ccgcccatcc cgcccctaac tccgcccagt tccgcccatt ctccgcccca tggctgacta
     2941 atttttttta tttatgcaga ggccgaggcc gcctcggcct ctgagctatt ccagaagtag
     3001 tgaggaggct tttttggagg cctaggcttt tgcaaaaagc ttaattaact gttgacaatt
     3061 aatcatcggc atagtatatc ggcatagtat aatacgacaa ggtgaggaac taaacccagg
     3121 aggcagatca tgattgaaca agatggattg cacgcaggtt ctccggccgc ttgggtggag
     3181 aggctattcg gctatgactg ggcacaacag acaatcggct gctctgatgc cgccgtgttc
     3241 cggctgtcag cgcaggggcg cccggttctt tttgtcaaga ccgacctgtc cggtgccctg
     3301 aatgaactgc aggacgaggc agcgcggcta tcgtggctgg ccacgacggg cgttccttgc
     3361 gcagctgtgc tcgacgttgt cactgaagcg ggaagggact ggctgctatt gggcgaagtg
     3421 ccggggcagg atctcctgtc atctcacctt gctcctgccg agaaagtatc catcatggct
     3481 gatgcaatgc ggcggctgca tacgcttgat ccggctacct gcccattcga ccaccaagcg
     3541 aaacatcgca tcgagcgagc acgtactcgg atggaagccg gtcttgtcga tcaggatgat
     3601 ctggacgaag agcatcaggg gctcgcgcca gccgaactgt tcgccaggct caaggcgcgc
     3661 atgcccgacg gcgaggatct cgtcgtgacc catggcgatg cctgcttgcc gaatatcatg
     3721 gtggaaaatg gccgcttttc tggattcatc gactgtggcc ggctgggtgt ggcggaccgc
     3781 tatcaggaca tagcgttggc tacccgtgat attgctgaag agcttggcgg cgaatgggct
     3841 gaccgcttcc tcgtgcttta cggtatcgcc gctcccgatt cgcagcgcat cgccttctat
     3901 cgccttcttg acgagttctt ctgagcggga ctctggggtt cgaaatgacc gaccaagcga
     3961 cgcccaacct gccatcacga tggccgcaat aaaatatctt tattttcatt acatctgtgt
     4021 gttggttttt tgtgtgaatc gatagcgata aggatcctct ttgcgcttgc gttttccctt
     4081 gtccagatag cccagtagct gacattcatc cggggtcagc accgtttctg cggactggct
     4141 ttctacccgg tatcagctca ctcaaaggcg gtaatacggt tatccacaga atcaggggat
     4201 aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc
     4261 gcgttgctgg cgtttttcca taggctccgc ccccctgacg agcatcacaa aaatcgacgc
     4321 tcaagtcaga ggtggcgaaa cccgacagga ctataaagat accaggcgtt tccccctgga
     4381 agctccctcg tgcgctctcc tgttccgacc ctgccgctta ccggatacct gtccgccttt
     4441 ctcccttcgg gaagcgtggc gctttctcat agctcacgct gtaggtatct cagttcggtg
     4501 taggtcgttc gctccaagct gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc
     4561 gccttatccg gtaactatcg tcttgagtcc aacccggtaa gacacgactt atcgccactg
     4621 gcagcagcca ctggtaacag gattagcaga gcgaggtatg taggcggtgc tacagagttc
     4681 ttgaagtggt ggcctaacta cggctacact agaaggacag tatttggtat ctgcgctctg
     4741 ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa acaaaccacc
     4801 gctggtagcg gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa aaaaggattt
     4861 caagaagatc ctttgatctt ttctacgggg tctgacgctc agtggaacga aaactcacgt
     4921 taagggattt tggtcatgag attatcaaaa aggatcttca cctagatcct tttatagtcc
     4981 ggaaatacag gaacgcacgc tggatggccc ttcgctggga tggtgaaacc atgaaaaatg
     5041 gcagcttcag tggattaagt gggggtaatg tggcctgtac cctctggttg cataggtatt
     5101 catacggtta aaatttatca ggcgcgattg cggcagtttt tcgggtggtt tgttgccatt
     5161 tttacctgtc tgctgccgtg atcgcgctga acgcgtttta gcggtgcgta caattaaggg
     5221 attatggtaa atccacttac tgtctgccct cgtagccatc gagataaacc gcagtactcc
     5281 ggccacgatg cgtccggcgt agaggatcga gatct
//
