LOCUS       sequence-473370-        6612 bp    DNA     circular SYN 28-MAY-2025
DEFINITION  Express HiBit-VPS4A in Mammalian Cells under a CMV promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6612)
  TITLE     Promega Early Access Collection
REFERENCE   2  (bases 1 to 6612)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 28, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"
COMMENT     Sequence Label: HiBit-VPS4A Fusion Vector
FEATURES             Location/Qualifiers
     source          1..6612
                     /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(2558..2576)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     polyA_signal    2613..2734
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2624..2643)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     2678..2697
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     promoter        2979..3336
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2979..3170
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3002..3022)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3187..3322
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3387..4181
                     /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(3441..3460)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4051..4070
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4245..4293
                     /note="synthetic polyadenylation signal"
     promoter        4449..4553
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4554..4622,4623..5414)
                     /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     complement(4772..4791)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      complement(5572..6160)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5652..5671)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     misc_feature    6276..6559
                     /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 gccatgacaa
     1141 cgtcaaccct ccagaaagcc attgatctgg tgacgaaagc cacagaggag gacaaagcca
     1201 agaactacga ggaggcgctg cggctgtacc agcatgcggt ggagtacttc ctccacgcta
     1261 tcaagtatga ggcccacagc gacaaggcca aggagagcat tcgagccaag tgcgtgcagt
     1321 acctagaccg ggccgagaag ctgaaggatt atttacgaag caaagagaaa cacggcaaga
     1381 agccagtcaa agagaaccag agtgagggca agggcagtga cagtgacagt gaaggggata
     1441 atccggagaa aaagaaactg caagaacagc tgatgggtgc cgtcgtgatg gagaagccca
     1501 acatacggtg gaacgacgtg gccgggctgg agggggccaa ggaggccctc aaagaagctg
     1561 tcattttgcc aatcaaattc ccacacttgt tcacaggcaa gcgcaccccc tggcggggga
     1621 ttctgctgtt cggaccccct ggcacaggga aatcctacct ggccaaagcc gtggcaacag
     1681 aggccaacaa ctccaccttc ttctctgtgt cctcctcaga tctgatgtcc aagtggctgg
     1741 gggagagtga aaagctggtc aagaacctgt ttgagctggc caggcagcac aagccctcca
     1801 tcatcttcat cgatgaggtg gattccctct gcgggtcccg aaatgaaaat gagagtgagg
     1861 ccgcccggag gatcaaaacg gagttcttgg tccagatgca gggggtgggg aataacaatg
     1921 atgggactct ggttcttgga gccacaaaca tcccatgggt gttggattcg gccatcagga
     1981 ggaggtttga aaaacgaatt tatatcccct tgccggagga agctgcccgc gcccagatgt
     2041 tccggttgca tctcgggagc actccccaca acctcacgga tgcaaacatc cacgagctgg
     2101 cccggaagac ggaaggctac tcgggcgcgg acatcagcat catcgtgcgg gactctctca
     2161 tgcagcccgt gaggaaggtg cagtcggcca cacacttcaa aaaggtctgt ggcccctctc
     2221 gcaccaaccc cagcatgatg attgatgacc tcctgactcc atgctcacca ggggacccag
     2281 gagccatgga gatgacttgg atggatgtcc ctggggacaa actcttagag cctgtggttt
     2341 gcatgtcgga catgctgcgg tctctggcca ccacccggcc cacggtgaat gcagacgacc
     2401 tcctgaaagt gaagaaattc tcagaggact ttgggcaaga gagtgtttaa acgaattcgg
     2461 gctcggtacc cggggatcct ctagagtcga cctgcaggca tgcaagctga tccggctgct
     2521 aacaaagccc gaaaggaagc tgagttggct gctgccaccg ctgagcaata actagcataa
     2581 ccccttgggg cggccgcttc gagcagacat gataagatac attgatgagt ttggacaaac
     2641 cacaactaga atgcagtgaa aaaaatgctt tatttgtgaa atttgtgatg ctattgcttt
     2701 atttgtaacc attataagct gcaataaaca agttaacaac aacaattgca ttcattttat
     2761 gtttcaggtt cagggggaga tgtgggaggt ttttttaagc aagtaaaacc tctacaaatg
     2821 tggtaaaatc gaattttaac aaaatattaa cgcttacaat ttcctgatgc ggtattttct
     2881 ccttacgcat ctgtgcggta tttcacaccg catacgcgga tctgcgcagc accatggcct
     2941 gaaataacct ctgaaagagg aacttggtta ggtaccttct gaggcggaaa gaaccagctg
     3001 tggaatgtgt gtcagttagg gtgtggaaag tccccaggct ccccagcagg cagaagtatg
     3061 caaagcatgc atctcaatta gtcagcaacc aggtgtggaa agtccccagg ctccccagca
     3121 ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa ccatagtccc gcccctaact
     3181 ccgcccatcc cgcccctaac tccgcccagt tccgcccatt ctccgcccca tggctgacta
     3241 atttttttta tttatgcaga ggccgaggcc gcctcggcct ctgagctatt ccagaagtag
     3301 tgaggaggct tttttggagg cctaggcttt tgcaaaaagc ttgattcttc tgacacaaca
     3361 gtctcgaact taaggctaga gccaccatga ttgaacaaga tggattgcac gcaggttctc
     3421 cggccgcttg ggtggagagg ctattcggct atgactgggc acaacagaca atcggctgct
     3481 ctgatgccgc cgtgttccgg ctgtcagcgc aggggcgccc ggttcttttt gtcaagaccg
     3541 acctgtccgg tgccctgaat gaactgcagg acgaggcagc gcggctatcg tggctggcca
     3601 cgacgggcgt tccttgcgca gctgtgctcg acgttgtcac tgaagcggga agggactggc
     3661 tgctattggg cgaagtgccg gggcaggatc tcctgtcatc tcaccttgct cctgccgaga
     3721 aagtatccat catggctgat gcaatgcggc ggctgcatac gcttgatccg gctacctgcc
     3781 cattcgacca ccaagcgaaa catcgcatcg agcgagcacg tactcggatg gaagccggtc
     3841 ttgtcgatca ggatgatctg gacgaagagc atcaggggct cgcgccagcc gaactgttcg
     3901 ccaggctcaa ggcgcgcatg cccgacggcg aggatctcgt cgtgacccat ggcgatgcct
     3961 gcttgccgaa tatcatggtg gaaaatggcc gcttttctgg attcatcgac tgtggccggc
     4021 tgggtgtggc ggaccgctat caggacatag cgttggctac ccgtgatatt gctgaagagc
     4081 ttggcggcga atgggctgac cgcttcctcg tgctttacgg tatcgccgct cccgattcgc
     4141 agcgcatcgc cttctatcgc cttcttgacg agttcttctg agcgggactc tggggttcga
     4201 aatgaccgac caagcgacgc ccaacctgcc atcacgatgg ccgcaataaa atatctttat
     4261 tttcattaca tctgtgtgtt ggttttttgt gtgaatcgat agcgataagg atcctctttg
     4321 cgcttgcgtt ttcccttgtc cagatagccc agtagctgac attcatccgg ggtcagcacc
     4381 gtttctgcgg actggctttc tacgtaatgg tttcttagac gtcaggtggc acttttcggg
     4441 gaaatgtgcg cggaacccct atttgtttat ttttctaaat acattcaaat atgtatccgc
     4501 tcatgagaca ataaccctga taaatgcttc aataatattg aaaaaggaag agtatgagta
     4561 ttcaacattt ccgtgtcgcc cttattccct tttttgcggc attttgcctt cctgtttttg
     4621 ctcacccaga aacgctggtg aaagtaaaag atgctgaaga tcagttgggt gcacgagtgg
     4681 gttacatcga actggatctc aacagcggta agatccttga gagttttcgc cccgaagaac
     4741 gttttccaat gatgagcact ttcaaagttc tgctatgtgg cgcggtatta tcccgtattg
     4801 acgccgggca agagcaactc ggtcgccgca tacactattc tcagaatgac ttggttgagt
     4861 actcaccagt cacagaaaag catcttacgg atggcatgac agtaagagaa ttatgcagtg
     4921 ctgccataac catgagtgat aacactgcgg ccaacttact tctgacaact atcggaggac
     4981 cgaaggagct aaccgctttt ttgcacaaca tgggggatca tgtaactcgc cttgatcgtt
     5041 gggaaccgga gctgaatgaa gccataccaa acgacgagcg tgacaccacg atgcctgtag
     5101 caatggcaac aacgttgcgc aaactattaa ctggcgaact acttactcta gcttcccggc
     5161 aacaattaat agactggatg gaggcggata aagttgcagg accacttctg cgctcggccc
     5221 ttccggctgg ctggtttatt gctgataaat ctggagccgg tgagcgtggg tctcgcggta
     5281 tcattgcagc actggggcca gatggtaagc cctcccgtat cgtagttatc tacacgacgg
     5341 ggagtcaggc aactatggat gaacgaaata gacagatcgc tgagataggt gcctcactga
     5401 ttaagcattg gtaattcgaa atgaccgacc aagcgacgcc caaccggtat cagctcactc
     5461 aaaggcggta atacggttat ccacagaatc aggggataac gcaggaaaga acatgtgagc
     5521 aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag
     5581 gctccgcccc cctgacgagc atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc
     5641 gacaggacta taaagatacc aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt
     5701 tccgaccctg ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct
     5761 ttctcatagc tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg
     5821 ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc ttatccggta actatcgtct
     5881 tgagtccaac ccggtaagac acgacttatc gccactggca gcagccactg gtaacaggat
     5941 tagcagagcg aggtatgtag gcggtgctac agagttcttg aagtggtggc ctaactacgg
     6001 ctacactaga aggacagtat ttggtatctg cgctctgctg aagccagtta ccttcggaaa
     6061 aagagttggt agctcttgat ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt
     6121 ttgcaagcag cagattacgc gcagaaaaaa aggatttcaa gaagatcctt tgatcttttc
     6181 tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt
     6241 atcaaaaagg atcttcacct agatcctttt atagtccgga aatacaggaa cgcacgctgg
     6301 atggcccttc gctgggatgg tgaaaccatg aaaaatggca gcttcagtgg attaagtggg
     6361 ggtaatgtgg cctgtaccct ctggttgcat aggtattcat acggttaaaa tttatcaggc
     6421 gcgattgcgg cagtttttcg ggtggtttgt tgccattttt acctgtctgc tgccgtgatc
     6481 gcgctgaacg cgttttagcg gtgcgtacaa ttaagggatt atggtaaatc cacttactgt
     6541 ctgccctcgt agccatcgag ataaaccgca gtactccggc cacgatgcgt ccggcgtaga
     6601 ggatcgagat ct
//
