LOCUS       sequence-420414-        8921 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  mammalian expression of human RPA32 N-terminally fused to mVenus and
            RPA70 and RPA14.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8921)
  TITLE     Pospiech Lab Plasmids 2023
REFERENCE   2  (bases 1 to 8921)
  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"
COMMENT     Sequence Label: pRPA70-IRES-Ven-RPA32-IRES-RPA14
FEATURES             Location/Qualifiers
     source          1..8921
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        69..372
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        373..576
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     527..547
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     CDS             join(3071..3073,3074..3076,3077..3787)
                     /codon_start=1
                     /product="Venus YFP with monomerizing A206K mutation (Nagai
                     et al., 2002; Kremers et al., 2006)"
                     /label=mVenus
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KLICTTGKLPVPWPTLVTTLGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYITADKQKNGIK
                     ANFKIRHNIEDGGVQLADHYQQNTPIGDGPVLLPDNHYLSYQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(3116..3137)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(3377..3396)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3724..3745
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    5717..5838
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5754..5773)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5808..5827
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(5845..6300)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5982..6003)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     6194..6213
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        6327..6431
                     /gene="bla"
                     /label=AmpR promoter
     promoter        6433..6790
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        6433..6624
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(6456..6476)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      6641..6776
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             6825..7619
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             6825..7619
                     /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
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(6879..6898)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     7489..7508
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    7851..7898
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      8227..8815
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     8716..8735
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 ccatgcatta gttattaata gtaatcaatt acggggtcat tagttcatag cccatatatg
       61 gagttccgcg ttacataact tacggtaaat ggcccgcctg gctgaccgcc caacgacccc
      121 cgcccattga cgtcaataat gacgtatgtt cccatagtaa cgccaatagg gactttccat
      181 tgacgtcaat gggtggagta tttacggtaa actgcccact tggcagtaca tcaagtgtat
      241 catatgccaa gtacgccccc tattgacgtc aatgacggta aatggcccgc ctggcattat
      301 gcccagtaca tgaccttatg ggactttcct acttggcagt acatctacgt attagtcatc
      361 gctattacca tggtgatgcg gttttggcag tacatcaatg ggcgtggata gcggtttgac
      421 tcacggggat ttccaagtct ccaccccatt gacgtcaatg ggagtttgtt ttggcaccaa
      481 aatcaacggg actttccaaa atgtcgtaac aactccgccc cattgacgca aatgggcggt
      541 aggcgtgtac ggtgggaggt ctatataagc agagctggtt tagtgaaccg tcagatccgc
      601 tagcgctacc ggactcagat ctcgagctcg tatggtcggc cagctgagcg agggggccat
      661 tgcggccatc atgcagaagg gggatacaaa cataaagccc atcctccaag tcatcaacat
      721 ccgtcccatt actacgggga atagtccgcc gcgttatcga ctgctcatga gtgatggatt
      781 gaacactcta tcctctttca tgttggcgac acagttgaac cctctcgtgg aggaagaaca
      841 attgtccagc aactgtgtat gccagattca cagatttatt gtgaacactc tgaaagacgg
      901 aaggagagta gttatcttga tggaattaga agttttgaag tcagctgaag cagttggagt
      961 gaagattggc aatccagtgc cctataatga aggactcggg cagccgcaag tagctcctcc
     1021 agcgccagca gccagcccag cagcaagcag caggccccag ccgcagaatg gaagctcggg
     1081 aatgggttct actgtttcta aggcttatgg tgcttcaaag acatttggaa aagctgcagg
     1141 tcccagcctg tcacacactt ctgggggaac acagtccaaa gtggtgccca ttgccagcct
     1201 cactccttac cagtccaagt ggaccatttg tgctcgtgtt accaacaaaa gtcagatccg
     1261 tacctggagc aactcccgag gggaagggaa gcttttctcc ctagaactgg ttgacgaaag
     1321 tggtgaaatc cgagctacag ctttcaatga gcaagtggac aagttctttc ctcttattga
     1381 agtgaacaag gtgtattatt tctcgaaagg caccctgaag attgctaaca agcagttcac
     1441 agctgttaaa aatgactacg agatgacctt caataacgag acttccgtca tgccctgtga
     1501 ggacgaccat catttaccta cggttcagtt tgatttcacg gggattgatg acctcgagaa
     1561 caagtcgaaa gactcacttg tagacatcat cgggatctgc aagagctatg aagacgccac
     1621 taaaatcaca gtgaggtcta acaacagaga agttgccaag aggaatatct acttgatgga
     1681 cacatctggg aaggtggtga ctgctacact gtggggggaa gatgctgata aatttgatgg
     1741 ttctagacag cccgtgttgg ctatcaaagg agcccgagtc tctgatttcg gtggacggag
     1801 cctctccgtg ctgtcttcaa gcactatcat tgcgaatcct gacatcccag aggcctataa
     1861 gcttcgtgga tggtttgacg cagaaggaca agccttagat ggtgtttcca tctctgatct
     1921 aaagagcggc ggagtcggag ggagtaacac caactggaaa accttgtatg aggtcaaatc
     1981 cgagaacctg ggccaaggcg acaagccgga ctactttagt tctgtggcca cagtggtgta
     2041 tcttcgcaaa gagaactgca tgtaccaagc ctgcccgact caggactgca ataagaaagt
     2101 gattgatcaa cagaatggat tgtaccgctg tgagaagtgc gacaccgaat ttcccaattt
     2161 caagtaccgc atgatcctgt cagtaaatat tgcagatttt caagagaatc agtgggtgac
     2221 ttgtttccag gagtctgctg aagctatcct tggacaaaat gctgcttatc ttggggaatt
     2281 aaaagacaag aatgaacagg catttgaaga agttttccag aatgccaact tccgatcttt
     2341 catattcaga gtcagggtca aagtggagac ctacaacgac gagtctcgaa ttaaggccac
     2401 tgtgatggac gtgaagcccg tggactacag agagtatggc cgaaggctgg tcatgagcat
     2461 caggagaagt gcattgatgt gaggatcttt cgcccctctc cctccccccc ccctaacgtt
     2521 actggccgaa gccgcttgga ataaggccgg tgtgcgtttg tctatatgtt attttccacc
     2581 atattgccgt cttttggcaa tgtgagggcc cggaaacctg gccctgtctt cttgacgagc
     2641 attcctaggg gtctttcccc tctcgccaaa ggaatgcaag gtctgttgaa tgtcgtgaag
     2701 gaagcagttc ctctggaagc ttcttgaaga caaacaacgt ctgtagcgac cctttgcagg
     2761 cagcggaacc ccccacctgg cgacaggtgc ctctgcggcc aaaagccacg tgtataagat
     2821 acacctgcaa aggcggcaca accccagtgc cacgttgtga gttggatagt tgtggaaaga
     2881 gtcaaatggc tctcctcaag cgtattcaac aaggggctga aggatgccca gaaggtaccc
     2941 cattgtatgg gatctgatct ggggcctcgg tgcacatgct ttacatgtgt ttagtcgagg
     3001 ttaaaaaaac gtctaggccc cccgaaccac ggggacgtgg ttttcctttg aaaaacacga
     3061 taagaattgc atggtgagca agggcgagga gctgttcacc ggggtggtgc ccatcctggt
     3121 cgagctggac ggcgacgtaa acggccacaa gttcagcgtg tccggcgagg gcgagggcga
     3181 tgccacctac ggcaagctga ccctgaagct gatctgcacc accggcaagc tgcccgtgcc
     3241 ctggcccacc ctcgtgacca ccctgggcta cggcctgcag tgcttcgccc gctaccccga
     3301 ccacatgaag cagcacgact tcttcaagtc cgccatgccc gaaggctacg tccaggagcg
     3361 caccatcttc ttcaaggacg acggcaacta caagacccgc gccgaggtga agttcgaggg
     3421 cgacaccctg gtgaaccgca tcgagctgaa gggcatcgac ttcaaggagg acggcaacat
     3481 cctggggcac aagctggagt acaactacaa cagccacaac gtctatatca ccgccgacaa
     3541 gcagaagaac ggcatcaagg ccaacttcaa gatccgccac aacatcgagg acggcggcgt
     3601 gcagctcgcc gaccactacc agcagaacac ccccatcggc gacggccccg tgctgctgcc
     3661 cgacaaccac tacctgagct accagtccaa gctgagcaaa gaccccaacg agaagcgcga
     3721 tcacatggtc ctgctggagt tcgtgaccgc cgccgggatc actctcggca tggacgagct
     3781 gtacaagtcc ggactcagat ctcgagctca agcttcgaat tctgccacca tgtggaacag
     3841 tggattcgaa agctatggca gctcctcata cgggggagcc ggcggctaca cgcagtcccc
     3901 ggggggcttt ggatcgcccg caccttctca agccgaaaag aaatcaagag cccgagccca
     3961 gcacattgtg ccctgtacta tatctcagct gctttctgcc actttggttg atgaagtgtt
     4021 cagaattggg aatgttgaga tttcacaggt cactattgtg gggatcatca gacatgcaga
     4081 gaaggctcca accaacattg tttacaaaat agatgacatg acagctgcac ccatggacgt
     4141 tcgccagtgg gttgacacag atgacaccag cagtgaaaac actgtggttc ctccagaaac
     4201 atatgtgaaa gtggcaggcc acctgagatc ttttcagaac aaaaagagcc tggtagcctt
     4261 taagatcatg cccctggagg atatgaatga gttcaccaca catattctgg aagtgatcaa
     4321 tgcacacatg gtactaagca aagccaacag ccagccctca gcagggagag cacctatcag
     4381 caatccagga atgagtgaag cagggaactt tggtgggaat agcttcatgc cagcaaatgg
     4441 cctcactgtg gcccaaaacc aggtgttgaa tttgattaag gcttgtccaa gacctgaagg
     4501 gttgaacttt caggatctca agaaccagct gaaacacatg tctgtatcct caatcaagca
     4561 agctgtggat tttctgagca atgaggggca catctattct actgtggatg atgaccattt
     4621 taaatccaca gatgcagaat aaggatccaa ttccgcccct ctccctcccc cccccctaac
     4681 gttactggcc gaagccgctt ggaataaggc cggtgtgcgt ttgtctatat gttattttcc
     4741 accatattgc cgtcttttgg caatgtgagg gcccggaaac ctggccctgt cttcttgacg
     4801 agcattccta ggggtctttc ccctctcgcc aaaggaatgc aaggtctgtt gaatgtcgtg
     4861 aaggaagcag ttcctctgga agcttcttga agacaaacaa cgtctgtagc gaccctttgc
     4921 aggcagcgga accccccacc tggcgacagg tgcctctgcg gccaaaagcc acgtgtataa
     4981 gatacacctg caaaggcggc acaaccccag tgccacgttg tgagttggat agttgtggaa
     5041 agagtcaaat ggctctcctc aagcgtattc aacaaggggc tgaaggatgc ccagaaggta
     5101 ccccattgta tgggatctga tctggggcct cggtgcacat gctttacatg tgtttagtcg
     5161 aggttaaaaa aacgtctagg ccccccgaac cacggggacg tggttttcct ttgaaaaaca
     5221 cgataaagat ctcatggtgg acatgatgga cttgcccagg tcgcgcatca acgccggcat
     5281 gctagctcaa ttcatcgaca agcctgtctg cttcgtaggg aggctggaaa agattcatcc
     5341 caccggaaaa atgtttattc tttcagatgg agaaggaaaa aatggaacca tcgagttgat
     5401 ggaacccctt gatgaagaaa tctctggaat tgtggaagtg gttggaagag taaccgccaa
     5461 ggccaccatc ttgtgtacat cttatgtcca gtttaaagaa gatagccatc cttttgatct
     5521 tggactttac aatgaagctg tgaaaattat ccatgacttc cctcagtttt atcctttagg
     5581 gattgtgcaa catgattgat ctagataact gatcataatc agccatacca catttgtaga
     5641 ggttttactt gctttaaaaa acctcccaca cctccccctg aacctgaaac ataaaatgaa
     5701 tgcaattgtt gttgttaact tgtttattgc agcttataat ggttacaaat aaagcaatag
     5761 catcacaaat ttcacaaata aagcattttt ttcactgcat tctagttgtg gtttgtccaa
     5821 actcatcaat gtatcttaac gcgtaaattg taagcgttaa tattttgtta aaattcgcgt
     5881 taaatttttg ttaaatcagc tcatttttta accaataggc cgaaatcggc aaaatccctt
     5941 ataaatcaaa agaatagacc gagatagggt tgagtgttgt tccagtttgg aacaagagtc
     6001 cactattaaa gaacgtggac tccaacgtca aagggcgaaa aaccgtctat cagggcgatg
     6061 gcccactacg tgaaccatca ccctaatcaa gttttttggg gtcgaggtgc cgtaaagcac
     6121 taaatcggaa ccctaaaggg agcccccgat ttagagcttg acggggaaag ccggcgaacg
     6181 tggcgagaaa ggaagggaag aaagcgaaag gagcgggcgc tagggcgctg gcaagtgtag
     6241 cggtcacgct gcgcgtaacc accacacccg ccgcgcttaa tgcgccgcta cagggcgcgt
     6301 caggtggcac ttttcgggga aatgtgcgcg gaacccctat ttgtttattt ttctaaatac
     6361 attcaaatat gtatccgctc atgagacaat aaccctgata aatgcttcaa taatattgaa
     6421 aaaggaagag tcctgaggcg gaaagaacca gctgtggaat gtgtgtcagt tagggtgtgg
     6481 aaagtcccca ggctccccag caggcagaag tatgcaaagc atgcatctca attagtcagc
     6541 aaccaggtgt ggaaagtccc caggctcccc agcaggcaga agtatgcaaa gcatgcatct
     6601 caattagtca gcaaccatag tcccgcccct aactccgccc atcccgcccc taactccgcc
     6661 cagttccgcc cattctccgc cccatggctg actaattttt tttatttatg cagaggccga
     6721 ggccgcctcg gcctctgagc tattccagaa gtagtgagga ggcttttttg gaggcctagg
     6781 cttttgcaaa gatcgatcaa gagacaggat gaggatcgtt tcgcatgatt gaacaagatg
     6841 gattgcacgc aggttctccg gccgcttggg tggagaggct attcggctat gactgggcac
     6901 aacagacaat cggctgctct gatgccgccg tgttccggct gtcagcgcag gggcgcccgg
     6961 ttctttttgt caagaccgac ctgtccggtg ccctgaatga actgcaagac gaggcagcgc
     7021 ggctatcgtg gctggccacg acgggcgttc cttgcgcagc tgtgctcgac gttgtcactg
     7081 aagcgggaag ggactggctg ctattgggcg aagtgccggg gcaggatctc ctgtcatctc
     7141 accttgctcc tgccgagaaa gtatccatca tggctgatgc aatgcggcgg ctgcatacgc
     7201 ttgatccggc tacctgccca ttcgaccacc aagcgaaaca tcgcatcgag cgagcacgta
     7261 ctcggatgga agccggtctt gtcgatcagg atgatctgga cgaagagcat caggggctcg
     7321 cgccagccga actgttcgcc aggctcaagg cgagcatgcc cgacggcgag gatctcgtcg
     7381 tgacccatgg cgatgcctgc ttgccgaata tcatggtgga aaatggccgc ttttctggat
     7441 tcatcgactg tggccggctg ggtgtggcgg accgctatca ggacatagcg ttggctaccc
     7501 gtgatattgc tgaagagctt ggcggcgaat gggctgaccg cttcctcgtg ctttacggta
     7561 tcgccgctcc cgattcgcag cgcatcgcct tctatcgcct tcttgacgag ttcttctgag
     7621 cgggactctg gggttcgaaa tgaccgacca agcgacgccc aacctgccat cacgagattt
     7681 cgattccacc gccgccttct atgaaaggtt gggcttcgga atcgttttcc gggacgccgg
     7741 ctggatgatc ctccagcgcg gggatctcat gctggagttc ttcgcccacc ctagggggag
     7801 gctaactgaa acacggaagg agacaatacc ggaaggaacc cgcgctatga cggcaataaa
     7861 aagacagaat aaaacgcacg gtgttgggtc gtttgttcat aaacgcgggg ttcggtccca
     7921 gggctggcac tctgtcgata ccccaccgag accccattgg ggccaatacg cccgcgtttc
     7981 ttccttttcc ccaccccacc ccccaagttc gggtgaaggc ccagggctcg cagccaacgt
     8041 cggggcggca ggccctgcca tagcctcagg ttactcatat atactttaga ttgatttaaa
     8101 acttcatttt taatttaaaa ggatctaggt gaagatcctt tttgataatc tcatgaccaa
     8161 aatcccttaa cgtgagtttt cgttccactg agcgtcagac cccgtagaaa agatcaaagg
     8221 atcttcttga gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc
     8281 gctaccagcg gtggtttgtt tgccggatca agagctacca actctttttc cgaaggtaac
     8341 tggcttcagc agagcgcaga taccaaatac tgttcttcta gtgtagccgt agttaggcca
     8401 ccacttcaag aactctgtag caccgcctac atacctcgct ctgctaatcc tgttaccagt
     8461 ggctgctgcc agtggcgata agtcgtgtct taccgggttg gactcaagac gatagttacc
     8521 ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc acacagccca gcttggagcg
     8581 aacgacctac accgaactga gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc
     8641 cgaagggaga aaggcggaca ggtatccggt aagcggcagg gtcggaacag gagagcgcac
     8701 gagggagctt ccagggggaa acgcctggta tctttatagt cctgtcgggt ttcgccacct
     8761 ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc
     8821 cagcaacgcg gcctttttac ggttcctggc cttttgctgg ccttttgctc acatgttctt
     8881 tcctgcgtta tcccctgatt ctgtggataa ccgtattacc g
//
