LOCUS       sequence-420425-        6968 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  mammalian expression of human RPA32 C-terminally fused to mVenus.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6968)
  TITLE     Pospiech Lab Plasmids 2023
REFERENCE   2  (bases 1 to 6968)
  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: pcDNA3-RPA32-mVen
FEATURES             Location/Qualifiers
     source          1..6968
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(72..91)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        263..642
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        643..846
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     797..817
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     891..910
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        891..909
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      1814..1823
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1820..1822,1823..1825,1826..2539)
                     /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(1865..1886)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2126..2145)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2473..2494
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     promoter        complement(2549..2567)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(2550..2567)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(2587..2604)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2593..2817
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2863..3291
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2950..2969)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3160..3181
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3300..3320)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3305..3634
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3485..3620
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3547..3566
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3701..4495
                     /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(3755..3774)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4365..4384
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4669..4790
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4706..4725)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4760..4779
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4839..4855)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4839..4855)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4852..4874)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4863..4879
                     /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(4887..4893,4894..4911,4912..4917))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4932..4953
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5070..5087)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5241..5829)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5321..5340)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6000..6791,6792..6860))
                     /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     6623..6642
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6861..6965)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 atttccccga aaagtgccac ctgacgtcga cggatcggga gatctcccga tcccctatgg
       61 tgcactctca gtacaatctg ctctgatgcc gcatagttaa gccagtatct gctccctgct
      121 tgtgtgttgg aggtcgctga gtagtgcgcg agcaaaattt aagctacaac aaggcaaggc
      181 ttgaccgaca attgcatgaa gaatctgctt agggttaggc gttttgcgct gcttcgcgat
      241 gtacgggcca gatatacgcg ttgacattga ttattgacta gttattaata gtaatcaatt
      301 acggggtcat tagttcatag cccatatatg gagttccgcg ttacataact tacggtaaat
      361 ggcccgcctg gctgaccgcc caacgacccc cgcccattga cgtcaataat gacgtatgtt
      421 cccatagtaa cgccaatagg gactttccat tgacgtcaat gggtggagta tttacggtaa
      481 actgcccact tggcagtaca tcaagtgtat catatgccaa gtacgccccc tattgacgtc
      541 aatgacggta aatggcccgc ctggcattat gcccagtaca tgaccttatg ggactttcct
      601 acttggcagt acatctacgt attagtcatc gctattacca tggtgatgcg gttttggcag
      661 tacatcaatg ggcgtggata gcggtttgac tcacggggat ttccaagtct ccaccccatt
      721 gacgtcaatg ggagtttgtt ttggcaccaa aatcaacggg actttccaaa atgtcgtaac
      781 aactccgccc cattgacgca aatgggcggt aggcgtgtac ggtgggaggt ctatataagc
      841 agagctctct ggctaactag agaacccact gcttactggc ttatcgaaat taatacgact
      901 cactataggg agacccaagc ttggtaccga gctcggatcc gccaccatgt ggaacagtgg
      961 attcgaaagc tatggcagct cctcatacgg gggagccggc ggctacacgc agtccccggg
     1021 gggctttgga tcgcccgcac cttctcaagc cgaaaagaaa tcaagagccc gagcccagca
     1081 cattgtgccc tgtactatat ctcagctgct ttctgccact ttggttgatg aagtgttcag
     1141 aattgggaat gttgagattt cacaggtcac tattgtgggg atcatcagac atgcagagaa
     1201 ggctccaacc aacattgttt acaaaataga tgacatgaca gctgcaccca tggacgttcg
     1261 ccagtgggtt gacacagatg acaccagcag tgaaaacact gtggttcctc cagaaacata
     1321 tgtgaaagtg gcaggccacc tgagatcttt tcagaacaaa aagagcctgg tagcctttaa
     1381 gatcatgccc ctggaggata tgaatgagtt caccacacat attctggaag tgatcaatgc
     1441 acacatggta ctaagcaaag ccaacagcca gccctcagca gggagagcac ctatcagcaa
     1501 tccaggaatg agtgaagcag ggaactttgg tgggaatagc ttcatgccag caaatggcct
     1561 cactgtggcc caaaaccagg tgttgaattt gattaaggct tgtccaagac ctgaagggtt
     1621 gaactttcag gatctcaaga accagctgaa acacatgtct gtatcctcaa tcaagcaagc
     1681 tgtggatttt ctgagcaatg aggggcacat ctattctact gtggatgatg accattttaa
     1741 atccacagat gcagaagaat tctgcagata tccatcacac tggcggccgc tcgagcatgc
     1801 atctagaccg gtcgccacca tggtgagcaa gggcgaggag ctgttcaccg gggtggtgcc
     1861 catcctggtc gagctggacg gcgacgtaaa cggccacaag ttcagcgtgt ccggcgaggg
     1921 cgagggcgat gccacctacg gcaagctgac cctgaagctg atctgcacca ccggcaagct
     1981 gcccgtgccc tggcccaccc tcgtgaccac cctgggctac ggcctgcagt gcttcgcccg
     2041 ctaccccgac cacatgaagc agcacgactt cttcaagtcc gccatgcccg aaggctacgt
     2101 ccaggagcgc accatcttct tcaaggacga cggcaactac aagacccgcg ccgaggtgaa
     2161 gttcgagggc gacaccctgg tgaaccgcat cgagctgaag ggcatcgact tcaaggagga
     2221 cggcaacatc ctggggcaca agctggagta caactacaac agccacaacg tctatatcac
     2281 cgccgacaag cagaagaacg gcatcaaggc caacttcaag atccgccaca acatcgagga
     2341 cggcggcgtg cagctcgccg accactacca gcagaacacc cccatcggcg acggccccgt
     2401 gctgctgccc gacaaccact acctgagcta ccagtccaag ctgagcaaag accccaacga
     2461 gaagcgcgat cacatggtcc tgctggagtt cgtgaccgcc gccgggatca ctctcggcat
     2521 ggacgagctg tacaagtaag ggccctattc tatagtgtca cctaaatgct agagctcgct
     2581 gatcagcctc gactgtgcct tctagttgcc agccatctgt tgtttgcccc tcccccgtgc
     2641 cttccttgac cctggaaggt gccactccca ctgtcctttc ctaataaaat gaggaaattg
     2701 catcgcattg tctgagtagg tgtcattcta ttctgggggg tggggtgggg caggacagca
     2761 agggggagga ttgggaagac aatagcaggc atgctgggga tgcggtgggc tctatggctt
     2821 ctgaggcgga aagaaccagc tggggctcta gggggtatcc ccacgcgccc tgtagcggcg
     2881 cattaagcgc ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt gccagcgccc
     2941 tagcgcccgc tcctttcgct ttcttccctt cctttctcgc cacgttcgcc ggctttcccc
     3001 gtcaagctct aaatcggggg ctccctttag ggttccgatt tagtgcttta cggcacctcg
     3061 accccaaaaa acttgattag ggtgatggtt cacgtagtgg gccatcgccc tgatagacgg
     3121 tttttcgccc tttgacgttg gagtccacgt tctttaatag tggactcttg ttccaaactg
     3181 gaacaacact caaccctatc tcggtctatt cttttgattt ataagggatt ttgccgattt
     3241 cggcctattg gttaaaaaat gagctgattt aacaaaaatt taacgcgaat taattctgtg
     3301 gaatgtgtgt cagttagggt gtggaaagtc cccaggctcc ccagcaggca gaagtatgca
     3361 aagcatgcat ctcaattagt cagcaaccag gtgtggaaag tccccaggct ccccagcagg
     3421 cagaagtatg caaagcatgc atctcaatta gtcagcaacc atagtcccgc ccctaactcc
     3481 gcccatcccg cccctaactc cgcccagttc cgcccattct ccgccccatg gctgactaat
     3541 tttttttatt tatgcagagg ccgaggccgc ctctgcctct gagctattcc agaagtagtg
     3601 aggaggcttt tttggaggcc taggcttttg caaaaagctc ccgggagctt gtatatccat
     3661 tttcggatct gatcaagaga caggatgagg atcgtttcgc atgattgaac aagatggatt
     3721 gcacgcaggt tctccggccg cttgggtgga gaggctattc ggctatgact gggcacaaca
     3781 gacaatcggc tgctctgatg ccgccgtgtt ccggctgtca gcgcaggggc gcccggttct
     3841 ttttgtcaag accgacctgt ccggtgccct gaatgaactg caggacgagg cagcgcggct
     3901 atcgtggctg gccacgacgg gcgttccttg cgcagctgtg ctcgacgttg tcactgaagc
     3961 gggaagggac tggctgctat tgggcgaagt gccggggcag gatctcctgt catctcacct
     4021 tgctcctgcc gagaaagtat ccatcatggc tgatgcaatg cggcggctgc atacgcttga
     4081 tccggctacc tgcccattcg accaccaagc gaaacatcgc atcgagcgag cacgtactcg
     4141 gatggaagcc ggtcttgtcg atcaggatga tctggacgaa gagcatcagg ggctcgcgcc
     4201 agccgaactg ttcgccaggc tcaaggcgcg catgcccgac ggcgaggatc tcgtcgtgac
     4261 ccatggcgat gcctgcttgc cgaatatcat ggtggaaaat ggccgctttt ctggattcat
     4321 cgactgtggc cggctgggtg tggcggaccg ctatcaggac atagcgttgg ctacccgtga
     4381 tattgctgaa gagcttggcg gcgaatgggc tgaccgcttc ctcgtgcttt acggtatcgc
     4441 cgctcccgat tcgcagcgca tcgccttcta tcgccttctt gacgagttct tctgagcggg
     4501 actctggggt tcgaaatgac cgaccaagcg acgcccaacc tgccatcacg agatttcgat
     4561 tccaccgccg ccttctatga aaggttgggc ttcggaatcg ttttccggga cgccggctgg
     4621 atgatcctcc agcgcgggga tctcatgctg gagttcttcg cccaccccaa cttgtttatt
     4681 gcagcttata atggttacaa ataaagcaat agcatcacaa atttcacaaa taaagcattt
     4741 ttttcactgc attctagttg tggtttgtcc aaactcatca atgtatctta tcatgtctgt
     4801 ataccgtcga cctctagcta gagcttggcg taatcatggt catagctgtt tcctgtgtga
     4861 aattgttatc cgctcacaat tccacacaac atacgagccg gaagcataaa gtgtaaagcc
     4921 tggggtgcct aatgagtgag ctaactcaca ttaattgcgt tgcgctcact gcccgctttc
     4981 cagtcgggaa acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc ggggagaggc
     5041 ggtttgcgta ttgggcgctc ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt
     5101 cggctgcggc gagcggtatc agctcactca aaggcggtaa tacggttatc cacagaatca
     5161 ggggataacg caggaaagaa catgtgagca aaaggccagc aaaaggccag gaaccgtaaa
     5221 aaggccgcgt tgctggcgtt tttccatagg ctccgccccc ctgacgagca tcacaaaaat
     5281 cgacgctcaa gtcagaggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc
     5341 cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc
     5401 gcctttctcc cttcgggaag cgtggcgctt tctcatagct cacgctgtag gtatctcagt
     5461 tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac
     5521 cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgacttatcg
     5581 ccactggcag cagccactgg taacaggatt agcagagcga ggtatgtagg cggtgctaca
     5641 gagttcttga agtggtggcc taactacggc tacactagaa gaacagtatt tggtatctgc
     5701 gctctgctga agccagttac cttcggaaaa agagttggta gctcttgatc cggcaaacaa
     5761 accaccgctg gtagcggtgg tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa
     5821 ggatctcaag aagatccttt gatcttttct acggggtctg acgctcagtg gaacgaaaac
     5881 tcacgttaag ggattttggt catgagatta tcaaaaagga tcttcaccta gatcctttta
     5941 aattaaaaat gaagttttaa atcaatctaa agtatatatg agtaaacttg gtctgacagt
     6001 taccaatgct taatcagtga ggcacctatc tcagcgatct gtctatttcg ttcatccata
     6061 gttgcctgac tccccgtcgt gtagataact acgatacggg agggcttacc atctggcccc
     6121 agtgctgcaa tgataccgcg agacccacgc tcaccggctc cagatttatc agcaataaac
     6181 cagccagccg gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag
     6241 tctattaatt gttgccggga agctagagta agtagttcgc cagttaatag tttgcgcaac
     6301 gttgttgcca ttgctacagg catcgtggtg tcacgctcgt cgtttggtat ggcttcattc
     6361 agctccggtt cccaacgatc aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg
     6421 gttagctcct tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt gttatcactc
     6481 atggttatgg cagcactgca taattctctt actgtcatgc catccgtaag atgcttttct
     6541 gtgactggtg agtactcaac caagtcattc tgagaatagt gtatgcggcg accgagttgc
     6601 tcttgcccgg cgtcaatacg ggataatacc gcgccacata gcagaacttt aaaagtgctc
     6661 atcattggaa aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc
     6721 agttcgatgt aacccactcg tgcacccaac tgatcttcag catcttttac tttcaccagc
     6781 gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca
     6841 cggaaatgtt gaatactcat actcttcctt tttcaatatt attgaagcat ttatcagggt
     6901 tattgtctca tgagcggata catatttgaa tgtatttaga aaaataaaca aataggggtt
     6961 ccgcgcac
//
