LOCUS       sequence-420497-        5526 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  mammalian expression of human RPA32 N-terminally fused to EGFP.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5526)
  TITLE     Pospiech Lab Plasmids 2023
REFERENCE   2  (bases 1 to 5526)
  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: pEGFP-RPA32
FEATURES             Location/Qualifiers
     source          1..5526
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        76..379
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        380..583
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     534..554
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      622..631
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(628..630,631..633,634..1344)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(628..630,631..633,634..1344)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(673..694)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(934..953)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1281..1302
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    2329..2450
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2366..2385)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2420..2439
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2457..2912)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2594..2615)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2806..2825
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2939..3043
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3045..3402
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3045..3236
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3068..3088)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3253..3388
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3437..4231
                     /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             3437..4231
                     /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(3491..3510)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4101..4120
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4419..4438)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4463..4510
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4839..5427
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5328..5347
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 attaccgcca tgcattagtt attaatagta atcaattacg gggtcattag ttcatagccc
       61 atatatggag ttccgcgtta cataacttac ggtaaatggc ccgcctggct gaccgcccaa
      121 cgacccccgc ccattgacgt caataatgac gtatgttccc atagtaacgc caatagggac
      181 tttccattga cgtcaatggg tggagtattt acggtaaact gcccacttgg cagtacatca
      241 agtgtatcat atgccaagta cgccccctat tgacgtcaat gacggtaaat ggcccgcctg
      301 gcattatgcc cagtacatga ccttatggga ctttcctact tggcagtaca tctacgtatt
      361 agtcatcgct attaccatgg tgatgcggtt ttggcagtac atcaatgggc gtggatagcg
      421 gtttgactca cggggatttc caagtctcca ccccattgac gtcaatggga gtttgttttg
      481 gcaccaaaat caacgggact ttccaaaatg tcgtaacaac tccgccccat tgacgcaaat
      541 gggcggtagg cgtgtacggt gggaggtcta tataagcaga gctggtttag tgaaccgtca
      601 gatccgctag cgctaccggt cgccaccatg gtgagcaagg gcgaggagct gttcaccggg
      661 gtggtgccca tcctggtcga gctggacggc gacgtaaacg gccacaagtt cagcgtgtcc
      721 ggcgagggcg agggcgatgc cacctacggc aagctgaccc tgaagttcat ctgcaccacc
      781 ggcaagctgc ccgtgccctg gcccaccctc gtgaccaccc tgacctacgg cgtgcagtgc
      841 ttcagccgct accccgacca catgaagcag cacgacttct tcaagtccgc catgcccgaa
      901 ggctacgtcc aggagcgcac catcttcttc aaggacgacg gcaactacaa gacccgcgcc
      961 gaggtgaagt tcgagggcga caccctggtg aaccgcatcg agctgaaggg catcgacttc
     1021 aaggaggacg gcaacatcct ggggcacaag ctggagtaca actacaacag ccacaacgtc
     1081 tatatcatgg ccgacaagca gaagaacggc atcaaggtga acttcaagat ccgccacaac
     1141 atcgaggacg gcagcgtgca gctcgccgac cactaccagc agaacacccc catcggcgac
     1201 ggccccgtgc tgctgcccga caaccactac ctgagcaccc agtccgccct gagcaaagac
     1261 cccaacgaga agcgcgatca catggtcctg ctggagttcg tgaccgccgc cgggatcact
     1321 ctcggcatgg acgagctgta caagtccgga ctcagatctc gagctcaagc ttcgaattct
     1381 gccaccatgt ggaacagtgg attcgaaagc tatggcagct cctcatacgg gggagccggc
     1441 ggctacacgc agtccccggg gggctttgga tcgcccgcac cttctcaagc cgaaaagaaa
     1501 tcaagagccc gagcccagca cattgtgccc tgtactatat ctcagctgct ttctgccact
     1561 ttggttgatg aagtgttcag aattgggaat gttgagattt cacaggtcac tattgtgggg
     1621 atcatcagac atgcagagaa ggctccaacc aacattgttt acaaaataga tgacatgaca
     1681 gctgcaccca tggacgttcg ccagtgggtt gacacagatg acaccagcag tgaaaacact
     1741 gtggttcctc cagaaacata tgtgaaagtg gcaggccacc tgagatcttt tcagaacaaa
     1801 aagagcctgg tagcctttaa gatcatgccc ctggaggata tgaatgagtt caccacacat
     1861 attctggaag tgatcaatgc acacatggta ctaagcaaag ccaacagcca gccctcagca
     1921 gggagagcac ctatcagcaa tccaggaatg agtgaagcag ggaactttgg tgggaatagc
     1981 ttcatgccag caaatggcct cactgtggcc caaaaccagg tgttgaattt gattaaggct
     2041 tgtccaagac ctgaagggtt gaactttcag gatctcaaga accagctgaa acacatgtct
     2101 gtatcctcaa tcaagcaagc tgtggatttt ctgagcaatg aggggcacat ctattctact
     2161 gtggatgatg accattttaa atccacagat gcagaataag gatccaccgg atctagataa
     2221 ctgatcataa tcagccatac cacatttgta gaggttttac ttgctttaaa aaacctccca
     2281 cacctccccc tgaacctgaa acataaaatg aatgcaattg ttgttgttaa cttgtttatt
     2341 gcagcttata atggttacaa ataaagcaat agcatcacaa atttcacaaa taaagcattt
     2401 ttttcactgc attctagttg tggtttgtcc aaactcatca atgtatctta acgcgtaaat
     2461 tgtaagcgtt aatattttgt taaaattcgc gttaaatttt tgttaaatca gctcattttt
     2521 taaccaatag gccgaaatcg gcaaaatccc ttataaatca aaagaataga ccgagatagg
     2581 gttgagtgtt gttccagttt ggaacaagag tccactatta aagaacgtgg actccaacgt
     2641 caaagggcga aaaaccgtct atcagggcga tggcccacta cgtgaaccat caccctaatc
     2701 aagttttttg gggtcgaggt gccgtaaagc actaaatcgg aaccctaaag ggagcccccg
     2761 atttagagct tgacggggaa agccggcgaa cgtggcgaga aaggaaggga agaaagcgaa
     2821 aggagcgggc gctagggcgc tggcaagtgt agcggtcacg ctgcgcgtaa ccaccacacc
     2881 cgccgcgctt aatgcgccgc tacagggcgc gtcaggtggc acttttcggg gaaatgtgcg
     2941 cggaacccct atttgtttat ttttctaaat acattcaaat atgtatccgc tcatgagaca
     3001 ataaccctga taaatgcttc aataatattg aaaaaggaag agtcctgagg cggaaagaac
     3061 cagctgtgga atgtgtgtca gttagggtgt ggaaagtccc caggctcccc agcaggcaga
     3121 agtatgcaaa gcatgcatct caattagtca gcaaccaggt gtggaaagtc cccaggctcc
     3181 ccagcaggca gaagtatgca aagcatgcat ctcaattagt cagcaaccat agtcccgccc
     3241 ctaactccgc ccatcccgcc cctaactccg cccagttccg cccattctcc gccccatggc
     3301 tgactaattt tttttattta tgcagaggcc gaggccgcct cggcctctga gctattccag
     3361 aagtagtgag gaggcttttt tggaggccta ggcttttgca aagatcgatc aagagacagg
     3421 atgaggatcg tttcgcatga ttgaacaaga tggattgcac gcaggttctc cggccgcttg
     3481 ggtggagagg ctattcggct atgactgggc acaacagaca atcggctgct ctgatgccgc
     3541 cgtgttccgg ctgtcagcgc aggggcgccc ggttcttttt gtcaagaccg acctgtccgg
     3601 tgccctgaat gaactgcaag acgaggcagc gcggctatcg tggctggcca cgacgggcgt
     3661 tccttgcgca gctgtgctcg acgttgtcac tgaagcggga agggactggc tgctattggg
     3721 cgaagtgccg gggcaggatc tcctgtcatc tcaccttgct cctgccgaga aagtatccat
     3781 catggctgat gcaatgcggc ggctgcatac gcttgatccg gctacctgcc cattcgacca
     3841 ccaagcgaaa catcgcatcg agcgagcacg tactcggatg gaagccggtc ttgtcgatca
     3901 ggatgatctg gacgaagagc atcaggggct cgcgccagcc gaactgttcg ccaggctcaa
     3961 ggcgagcatg cccgacggcg aggatctcgt cgtgacccat ggcgatgcct gcttgccgaa
     4021 tatcatggtg gaaaatggcc gcttttctgg attcatcgac tgtggccggc tgggtgtggc
     4081 ggaccgctat caggacatag cgttggctac ccgtgatatt gctgaagagc ttggcggcga
     4141 atgggctgac cgcttcctcg tgctttacgg tatcgccgct cccgattcgc agcgcatcgc
     4201 cttctatcgc cttcttgacg agttcttctg agcgggactc tggggttcga aatgaccgac
     4261 caagcgacgc ccaacctgcc atcacgagat ttcgattcca ccgccgcctt ctatgaaagg
     4321 ttgggcttcg gaatcgtttt ccgggacgcc ggctggatga tcctccagcg cggggatctc
     4381 atgctggagt tcttcgccca ccctaggggg aggctaactg aaacacggaa ggagacaata
     4441 ccggaaggaa cccgcgctat gacggcaata aaaagacaga ataaaacgca cggtgttggg
     4501 tcgtttgttc ataaacgcgg ggttcggtcc cagggctggc actctgtcga taccccaccg
     4561 agaccccatt ggggccaata cgcccgcgtt tcttcctttt ccccacccca ccccccaagt
     4621 tcgggtgaag gcccagggct cgcagccaac gtcggggcgg caggccctgc catagcctca
     4681 ggttactcat atatacttta gattgattta aaacttcatt tttaatttaa aaggatctag
     4741 gtgaagatcc tttttgataa tctcatgacc aaaatccctt aacgtgagtt ttcgttccac
     4801 tgagcgtcag accccgtaga aaagatcaaa ggatcttctt gagatccttt ttttctgcgc
     4861 gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg tttgccggat
     4921 caagagctac caactctttt tccgaaggta actggcttca gcagagcgca gataccaaat
     4981 actgttcttc tagtgtagcc gtagttaggc caccacttca agaactctgt agcaccgcct
     5041 acatacctcg ctctgctaat cctgttacca gtggctgctg ccagtggcga taagtcgtgt
     5101 cttaccgggt tggactcaag acgatagtta ccggataagg cgcagcggtc gggctgaacg
     5161 gggggttcgt gcacacagcc cagcttggag cgaacgacct acaccgaact gagataccta
     5221 cagcgtgagc tatgagaaag cgccacgctt cccgaaggga gaaaggcgga caggtatccg
     5281 gtaagcggca gggtcggaac aggagagcgc acgagggagc ttccaggggg aaacgcctgg
     5341 tatctttata gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc
     5401 tcgtcagggg ggcggagcct atggaaaaac gccagcaacg cggccttttt acggttcctg
     5461 gccttttgct ggccttttgc tcacatgttc tttcctgcgt tatcccctga ttctgtggat
     5521 aaccgt
//
