LOCUS       sequence-416902-        5526 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  expression of GFP-tagged hsRPA2-Asp8 in human cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5526)
  TITLE     Wold 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-hsRPA32-Asp8 (MSW#507)
FEATURES             Location/Qualifiers
     source          1..5526
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        68..371
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        372..575
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     526..546
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      614..623
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(620..622,623..625,626..1336)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(620..622,623..625,626..1336)
                     /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(665..686)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(926..945)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1273..1294
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    2321..2442
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2358..2377)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2412..2431
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2449..2904)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2586..2607)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2798..2817
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2931..3035
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3037..3394
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3037..3228
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3060..3080)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3245..3380
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3429..4223
                     /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             3429..4223
                     /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(3483..3502)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4093..4112
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4411..4430)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4455..4502
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4831..5419
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5320..5339
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 catgcattag ttattaatag taatcaatta cggggtcatt agttcatagc ccatatatgg
       61 agttccgcgt tacataactt acggtaaatg gcccgcctgg ctgaccgccc aacgaccccc
      121 gcccattgac gtcaataatg acgtatgttc ccatagtaac gccaataggg actttccatt
      181 gacgtcaatg ggtggagtat ttacggtaaa ctgcccactt ggcagtacat caagtgtatc
      241 atatgccaag tacgccccct attgacgtca atgacggtaa atggcccgcc tggcattatg
      301 cccagtacat gaccttatgg gactttccta cttggcagta catctacgta ttagtcatcg
      361 ctattaccat ggtgatgcgg ttttggcagt acatcaatgg gcgtggatag cggtttgact
      421 cacggggatt tccaagtctc caccccattg acgtcaatgg gagtttgttt tggcaccaaa
      481 atcaacggga ctttccaaaa tgtcgtaaca actccgcccc attgacgcaa atgggcggta
      541 ggcgtgtacg gtgggaggtc tatataagca gagctggttt agtgaaccgt cagatccgct
      601 agcgctaccg gtcgccacca tggtgagcaa gggcgaggag ctgttcaccg gggtggtgcc
      661 catcctggtc gagctggacg gcgacgtaaa cggccacaag ttcagcgtgt ccggcgaggg
      721 cgagggcgat gccacctacg gcaagctgac cctgaagttc atctgcacca ccggcaagct
      781 gcccgtgccc tggcccaccc tcgtgaccac cctgacctac ggcgtgcagt gcttcagccg
      841 ctaccccgac cacatgaagc agcacgactt cttcaagtcc gccatgcccg aaggctacgt
      901 ccaggagcgc accatcttct tcaaggacga cggcaactac aagacccgcg ccgaggtgaa
      961 gttcgagggc gacaccctgg tgaaccgcat cgagctgaag ggcatcgact tcaaggagga
     1021 cggcaacatc ctggggcaca agctggagta caactacaac agccacaacg tctatatcat
     1081 ggccgacaag cagaagaacg gcatcaaggt gaacttcaag atccgccaca acatcgagga
     1141 cggcagcgtg cagctcgccg accactacca gcagaacacc cccatcggcg acggccccgt
     1201 gctgctgccc gacaaccact acctgagcac ccagtccgcc ctgagcaaag accccaacga
     1261 gaagcgcgat cacatggtcc tgctggagtt cgtgaccgcc gccgggatca ctctcggcat
     1321 ggacgagctg tacaagtccg gactcagatc tcgaggtgga ggcatgtgga acagtggatt
     1381 cgaagactat ggcgacgacg actacggggg agccggcggc tacgaccagg atccgggggg
     1441 ctttggagac ccagcacctg atcaagccga aaagaaatca agagcccgag cccagcacat
     1501 tgtgccctgt actatatctc agctgctttc tgccactttg gttgatgaag tgttcagaat
     1561 tgggaatgtt gagatttcac aggtcactat tgtggggatc atcagacatg cagagaaggc
     1621 tccaatcaac attgtttaca aaatagatga catgacagct gcacccatgg acgttcgcca
     1681 gtgggttgac acagatgaca ccagcagtga aaacactgtg gttcctccag aaacatatgt
     1741 gaaagtggca ggccacctga gatcttttca gaacaaaaag agcctggtag cctttaagat
     1801 catgcccctg gaggatatga atgagttcac cacacatatt ctggaagtga tcaatgcaca
     1861 catggtacta agcaaagcca acagccagcc ctcagcaggg agagcaccta tcagcaatcc
     1921 aggaatgagt gaagcaggga actttggtgg gaatagcttc atgccagcaa atggcctcac
     1981 tgtggcccaa aaccaggtgt tgaatttgat taaggcttgt ccaagacctg aagggttgaa
     2041 ctttcaggat ctcaagaacc agctgaaaca catgtctgta tcctcaatca agcaagctgt
     2101 ggattttctg agcaatgagg ggcacatcta ttctactgtg gatgatgacc attttaaatc
     2161 cacagatgca gaataaggta ccgcgggccc gggatccacc ggatctagat aactgatcat
     2221 aatcagccat accacatttg tagaggtttt acttgcttta aaaaacctcc cacacctccc
     2281 cctgaacctg aaacataaaa tgaatgcaat tgttgttgtt aacttgttta ttgcagctta
     2341 taatggttac aaataaagca atagcatcac aaatttcaca aataaagcat ttttttcact
     2401 gcattctagt tgtggtttgt ccaaactcat caatgtatct taacgcgtaa attgtaagcg
     2461 ttaatatttt gttaaaattc gcgttaaatt tttgttaaat cagctcattt tttaaccaat
     2521 aggccgaaat cggcaaaatc ccttataaat caaaagaata gaccgagata gggttgagtg
     2581 ttgttccagt ttggaacaag agtccactat taaagaacgt ggactccaac gtcaaagggc
     2641 gaaaaaccgt ctatcagggc gatggcccac tacgtgaacc atcaccctaa tcaagttttt
     2701 tggggtcgag gtgccgtaaa gcactaaatc ggaaccctaa agggagcccc cgatttagag
     2761 cttgacgggg aaagccggcg aacgtggcga gaaaggaagg gaagaaagcg aaaggagcgg
     2821 gcgctagggc gctggcaagt gtagcggtca cgctgcgcgt aaccaccaca cccgccgcgc
     2881 ttaatgcgcc gctacagggc gcgtcaggtg gcacttttcg gggaaatgtg cgcggaaccc
     2941 ctatttgttt atttttctaa atacattcaa atatgtatcc gctcatgaga caataaccct
     3001 gataaatgct tcaataatat tgaaaaagga agagtcctga ggcggaaaga accagctgtg
     3061 gaatgtgtgt cagttagggt gtggaaagtc cccaggctcc ccagcaggca gaagtatgca
     3121 aagcatgcat ctcaattagt cagcaaccag gtgtggaaag tccccaggct ccccagcagg
     3181 cagaagtatg caaagcatgc atctcaatta gtcagcaacc atagtcccgc ccctaactcc
     3241 gcccatcccg cccctaactc cgcccagttc cgcccattct ccgccccatg gctgactaat
     3301 tttttttatt tatgcagagg ccgaggccgc ctcggcctct gagctattcc agaagtagtg
     3361 aggaggcttt tttggaggcc taggcttttg caaagatcga tcaagagaca ggatgaggat
     3421 cgtttcgcat gattgaacaa gatggattgc acgcaggttc tccggccgct tgggtggaga
     3481 ggctattcgg ctatgactgg gcacaacaga caatcggctg ctctgatgcc gccgtgttcc
     3541 ggctgtcagc gcaggggcgc ccggttcttt ttgtcaagac cgacctgtcc ggtgccctga
     3601 atgaactgca agacgaggca gcgcggctat cgtggctggc cacgacgggc gttccttgcg
     3661 cagctgtgct cgacgttgtc actgaagcgg gaagggactg gctgctattg ggcgaagtgc
     3721 cggggcagga tctcctgtca tctcaccttg ctcctgccga gaaagtatcc atcatggctg
     3781 atgcaatgcg gcggctgcat acgcttgatc cggctacctg cccattcgac caccaagcga
     3841 aacatcgcat cgagcgagca cgtactcgga tggaagccgg tcttgtcgat caggatgatc
     3901 tggacgaaga gcatcagggg ctcgcgccag ccgaactgtt cgccaggctc aaggcgagca
     3961 tgcccgacgg cgaggatctc gtcgtgaccc atggcgatgc ctgcttgccg aatatcatgg
     4021 tggaaaatgg ccgcttttct ggattcatcg actgtggccg gctgggtgtg gcggaccgct
     4081 atcaggacat agcgttggct acccgtgata ttgctgaaga gcttggcggc gaatgggctg
     4141 accgcttcct cgtgctttac ggtatcgccg ctcccgattc gcagcgcatc gccttctatc
     4201 gccttcttga cgagttcttc tgagcgggac tctggggttc gaaatgaccg accaagcgac
     4261 gcccaacctg ccatcacgag atttcgattc caccgccgcc ttctatgaaa ggttgggctt
     4321 cggaatcgtt ttccgggacg ccggctggat gatcctccag cgcggggatc tcatgctgga
     4381 gttcttcgcc caccctaggg ggaggctaac tgaaacacgg aaggagacaa taccggaagg
     4441 aacccgcgct atgacggcaa taaaaagaca gaataaaacg cacggtgttg ggtcgtttgt
     4501 tcataaacgc ggggttcggt cccagggctg gcactctgtc gataccccac cgagacccca
     4561 ttggggccaa tacgcccgcg tttcttcctt ttccccaccc caccccccaa gttcgggtga
     4621 aggcccaggg ctcgcagcca acgtcggggc ggcaggccct gccatagcct caggttactc
     4681 atatatactt tagattgatt taaaacttca tttttaattt aaaaggatct aggtgaagat
     4741 cctttttgat aatctcatga ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc
     4801 agaccccgta gaaaagatca aaggatcttc ttgagatcct ttttttctgc gcgtaatctg
     4861 ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt tgtttgccgg atcaagagct
     4921 accaactctt tttccgaagg taactggctt cagcagagcg cagataccaa atactgttct
     4981 tctagtgtag ccgtagttag gccaccactt caagaactct gtagcaccgc ctacatacct
     5041 cgctctgcta atcctgttac cagtggctgc tgccagtggc gataagtcgt gtcttaccgg
     5101 gttggactca agacgatagt taccggataa ggcgcagcgg tcgggctgaa cggggggttc
     5161 gtgcacacag cccagcttgg agcgaacgac ctacaccgaa ctgagatacc tacagcgtga
     5221 gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg
     5281 cagggtcgga acaggagagc gcacgaggga gcttccaggg ggaaacgcct ggtatcttta
     5341 tagtcctgtc gggtttcgcc acctctgact tgagcgtcga tttttgtgat gctcgtcagg
     5401 ggggcggagc ctatggaaaa acgccagcaa cgcggccttt ttacggttcc tggccttttg
     5461 ctggcctttt gctcacatgt tctttcctgc gttatcccct gattctgtgg ataaccgtat
     5521 taccgc
//
