LOCUS       sequence-420424-        6971 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  mammalian expression of human RPA32 C-terminally fused to mCerulean.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6971)
  TITLE     Pospiech Lab Plasmids 2023
REFERENCE   2  (bases 1 to 6971)
  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-mCer
FEATURES             Location/Qualifiers
     source          1..6971
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(60..79)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        251..630
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        631..834
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     785..805
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     879..898
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        879..897
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      1805..1814
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1811..1813,1814..1816,1817..2530)
                     /codon_start=1
                     /product="enhanced monomeric variant of CFP (Rizzo et al., 
                     2004)"
                     /label=mCerulean
                     /note=" mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTWGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNAISDNVYITADKQKNGIK
                     ANFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1856..1877)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2117..2136)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2464..2485
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     promoter        complement(2540..2558)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(2541..2558)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(2578..2595)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2584..2808
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2854..3282
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2941..2960)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3151..3172
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3291..3311)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3296..3625
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3476..3611
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3538..3557
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3692..4486
                     /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(3746..3765)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4356..4375
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4660..4781
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4697..4716)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4751..4770
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4830..4846)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4830..4846)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4843..4865)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4854..4870
                     /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(4878..4884,4885..4902,4903..4908))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4923..4944
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5061..5078)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5232..5820)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5312..5331)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5991..6782,6783..6851))
                     /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     6614..6633
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6852..6956)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 agtgccacct gacgtcgacg gatcgggaga tctcccgatc ccctatggtg cactctcagt
       61 acaatctgct ctgatgccgc atagttaagc cagtatctgc tccctgcttg tgtgttggag
      121 gtcgctgagt agtgcgcgag caaaatttaa gctacaacaa ggcaaggctt gaccgacaat
      181 tgcatgaaga atctgcttag ggttaggcgt tttgcgctgc ttcgcgatgt acgggccaga
      241 tatacgcgtt gacattgatt attgactagt tattaatagt aatcaattac ggggtcatta
      301 gttcatagcc catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc
      361 tgaccgccca acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg
      421 ccaataggga ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg
      481 gcagtacatc aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa
      541 tggcccgcct ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac
      601 atctacgtat tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg
      661 cgtggatagc ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg
      721 agtttgtttt ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca
      781 ttgacgcaaa tgggcggtag gcgtgtacgg tgggaggtct atataagcag agctctctgg
      841 ctaactagag aacccactgc ttactggctt atcgaaatta atacgactca ctatagggag
      901 acccaagctt ggtaccgagc tcggatccgc caccatgtgg aacagtggat tcgaaagcta
      961 tggcagctcc tcatacgggg gagccggcgg ctacacgcag tccccggggg gctttggatc
     1021 gcccgcacct tctcaagccg aaaagaaatc aagagcccga gcccagcaca ttgtgccctg
     1081 tactatatct cagctgcttt ctgccacttt ggttgatgaa gtgttcagaa ttgggaatgt
     1141 tgagatttca caggtcacta ttgtggggat catcagacat gcagagaagg ctccaaccaa
     1201 cattgtttac aaaatagatg acatgacagc tgcacccatg gacgttcgcc agtgggttga
     1261 cacagatgac accagcagtg aaaacactgt ggttcctcca gaaacatatg tgaaagtggc
     1321 aggccacctg agatcttttc agaacaaaaa gagcctggta gcctttaaga tcatgcccct
     1381 ggaggatatg aatgagttca ccacacatat tctggaagtg atcaatgcac acatggtact
     1441 aagcaaagcc aacagccagc cctcagcagg gagagcacct atcagcaatc caggaatgag
     1501 tgaagcaggg aactttggtg ggaatagctt catgccagca aatggcctca ctgtggccca
     1561 aaaccaggtg ttgaatttga ttaaggcttg tccaagacct gaagggttga actttcagga
     1621 tctcaagaac cagctgaaac acatgtctgt atcctcaatc aagcaagctg tggattttct
     1681 gagcaatgag gggcacatct attctactgt ggatgatgac cattttaaat ccacagatgc
     1741 agaagaattc tgcagatatc catcacactg gcggccgctc gagcatgcat ctagaagatc
     1801 catcgccacc atggtgagca agggcgagga gctgttcacc ggggtggtgc ccatcctggt
     1861 cgagctggac ggcgacgtaa acggccacaa gttcagcgtg tccggcgagg gcgagggcga
     1921 tgccacctac ggcaagctga ccctgaagtt catctgcacc accggcaagc tgcccgtgcc
     1981 ctggcccacc ctcgtgacca ccctgacctg gggcgtgcag tgcttcgccc gctaccccga
     2041 ccacatgaag cagcacgact tcttcaagtc cgccatgccc gaaggctacg tccaggagcg
     2101 caccatcttc ttcaaggacg acggcaacta caagacccgc gccgaggtga agttcgaggg
     2161 cgacaccctg gtgaaccgca tcgagctgaa gggcatcgac ttcaaggagg acggcaacat
     2221 cctggggcac aagctggagt acaacgccat cagcgacaac gtctatatca ccgccgacaa
     2281 gcagaagaac ggcatcaagg ccaacttcaa gatccgccac aacatcgagg acggcagcgt
     2341 gcagctcgcc gaccactacc agcagaacac ccccatcggc gacggccccg tgctgctgcc
     2401 cgacaaccac tacctgagca cccagtccaa gctgagcaaa gaccccaacg agaagcgcga
     2461 tcacatggtc ctgctggagt tcgtgaccgc cgccgggatc actctcggca tggacgagct
     2521 gtacaagtaa gggccctatt ctatagtgtc acctaaatgc tagagctcgc tgatcagcct
     2581 cgactgtgcc ttctagttgc cagccatctg ttgtttgccc ctcccccgtg ccttccttga
     2641 ccctggaagg tgccactccc actgtccttt cctaataaaa tgaggaaatt gcatcgcatt
     2701 gtctgagtag gtgtcattct attctggggg gtggggtggg gcaggacagc aagggggagg
     2761 attgggaaga caatagcagg catgctgggg atgcggtggg ctctatggct tctgaggcgg
     2821 aaagaaccag ctggggctct agggggtatc cccacgcgcc ctgtagcggc gcattaagcg
     2881 cggcgggtgt ggtggttacg cgcagcgtga ccgctacact tgccagcgcc ctagcgcccg
     2941 ctcctttcgc tttcttccct tcctttctcg ccacgttcgc cggctttccc cgtcaagctc
     3001 taaatcgggg gctcccttta gggttccgat ttagtgcttt acggcacctc gaccccaaaa
     3061 aacttgatta gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg gtttttcgcc
     3121 ctttgacgtt ggagtccacg ttctttaata gtggactctt gttccaaact ggaacaacac
     3181 tcaaccctat ctcggtctat tcttttgatt tataagggat tttgccgatt tcggcctatt
     3241 ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa ttaattctgt ggaatgtgtg
     3301 tcagttaggg tgtggaaagt ccccaggctc cccagcaggc agaagtatgc aaagcatgca
     3361 tctcaattag tcagcaacca ggtgtggaaa gtccccaggc tccccagcag gcagaagtat
     3421 gcaaagcatg catctcaatt agtcagcaac catagtcccg cccctaactc cgcccatccc
     3481 gcccctaact ccgcccagtt ccgcccattc tccgccccat ggctgactaa ttttttttat
     3541 ttatgcagag gccgaggccg cctctgcctc tgagctattc cagaagtagt gaggaggctt
     3601 ttttggaggc ctaggctttt gcaaaaagct cccgggagct tgtatatcca ttttcggatc
     3661 tgatcaagag acaggatgag gatcgtttcg catgattgaa caagatggat tgcacgcagg
     3721 ttctccggcc gcttgggtgg agaggctatt cggctatgac tgggcacaac agacaatcgg
     3781 ctgctctgat gccgccgtgt tccggctgtc agcgcagggg cgcccggttc tttttgtcaa
     3841 gaccgacctg tccggtgccc tgaatgaact gcaggacgag gcagcgcggc tatcgtggct
     3901 ggccacgacg ggcgttcctt gcgcagctgt gctcgacgtt gtcactgaag cgggaaggga
     3961 ctggctgcta ttgggcgaag tgccggggca ggatctcctg tcatctcacc ttgctcctgc
     4021 cgagaaagta tccatcatgg ctgatgcaat gcggcggctg catacgcttg atccggctac
     4081 ctgcccattc gaccaccaag cgaaacatcg catcgagcga gcacgtactc ggatggaagc
     4141 cggtcttgtc gatcaggatg atctggacga agagcatcag gggctcgcgc cagccgaact
     4201 gttcgccagg ctcaaggcgc gcatgcccga cggcgaggat ctcgtcgtga cccatggcga
     4261 tgcctgcttg ccgaatatca tggtggaaaa tggccgcttt tctggattca tcgactgtgg
     4321 ccggctgggt gtggcggacc gctatcagga catagcgttg gctacccgtg atattgctga
     4381 agagcttggc ggcgaatggg ctgaccgctt cctcgtgctt tacggtatcg ccgctcccga
     4441 ttcgcagcgc atcgccttct atcgccttct tgacgagttc ttctgagcgg gactctgggg
     4501 ttcgaaatga ccgaccaagc gacgcccaac ctgccatcac gagatttcga ttccaccgcc
     4561 gccttctatg aaaggttggg cttcggaatc gttttccggg acgccggctg gatgatcctc
     4621 cagcgcgggg atctcatgct ggagttcttc gcccacccca acttgtttat tgcagcttat
     4681 aatggttaca aataaagcaa tagcatcaca aatttcacaa ataaagcatt tttttcactg
     4741 cattctagtt gtggtttgtc caaactcatc aatgtatctt atcatgtctg tataccgtcg
     4801 acctctagct agagcttggc gtaatcatgg tcatagctgt ttcctgtgtg aaattgttat
     4861 ccgctcacaa ttccacacaa catacgagcc ggaagcataa agtgtaaagc ctggggtgcc
     4921 taatgagtga gctaactcac attaattgcg ttgcgctcac tgcccgcttt ccagtcggga
     4981 aacctgtcgt gccagctgca ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt
     5041 attgggcgct cttccgcttc ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg
     5101 cgagcggtat cagctcactc aaaggcggta atacggttat ccacagaatc aggggataac
     5161 gcaggaaaga acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg
     5221 ttgctggcgt ttttccatag gctccgcccc cctgacgagc atcacaaaaa tcgacgctca
     5281 agtcagaggt ggcgaaaccc gacaggacta taaagatacc aggcgtttcc ccctggaagc
     5341 tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg gatacctgtc cgcctttctc
     5401 ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta ggtatctcag ttcggtgtag
     5461 gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc
     5521 ttatccggta actatcgtct tgagtccaac ccggtaagac acgacttatc gccactggca
     5581 gcagccactg gtaacaggat tagcagagcg aggtatgtag gcggtgctac agagttcttg
     5641 aagtggtggc ctaactacgg ctacactaga agaacagtat ttggtatctg cgctctgctg
     5701 aagccagtta ccttcggaaa aagagttggt agctcttgat ccggcaaaca aaccaccgct
     5761 ggtagcggtg gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa
     5821 gaagatcctt tgatcttttc tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa
     5881 gggattttgg tcatgagatt atcaaaaagg atcttcacct agatcctttt aaattaaaaa
     5941 tgaagtttta aatcaatcta aagtatatat gagtaaactt ggtctgacag ttaccaatgc
     6001 ttaatcagtg aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga
     6061 ctccccgtcg tgtagataac tacgatacgg gagggcttac catctggccc cagtgctgca
     6121 atgataccgc gagacccacg ctcaccggct ccagatttat cagcaataaa ccagccagcc
     6181 ggaagggccg agcgcagaag tggtcctgca actttatccg cctccatcca gtctattaat
     6241 tgttgccggg aagctagagt aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc
     6301 attgctacag gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt cagctccggt
     6361 tcccaacgat caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc
     6421 ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag tgttatcact catggttatg
     6481 gcagcactgc ataattctct tactgtcatg ccatccgtaa gatgcttttc tgtgactggt
     6541 gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg
     6601 gcgtcaatac gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga
     6661 aaacgttctt cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg
     6721 taacccactc gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg
     6781 tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt
     6841 tgaatactca tactcttcct ttttcaatat tattgaagca tttatcaggg ttattgtctc
     6901 atgagcggat acatatttga atgtatttag aaaaataaac aaataggggt tccgcgcaca
     6961 tttccccgaa a
//
