LOCUS       sequence-197482-        9822 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of human RPA with an alanine substituted phosphorylation 
            domain (with the following mutations in RPA32 
            :S8,11-13,23,29,33,39A,T21A)..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9822)
  TITLE     p11d-tRPA-32Ala9
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 9822)
  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"; journalName: 
            "Unpublished"
COMMENT     Sequence Label: p11d-tRPA-32Ala9
FEATURES             Location/Qualifiers
     source          1..9822
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(8..26)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        94..198
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(199..267,268..1059)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(417..436)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      1230..1818
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1719..1738
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     1972..1989
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    2004..2143
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(2069..2088)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2229..2251
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(2245..2436)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     4791..4810
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(4830..4907)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     5113..5132
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     5216..5235
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        5216..5234
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    5235..5259
                     /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)."
     RBS             5274..5296
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     RBS             7662..7684
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     RBS             8391..8413
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     primer_bind     complement(9348..9366)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      9362..9409
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     promoter        complement(join(9775..9780,9781..9797,9798..9803))
                     /label=tet promoter
                     /note="E. coli promoter for tetracycline efflux protein 
                     gene"
ORIGIN
        1 cgaaagggcc tcgtgatacg cctattttta taggttaatg tcatgataat aatggtttct
       61 tagacgtcag gtggcacttt tcggggaaat gtgcgcggaa cccctatttg tttatttttc
      121 taaatacatt caaatatgta tccgctcatg agacaataac cctgataaat gcttcaataa
      181 tattgaaaaa ggaagagtat gagtattcaa catttccgtg tcgcccttat tccctttttt
      241 gcggcatttt gccttcctgt ttttgctcac ccagaaacgc tggtgaaagt aaaagatgct
      301 gaagatcagt tgggtgcacg agtgggttac atcgaactgg atctcaacag cggtaagatc
      361 cttgagagtt ttcgccccga agaacgtttt ccaatgatga gcacttttaa agttctgcta
      421 tgtggcgcgg tattatcccg tgttgacgcc gggcaagagc aactcggtcg ccgcatacac
      481 tattctcaga atgacttggt tgagtactca ccagtcacag aaaagcatct tacggatggc
      541 atgacagtaa gagaattatg cagtgctgcc ataaccatga gtgataacac tgcggccaac
      601 ttacttctga caacgatcgg aggaccgaag gagctaaccg cttttttgca caacatgggg
      661 gatcatgtaa ctcgccttga tcgttgggaa ccggagctga atgaagccat accaaacgac
      721 gagcgtgaca ccacgatgcc tgcagcaatg gcaacaacgt tgcgcaaact attaactggc
      781 gaactactta ctctagcttc ccggcaacaa ttaatagact ggatggaggc ggataaagtt
      841 gcaggaccac ttctgcgctc ggcccttccg gctggctggt ttattgctga taaatctgga
      901 gccggtgagc gtgggtctcg cggtatcatt gcagcactgg ggccagatgg taagccctcc
      961 cgtatcgtag ttatctacac gacggggagt caggcaacta tggatgaacg aaatagacag
     1021 atcgctgaga taggtgcctc actgattaag cattggtaac tgtcagacca agtttactca
     1081 tatatacttt agattgattt aaaacttcat ttttaattta aaaggatcta ggtgaagatc
     1141 ctttttgata atctcatgac caaaatccct taacgtgagt tttcgttcca ctgagcgtca
     1201 gaccccgtag aaaagatcaa aggatcttct tgagatcctt tttttctgcg cgtaatctgc
     1261 tgcttgcaaa caaaaaaacc accgctacca gcggtggttt gtttgccgga tcaagagcta
     1321 ccaactcttt ttccgaaggt aactggcttc agcagagcgc agataccaaa tactgtcctt
     1381 ctagtgtagc cgtagttagg ccaccacttc aagaactctg tagcaccgcc tacatacctc
     1441 gctctgctaa tcctgttacc agtggctgct gccagtggcg ataagtcgtg tcttaccggg
     1501 ttggactcaa gacgatagtt accggataag gcgcagcggt cgggctgaac ggggggttcg
     1561 tgcacacagc ccagcttgga gcgaacgacc tacaccgaac tgagatacct acagcgtgag
     1621 ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc
     1681 agggtcggaa caggagagcg cacgagggag cttccagggg gaaacgcctg gtatctttat
     1741 agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg
     1801 gggcggagcc tatggaaaaa cgccagcaac gcggcctttt tacggttcct ggccttttgc
     1861 tggccttttg ctcacatgtt ctttcctgcg ttatcccctg attctgtgga taaccgtatt
     1921 accgcctttg agtgagctga taccgctcgc cgcagccgaa cgaccgagcg cagcgagtca
     1981 gtgagcgagg aagcggaaga gcgcctgatg cggtattttc tccttacgca tctgtgcggt
     2041 atttcacacc gcaatggtgc actctcagta caatctgctc tgatgccgca tagttaagcc
     2101 agtatacact ccgctatcgc tacgtgactg ggtcatggct gcgccccgac acccgccaac
     2161 acccgctgac gcgccctgac gggcttgtct gctcccggca tccgcttaca gacaagctgt
     2221 gaccgtctcc gggagctgca tgtgtcagag gttttcaccg tcatcaccga aacgcgcgag
     2281 gcagctgcgg taaagctcat cagcgtggtc gtgaagcgat tcacagatgt ctgcctgttc
     2341 atccgcgtcc agctcgttga gtttctccag aagcgttaat gtctggcttc tgataaagcg
     2401 ggccatgtta agggcggttt tttcctgttt ggtcactgat gcctccgtgt aagggggatt
     2461 tctgttcatg ggggtaatga taccgatgaa acgagagagg atgctcacga tacgggttac
     2521 tgatgatgaa catgcccggt tactggaacg ttgtgagggt aaacaactgg cggtatggat
     2581 gcggcgggac cagagaaaaa tcactcaggg tcaatgccag cgcttcgtta atacagatgt
     2641 aggtgttcca cagggtagcc agcagcatcc tgcgatgcag atccggaaca taatggtgca
     2701 gggcgctgac ttccgcgttt ccagacttta cgaaacacgg aaaccgaaga ccattcatgt
     2761 tgttgctcag gtcgcagacg ttttgcagca gcagtcgctt cacgttcgct cgcgtatcgg
     2821 tgattcattc tgctaaccag taaggcaacc ccgccagcct agccgggtcc tcaacgacag
     2881 gagcacgatc atgcgcaccc gtggccagga cccaacgctg cccgagatgc gccgcgtgcg
     2941 gctgctggag atggcggacg cgatggatat gttctgccaa gggttggttt gcgcattcac
     3001 agttctccgc aagaattgat tggctccaat tcttggagtg gtgaatccgt tagcgaggtg
     3061 ccgccggctt ccattcaggt cgaggtggcc cggctccatg caccgcgacg caacgcgggg
     3121 aggcagacaa ggtatagggc ggcgcctaca atccatgcca acccgttcca tgtgctcgcc
     3181 gaggcggcat aaatcgccgt gacgatcagc ggtccaatga tcgaagttag gctggtaaga
     3241 gccgcgagcg atccttgaag ctgtccctga tggtcgtcat ctacctgcct ggacagcatg
     3301 gcctgcaacg cgggcatccc gatgccgccg gaagcgagaa gaatcataat ggggaaggcc
     3361 atccagcctc gcgtcgcgaa cgccagcaag acgtagccca gcgcgtcggc cgccatgccg
     3421 gcgataatgg cctgcttctc gccgaaacgt ttggtggcgg gaccagtgac gaaggcttga
     3481 gcgagggcgt gcaagattcc gaataccgca agcgacaggc cgatcatcgt cgcgctccag
     3541 cgaaagcggt cctcgccgaa aatgacccag agcgctgccg gcacctgtcc tacgagttgc
     3601 atgataaaga agacagtcat aagtgcggcg acgatagtca tgccccgcgc ccaccggaag
     3661 gagctgactg ggttgaaggc tctcaagggc atcggtcgag atcccggtgc ctaatgagtg
     3721 agctaactta cattaattgc gttgcgctca ctgcccgctt tccagtcggg aaactgtcgt
     3781 gccagctgca ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt attgggcgcc
     3841 agggtggttt ttcttttcac cagtgagacg ggcaacagct gattgccctt caccgcctgg
     3901 ccctgagaga gttgcagcaa gcggtccacg ctggtttgcc ccagcaggcg aaaatcctgt
     3961 ttgatggtgg ttaacggcgg gatataacat gagctgtctt cggtatcgtc gtatcccact
     4021 accgagatat ccgcaccaac gcgcagcccg gactcggtaa tggcgcgcat tgcgcccagc
     4081 gccatctgat cgttggcaac cagcatcgca gtgggaacga tgccctcatt cagcatttgc
     4141 atggtttgtt gaaaaccgga catggcactc cagtcgcctt cccgttccgc tatcggctga
     4201 atttgattgc gagtgagata tttatgccag ccagccagac gcagacgcgc cgagacagaa
     4261 cttaatgggc ccgctaacag cgcgatttgc tggtgaccca atgcgaccag atgctccacg
     4321 cccagtcgcg taccgtcttc atgggagaaa ataatactgt tgatgggtgt ctggtcagag
     4381 acatcaagaa ataacgccgg aacattagtg caggcagctt ccacagcaat ggcatcctgg
     4441 tcatccagcg gatagttaat gatcagccca ctgacgcgtt gcgcgagaag attgtgcacc
     4501 gccgctttac aggcttcgac gccgcttcgt tctaccatcg acaccaccac gctggcaccc
     4561 agttgatcgg cgcgagattt aatcgccgcg acaatttgcg acggcgcgtg cagggccaga
     4621 ctggaggtgg caacgccaat cagcaacgac tgtttgcccg ccagttgttg tgccacgcgg
     4681 ttgggaatgt aattcagctc cgccatcgcc gcttccactt tttcccgcgt tttcgcagaa
     4741 acgtggctgg cctggttcac cacgcgggaa acggtctgat aagagacacc ggcatactct
     4801 gcgacatcgt ataacgttac tggtttcaca ttcaccaccc tgaattgact ctcttccggg
     4861 cgctatcatg ccataccgcg aaaggttttg cgccattcga tggtgtccgg gatctcgacg
     4921 ctctccctta tgcgactcct gcattaggaa gcagcccagt agtaggttga ggccgttgag
     4981 caccgccgcc gcaaggaatg gtgcatgcaa ggagatggcg cccaacagtc ccccggccac
     5041 ggggcctgcc accataccca cgccgaaaca agcgctcatg agcccgaagt ggcgagcccg
     5101 atcttcccca tcggtgatgt cggcgatata ggcgccagca accgcacctg tggcgccggt
     5161 gatgccggcc acgatgcgtc cggcgtagag gatcgagatc tcgatcccgc gaaattaata
     5221 cgactcacta taggggaatt gtgagcggat aacaattccc ctctagaaat aattttgttt
     5281 aactttaaga aggagatata ccatggtcgg ccagctgagc gagggggcca ttgcggccat
     5341 catgcagaag ggggatacaa acataaagcc catcctccaa gtcatcaaca tccgtcccat
     5401 tactacgggg aatagtccgc cgcgttatcg actgctcatg agtgatggat tgaacactct
     5461 atcctctttc atgttggcga cacagttgaa ccctctcgtg gaggaagaac aattgtccag
     5521 caactgtgta tgccagattc acagatttat tgtgaacact ctgaaagacg gaaggagagt
     5581 agttatcttg atggaattag aagttttgaa gtcagctgaa gcagttggag tgaagattgg
     5641 caatccagtg ccctataatg aaggactcgg gcagccgcaa gtagctcctc cagcgccagc
     5701 agccagccca gcagcaagca gcaggcccca gccgcagaat ggaagctcgg gaatgggttc
     5761 tactgtttct aaggcttatg gtgcttcaaa gacatttgga aaagctgcag gtcccagcct
     5821 gtcacacact tctgggggaa cacagtccaa agtggtgccc attgccagcc tcactcctta
     5881 ccagtccaag tggaccattt gtgctcgtgt taccaacaaa agtcagatcc gtacctggag
     5941 caactcccga ggggaaggga agcttttctc cctagaactg gttgacgaaa gtggtgaaat
     6001 ccgagctaca gctttcaatg agcaagtgga caagttcttt cctcttattg aagtgaacaa
     6061 ggtgtattat ttctcgaaag gcaccctgaa gattgctaac aagcagttca cagctgttaa
     6121 aaatgactac gagatgacct tcaataacga gacttccgtc atgccctgtg aggacgacca
     6181 tcatttacct acggttcagt ttgatttcac ggggattgat gacctcgaga acaagtcgaa
     6241 agactcactt gtagacatca tcgggatctg caagagctat gaagacgcca ctaaaatcac
     6301 agtgaggtct aacaacagag aagttgccaa gaggaatatc tacttgatgg acacatccgg
     6361 gaaggtggtg actgctacac tgtgggggga agatgctgat aaatttgatg gttctagaca
     6421 gcccgtgttg gctatcaaag gagcccgagt ctctgatttc ggtggacgga gcctctccgt
     6481 gctgtcttca agcactatca ttgcgaatcc tgacatccca gaggcctata agcttcgtgg
     6541 atggtttgac gcagaaggac aagccttaga tggtgtttcc atctctgatc taaagagcgg
     6601 cggagtcgga gggagtaaca ccaactggaa aaccttgtat gaggtcaaat ccgagaacct
     6661 gggccaaggc gacaagccgg actactttag ttctgtggcc acagtggtgt atcttcgcaa
     6721 agagaactgc atgtaccaag cctgcccgac tcaggactgc aataagaaag tgattgatca
     6781 acagaatgga ttgtaccgct gtgagaagtg cgacaccgaa tttcccaatt tcaagtaccg
     6841 catgatcctg tcagtaaata ttgcagattt tcaagagaat cagtgggtga cttgtttcca
     6901 ggagtctgct gaagctatcc ttggacaaaa tgctgcttat cttggggaat taaaagacaa
     6961 gaatgaacag gcatttgaag aagttttcca gaatgccaac ttccgatctt tcatattcag
     7021 agtcagggtc aaagtggaga cctacaacga cgagtctcga attaaggcca ctgtgatgga
     7081 cgtgaagccc gtggactaca gagagtatgg ccgaaggctg gtcatgagca tcaggagaag
     7141 tgcattgatg tgagaggagc agtgccaatc gggcagaagt ttgcaaatag gcagaatgga
     7201 atcgatttcc tcccacctcc gtgtgacgat cccatgttag ctacacagtg cagaggctct
     7261 tgatggtgga ctaagcaatt tcctccctcg tgcgcatctc agaacccatc ggtaggcaaa
     7321 ggaaaatacg ctcaggtggt tgtggtgtag actgtgtcag gcctacggag tcagccagtg
     7381 gctagcgcaa gaccagtcac tccctctgcc ttcaggcttc tgtcaatttc attatcatca
     7441 agcaggaatt atgtcgtaag tcactgaccc taactgcaga ccatgaagta aattatgtaa
     7501 ctaggttttt gcttctccag tggtgaccac ccccccccat ccccgctcac aacttgggtt
     7561 cttctcagcg gggcgagctg agaagcggtc atgagcacct ggggatttta gtaagtgtgt
     7621 cttcctagaa ttcctgcagc ccgggggatc ctagaaataa ttttgtttaa ctttaagaag
     7681 gagatatacc atggtggaca tgatggactt gcccaggtcg cgcatcaacg ccggcatgct
     7741 agctcaattc atcgacaagc ctgtctgctt cgtagggagg ctggaaaaga ttcatcccac
     7801 cggaaaaatg tttattcttt cagatggaga aggaaaaaat ggaaccatcg agttgatgga
     7861 accccttgat gaagaaatct ctggaattgt ggaagtggtt ggaagagtaa ccgccaaggc
     7921 caccatcttg tgtacatctt atgtccagtt taaagaagat agccatcctt ttgatcttgg
     7981 actttacaat gaagctgtga aaattatcca tgacttccct cagttttatc ctttagggat
     8041 tgtgcaacat gattgatctt gatggatttt catacgattg taaatgagct atattaaagt
     8101 ctattaaagg aagcccttct tgtttgaggg agagatttct gtgctttctc atatttaatt
     8161 tgctgttttt aagatattcc aacctagagt ttttgatgga actgatatat tgacagttct
     8221 caccgaagtc cttttataaa gaattgctac tccaatatat ggtcagatta gatgcaagaa
     8281 taaagcagtt gtccgagtct aagtttctat tttattaata aaaactaaaa tggtacgtac
     8341 aaaaaaaaaa ccggaattcg agctcggtac ccggggatcc tagaaataat tttgtttaac
     8401 tttaagaagg agatatacat atgtggaaca gtggattcga agcctatggc gccgcggcat
     8461 acgggggagc cggcggctac gcgcaggcgc cggggggctt tggagcgccc gcacctgctc
     8521 aagccgaaaa gaaagcaaga gcccgagccc agcacattgt gccctgtact atatctcagc
     8581 tgctttctgc cactttggtt gatgaagtgt tcagaattgg gaatgttgag atttcacagg
     8641 tcactattgt ggggatcatc agacatgcag agaaggctcc aaccaacatt gtttacaaaa
     8701 tagatgacat gacagctgca cccatggacg ttcgccagtg ggttgacaca gatgacacca
     8761 gcagtgaaaa cactgtggtt cctccagaaa catatgtgaa agtggcaggc cacctgagat
     8821 cttttcagaa caaaaagagc ctggtagcct ttaagatcat gcccctggag gatatgaatg
     8881 agttcaccac acatattctg gaagtgatca atgcacacat ggtactaagc aaagccaaca
     8941 gccagccctc agcagggaga gcacctatca gcaatccagg aatgagtgaa gcagggaact
     9001 ttggtgggaa tagcttcatg ccagcaaatg gcctcactgt ggcccaaaac caggtgttga
     9061 atttgattaa ggcttgtcca agacctgaag ggttgaactt tcaggatctc aagaaccagc
     9121 tgaaacacat gtctgtatcc tcaatcaagc aagctgtgga ttttctgagc aatgaggggc
     9181 acatctattc tactgtggat gatgaccatt ttaaatccac agatgcagaa taactggatc
     9241 taactgggta cctgagatat tttacagctg gacctagttt cacaatctgt tgtctccgga
     9301 tccggctgct aacaaagccc gaaaggaagc tgagttggct gctgccaccg ctgagcaata
     9361 actagcataa ccccttgggg cctctaaacg ggtcttgagg ggttttttgc tgaaaggagg
     9421 aactatatcc ggatatccac aggacgggtg tggtcgccat gatcgcgtag tcgatagtgg
     9481 ctccaagtag cgaagcgagc aggactgggc ggcggccaaa gcggtcggac agtgctccga
     9541 gaacgggtgc gcatagaaat tgcatcaacg catatagcgc tagcagcacg ccatagtgac
     9601 tggcgatgct gtcggaatgg acgatatccc gcaagaggcc cggcagtacc ggcataacca
     9661 agcctatgcc tacagcatcc agggtgacgg tgccgaggat gacgatgagc gcattgttag
     9721 atttcataca cggtgcctga ctgcgttagc aatttaactg tgataaacta ccgcattaaa
     9781 gcttatcgat gataagctgt caaacatgag aattcttgaa ga
//
