LOCUS       sequence-431430-        8224 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  for stable fly cell lines; Hygromycin resistance gene; for genome 
            integration by spontaneous insertion, destination vector.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8224)
  AUTHORS   McGee AV, Liu YV, Griffith AL, Szegletes ZM, Wen B, Kraus C, Miller 
            NW, Steger RJ, Escude Velasco B, Bosch JA, Zirin JD, Viswanatha R, 
            Sontheimer EJ, Goodale A, Greene MA, Green TM, Doench JG
  TITLE     Modular vector assembly enables rapid assessment of emerging CRISPR 
            technologies.
  JOURNAL   Cell Genom. 2024 Mar 13;4(3):100519. doi: 
            10.1016/j.xgen.2024.100519.
  PUBMED    38484704
REFERENCE   2  (bases 1 to 8224)
  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: "Cell 
            Genom."; date: "2024-03-13"; volume: "4(3):100519. doi"; pages: " 
            10.1016/j.xgen.2024.100519"
COMMENT     Sequence Label: pRDB_052
FEATURES             Location/Qualifiers
     source          1..8224
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     polyA_signal    296..430
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(333..352)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     387..406
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(2790..2807)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2961..3549)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3041..3060)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3720..4511,4512..4580))
                     /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     4343..4362
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4581..4685)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     4753..4771
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        6167..6446
                     /label=copia promoter
                     /note="strong promoter from the Drosophila transposable 
                     element copia (Sinclair et al., 1986)"
     CDS             6492..7517
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     E"
     CDS             6492..7517
                     /codon_start=1
                     /gene="aph(4)-Ia"
                     /product="aminoglycoside phosphotransferase from E. coli"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     E"
     intron          7880..7945
                     /label=small t intron
                     /note="SV40 (simian virus 40) small t antigen intron"
     CDS             8075..8095
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
ORIGIN
        1 atggaaaaat attctgtaac ctttataagt aggcataaca gttataatca taacatactg
       61 ttttttctta ctccacacag gcatagagtg tctgctatta ataactatgc tcaaaaattg
      121 tgtaccttta gctttttaat ttgtaaaggg gttaataagg aatatttgat gtatagtgcc
      181 ttgactagag atcataatca gccataccac atttgtagag gttttacttg ctttaaaaaa
      241 cctcccacac ctccccctga acctgaaaca taaaatgaat gcaattgttg ttgttaactt
      301 gtttattgca gcttataatg gttacaaata aagcaatagc atcacaaatt tcacaaataa
      361 agcatttttt tcactgcatt ctagttgtgg tttgtccaaa ctcatcaatg tatcttatca
      421 tgtctggatc cagaacgtct tcaccagtaa tttacgtggg aaaggtgctt gtgtcccaac
      481 aggctaggat ataatgctga aaccgtcccc caagcgttca gggtggggtt tgctacaact
      541 tccgagtcca atgtgtccgt gttcatgata tatatgctca agggcgagaa ttggacctag
      601 ctttcgtgtt agtacgtagc atggtgacac aagcacagta gatcctgccc gccgatcgta
      661 tatattaagt taattcttat ggaatataat aacatgtgga tggccagtgg tcggttgtta
      721 cacgcctacc gcgatgctga atgacccgga ctagagtggc gaaatttatg gcgtgtgacc
      781 cgttatgctc catttcggtc agtgggtcat tgctagtagt cgattgcatt gccattctcc
      841 gagtgattta tggaccccag tgaagacgca ttcactagtt ctagtcgacc atgaagatca
      901 agatcattgc cccgccagag cgcaagtact ctgtctggat cggtggctcc atcctggctt
      961 cgctgtccac cttccagcag atgtggatct ccaagcagga gtacgacgag tccggcccct
     1021 ccattgtgca ccgcaagtgc ttctaagaag gatcgcttgt ctgggcaaga ggatcaggat
     1081 cgggatggtc ttgattctgc tggaggagga ggaggagaag tcgaggaagc agcagcgaaa
     1141 gtgcaagtgc gagtggtgga agtttggagt gcagcacaac aaaatcaaca acaacaccaa
     1201 ctacaagatg aaaagagcgg aaccacctgc acaccatcat cactatcatc atcgttttgg
     1261 gcgcatgttg tgtggttcca gcgtattaat ataattaatt tattccacat gagatatgat
     1321 atgatatact atgtattttt tgtttttttt ttatttgtaa acctttaata taacaagaac
     1381 tacaaaaaat gaaaatgagc gaaaatgcat attctgccat tccacacaca caccaacaac
     1441 acccaacaca cgcacaccca caagcttaca cacacacatt cgcggcatga caaggacatc
     1501 aagataaaga agaacttaaa gaagatattt cccaaagcgc aaaaagaaca cacacacatt
     1561 gcaaaacaca aacaacacac tagcgttttg tacaattcgt cagcaacctt atgtattatt
     1621 ttttaattat gatgtaatta taaacaaagt gaaacaaaaa tatgaaaaca aaaaggaaaa
     1681 tcaaatctct cttctctttc tcccgctctc ctcgctctct gctgctaacc tcgccctctc
     1741 ctctctcatc tttttgtctg tctctcttcc acatttttgc cggccggcaa aataataacc
     1801 cacacacact cacacttggc tgcagtttcg cgtgcgatat tcacacacat tcaagcatac
     1861 aagcatacat acatatgtat ttttttttta tttgtacact tttctaattg catgcgtatc
     1921 gattgataag tttacgcctg aaaatgttaa ttaaaatgtg aaaatgcaac tgaaaaactg
     1981 atgaaatgaa acaacaacaa gcgaacaatt tgctacatgt gtattgtcta acaaccgtta
     2041 ctgcaacggt tgcttcggaa aagggtgaag aggaagaggg cacgaggtcg actagagcgg
     2101 ccgccgacgc gaggctggat ggccttcccc attatgattc ttctcgcttc cggcggcatc
     2161 gggatgcccg cgttgcaggc catgctgtcc aggcaggtag atgacgacca tcagggacag
     2221 cttcaaggat cgctcgcggc tcttaccagc ctaacttcga tcattggacc gctgatcgtc
     2281 acggcgattt atgccgcctc ggcgagcaca tggaacgggt tggcatggat tgtaggcgcc
     2341 gccctatacc ttgtctgcct ccccgcgttg cgtcgcggtg catggagccg ggccacctcg
     2401 acctgaatgg aagccggcgg cacctcgcta acggattcac cactccaaga attggagcca
     2461 atcaattctt gcggagaact gtgaatgcgc aaaccaaccc ttggcagaac atatccatcg
     2521 cgtccgccat ctccagcagc cgcacgcggc gcatctcggg cagcgttggg tcctggccac
     2581 gggtgcgcat gatcgtgctc ctgtcgttga ggacccggct aggctggcgg ggttgcctta
     2641 ctggttagca gaatgaatca ccgatacgcg agcgaacgtg aagcgactgc tgctgcaaaa
     2701 cgtctgcgac ctgagcaaca acatgaatgc tcttcggttt ccgtgtttcg taaagtctgg
     2761 aaacgcggaa gtcagcgctc ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt
     2821 cggctgcggc gagcggtatc agctcactca aaggcggtaa tacggttatc cacagaatca
     2881 ggggataacg caggaaagaa catgtgagca aaaggccagc aaaaggccag gaaccgtaaa
     2941 aaggccgcgt tgctggcgtt tttccatagg ctccgccccc ctgacgagca tcacaaaaat
     3001 cgacgctcaa gtcagaggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc
     3061 cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc
     3121 gcctttctcc cttcgggaag cgtggcgctt tctcatagct cacgctgtag gtatctcagt
     3181 tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac
     3241 cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgacttatcg
     3301 ccactggcag cagccactgg taacaggatt agcagagcga ggtatgtagg cggtgctaca
     3361 gagttcttga agtggtggcc taactacggc tacactagaa ggacagtatt tggtatctgc
     3421 gctctgctga agccagttac cttcggaaaa agagttggta gctcttgatc cggcaaacaa
     3481 accaccgctg gtagcggtgg tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa
     3541 ggatctcaag aagatccttt gatcttttct acggggtctg acgctcagtg gaacgaaaac
     3601 tcacgttaag ggattttggt catgagatta tcaaaaagga tcttcaccta gatcctttta
     3661 aattaaaaat gaagttttaa atcaatctaa agtatatatg agtaaacttg gtctgacagt
     3721 taccaatgct taatcagtga ggcacctatc tcagcgatct gtctatttcg ttcatccata
     3781 gttgcctgac tccccgtcgt gtagataact acgatacggg agggcttacc atctggcccc
     3841 agtgctgcaa tgataccgcg agacccacgc tcaccggctc cagatttatc agcaataaac
     3901 cagccagccg gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag
     3961 tctattaatt gttgccggga agctagagta agtagttcgc cagttaatag tttgcgcaac
     4021 gttgttgcca ttgctgcagg catcgtggtg tcacgctcgt cgtttggtat ggcttcattc
     4081 agctccggtt cccaacgatc aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg
     4141 gttagctcct tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt gttatcactc
     4201 atggttatgg cagcactgca taattctctt actgtcatgc catccgtaag atgcttttct
     4261 gtgactggtg agtactcaac caagtcattc tgagaatagt gtatgcggcg accgagttgc
     4321 tcttgcccgg cgtcaacacg ggataatacc gcgccacata gcagaacttt aaaagtgctc
     4381 atcattggaa aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc
     4441 agttcgatgt aacccactcg tgcacccaac tgatcttcag catcttttac tttcaccagc
     4501 gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca
     4561 cggaaatgtt gaatactcat actcttcctt tttcaatatt attgaagcat ttatcagggt
     4621 tattgtctca tgagcggata catatttgaa tgtatttaga aaaataaaca aataggggtt
     4681 ccgcgcacat ttccccgaaa agtgccacct gacgtctaag aaaccattat tatcatgaca
     4741 ttaacctata aaaataggcg tatcacgagg ccctttcctc ttcaagaatt cacatttgta
     4801 cgaatttttt ttttatcaaa agttcgagtt tttcaccaat ttcctcatca accgagcaag
     4861 gcaaacggct ttgaataata tggtgttata tatacatata tcaaatcgct gctgactgcg
     4921 tgattgatgg ccccaagatt acatattatc gaatcaggat tcagaaggag atcaatgtca
     4981 aatgcggaca ggaacatgaa agacgcctgt tatgcgcaat taaaaatttg ggtttaattg
     5041 ctgtggaaac tgttgttggc ggcatcttaa gttcctgttt aacaacatca actacttatg
     5101 tacgtagaag cgtttaagcc atttgcatac agatgagaac tggcttttgt gctaatcagt
     5161 caagatgact ccgatgatga tgactcatta cctgaccagt tttcgctgct ttcttttcaa
     5221 caactacttg tatatgtatt gtatccaata gcaatacatt gaatttccat ggtctagtca
     5281 cgtattatca tttaattgac accaagtcgt gttattgttg agctatcgag ttcagctcaa
     5341 acatttctta ttcccatgaa taagccggca aaaatatgca atctatgaaa gttaatataa
     5401 gcaaacctta ctttgactca ataccaatgc actttgtgtc gataggttca cgcaattgag
     5461 gcgattattc cgataaccca agcgattgac tgttcccgtt tcgattccaa ttgaaatttg
     5521 gaaatgtaca atagttttgc tatatgctgt caagtacgct cttatcttct ctgggttttc
     5581 ttcagagttt cgaaacgctt cttctttttt ttgttttttt ttttttggaa tctcgtattt
     5641 tggaaggggc tcccctctgg aatttgttac actgtcgtta tcattgcgaa caagcggccc
     5701 gaagctatca gcgactttaa catttacaat gcactttttt acgaccaatt aaatgtacat
     5761 tttcctttct tcgcccgttg ataagcgaac gcgatgtggc gcaggcaatg tgttgctctt
     5821 gcgacacaaa cgcaatcaaa atggattcaa tttcgctttt tcccagtgaa acgaagaacg
     5881 aaccgaccat catgatatgc tcctctgcat gttgcgtatt gaatcaatga caatttcaat
     5941 taagccgccc gttcgtcatg cgttttcgtg cgcttcgaaa tgctgataac gctgctgtcc
     6001 tccaactgct ttgcatgtgg acacaattcc atttatttaa ttcttttatt tggatcggtt
     6061 aaattaaaaa gcgccttgtt acgcatttaa cgttgtttcc ggtgcgtggt ggtttcatgc
     6121 ttctgggaac ggcaaatggg tttaggattg ggaacccctc atcatctgtt ggaatatact
     6181 attcaaccta caaaagtaac gttaaacaac actactttat atttgatatg aatggccaca
     6241 ccttttatgc cataaaacat attgtaagag aataccactc tttttattcc ttctttcctt
     6301 cttgtacgtt ttttgctgta agtaggtcgt ggtgctggtg ttgcagttga aataacttaa
     6361 aatataaatc ataaaactca aacataaact tgactattta tttatttatt aagaaaggaa
     6421 atataaatta taaattacaa caggttatgg acctgcagcc aagcttggcg ctcgtccggg
     6481 ggcaatgaga tatgaaaaag cctgaactca ccgcgacgtc tgtcgagaag tttctgatcg
     6541 aaaagttcga cagcgtgtcc gacctgatgc agctctcgga gggcgaagaa tctcgtgctt
     6601 tcagcttcga tgtaggaggg cgtggatatg tcctgcgggt aaatagctgc gccgatggtt
     6661 tctacaaaga tcgttatgtt tatcggcact ttgcatcggc cgcgctcccg attccggaag
     6721 tgcttgacat tggggaattc agcgagagcc tgacctattg catctcccgc cgtgcacagg
     6781 gtgtcacgtt gcaagacctg cctgaaaccg aactgcccgc tgttctgcag ccggtcgcgg
     6841 aggccatgga tgcgatcgct gcggccgatc ttagccagac gagcgggttc ggcccattcg
     6901 gaccgcaagg aatcggtcaa tacactacat ggcgtgattt catatgcgcg attgctgatc
     6961 cccatgtgta tcactggcaa actgtgatgg acgacaccgt cagtgcgtcc gtcgcgcagg
     7021 ctctcgatga gctgatgctt tgggccgagg actgccccga agtccggcac ctcgtgcacg
     7081 cggatttcgg ctccaacaat gtcctgacgg acaatggccg cataacagcg gtcattgact
     7141 ggagcgaggc gatgttcggg gattcccaat acgaggtcgc caacatcttc ttctggaggc
     7201 cgtggttggc ttgtatggag cagcagacgc gctacttcga gcggaggcat ccggagcttg
     7261 caggatcgcc gcggctccgg gcgtatatgc tccgcattgg tcttgaccaa ctctatcaga
     7321 gcttggttga cggcaatttc gatgatgcag cttgggcgca gggtcgatgc gacgcaatcg
     7381 tccgatccgg agccgggact gtcgggcgta cacaaatcgc ccgcagaagc gcggccgtct
     7441 ggaccgatgg ctgtgtagaa gtactcgccg atagtggaaa ccgacgcccc agcactcgtc
     7501 cgagggcaaa ggaatagagt agatgccgac cgaacaagag ctgatttcga gaacgcctca
     7561 gccagcaact cgcgcgagcc tagcaaggca aatgcgagag aacggcctta cgcttggtgg
     7621 cacagttctc gtccacagtt cgctaagctc gctcggctgg gtcgcgggag ggccggtcgc
     7681 agtgattcag gcccttctgg attgtgttgg tccccagggc acgattgtca tgcccacgca
     7741 ctcgggtgat ctgactgatc ccgcagattg gagatcgccg cccgtgcctg ccgattgggt
     7801 gcagatcttt gtgaaggaac cttacttctg tggtgtgaca taattggaca aactacctac
     7861 agagatttaa agctctaagg taaatataaa atttttaagt gtataatgtg ttaaactact
     7921 gattctaatt gtttgtgtat tttagattcc aacctatgga actgatgaat gggagcagtg
     7981 gtggaatgcc tttaatgagg aaaacctgtt ttgctcagaa gaaatgccat ctagtgatga
     8041 tgaggctact gctgactctc aacattctac tcctccaaaa aagaagagaa aggtagaaga
     8101 tcccaaggac tttccttcag aattgctaag ttttttgagt catgctgtgt ttagtaatag
     8161 aactcttgct tgctttgcta tttacaccac aaaggaaaaa gctgcactgc tatacaagaa
     8221 aatt
//
