pDRGFP

LOCUS       pDRGFP               8646 bp ds-DNA     circular SYN 06-AUG-2018
DEFINITION  In vivo homologous recombination substrate composed of two 
            differentially mutated GFP genes oriented as direct repeats and 
            separated by a drug selection marker .
ACCESSION   .
VERSION     .
KEYWORDS    pDRGFP
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8646)
  AUTHORS   Pierce AJ, Johnson RD, Thompson LH, Jasin M
  TITLE     XRCC3 promotes homology-directed repair of DNA damage in mammalian 
            cells.
  JOURNAL   Genes Dev. 1999 Oct 15. 13(20):2633-8.
  PUBMED    10541549
REFERENCE   2  (bases 1 to 8646)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Aug 6, 2018 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..8646
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     enhancer        4..383
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        386..661
                     /label=chicken beta-actin promoter
     intron          662..1670
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     1594..1616
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1678..1697
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     CDS             1755..1775
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             1797..1817
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     regulatory      2031..2040
                     /regulatory_class="other"
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     primer_bind     2690..2711
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    2879..3000
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     CDS             complement(3661..4260)
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(3744..3764)
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     primer_bind     4241..4260
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     promoter        complement(4279..4776)
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     complement(4327..4346)
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     primer_bind     4718..4741
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     primer_bind     complement(4864..4883)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_site      4929..4984
                     /label=-beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal"
     primer_bind     complement(4983..5002)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     CDS             5346..5366
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             5388..5408
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     regulatory      5622..5631
                     /regulatory_class="other"
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     primer_bind     complement(6149..6165)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6149..6165)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(6162..6184)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    6173..6189
                     /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(6197..6227)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6242..6263
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        6322..6517
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      6368..6503
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     6430..6449
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     polyA_signal    6523..6657
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6725..6742)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6896..7484)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6976..6995)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(7655..8515)
                     /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     8278..8297
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(8516..8620)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gtcgacattg attattgact agttattaat agtaatcaat tacggggtca ttagttcata
       61 gcccatatat ggagttccgc gttacataac ttacggtaaa tggcccgcct ggctgaccgc
      121 ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt tcccatagta acgccaatag
      181 ggactttcca ttgacgtcaa tgggtggagt atttacggta aactgcccac ttggcagtac
      241 atcaagtgta tcatatgcca agtccgcccc ctattgacgt caatgacggt aaatggcccg
      301 cctggcatta tgcccagtac atgaccttac gggactttcc tacttggcag tacatctacg
      361 tattagtcat cgctattacc atgggtcgag gtgagcccca cgttctgctt cactctcccc
      421 atctcccccc cctccccacc cccaattttg tatttattta ttttttaatt attttgtgca
      481 gcgatggggg cggggggggg gggggcgcgc gccaggcggg gcggggcggg gcgaggggcg
      541 gggcggggcg aggcggagag gtgcggcggc agccaatcag agcggcgcgc tccgaaagtt
      601 tccttttatg gcgaggcggc ggcggcggcg gccctataaa aagcgaagcg cgcggcgggc
      661 gggagtcgct gcgttgcctt cgccccgtgc cccgctccgc gccgcctcgc gccgcccgcc
      721 ccggctctga ctgaccgcgt tactcccaca ggtgagcggg cgggacggcc cttctcctcc
      781 gggctgtaat tagcgcttgg tttaatgacg gctcgtttct tttctgtggc tgcgtgaaag
      841 ccttaaaggg ctccgggagg gccctttgtg cgggggggag cggctcgggg ggtgcgtgcg
      901 tgtgtgtgtg cgtggggagc gccgcgtgcg gcccgcgctg cccggcggct gtgagcgctg
      961 cgggcgcggc gcggggcttt gtgcgctccg cgtgtgcgcg aggggagcgc ggccgggggc
     1021 ggtgccccgc ggtgcggggg ggctgcgagg ggaacaaagg ctgcgtgcgg ggtgtgtgcg
     1081 tgggggggtg agcagggggt gtgggcgcgg cggtcgggct gtaacccccc cctgcacccc
     1141 cctccccgag ttgctgagca cggcccggct tcgggtgcgg ggctccgtgc ggggcgtggc
     1201 gcggggctcg ccgtgccggg cggggggtgg cggcaggtgg gggtgccggg cggggcgggg
     1261 ccgcctcggg ccggggaggg ctcgggggag gggcgcggcg gccccggagc gccggcggct
     1321 gtcgaggcgc ggcgagccgc agccattgcc ttttatggta atcgtgcgag agggcgcagg
     1381 gacttccttt gtcccaaatc tggcggagcc gaaatctggg aggcgccgcc gcaccccctc
     1441 tagcgggcgc gggcgaagcg gtgcggcgcc ggcaggaagg aaatgggcgg ggagggcctt
     1501 cgtgcgtcgc cgcgccgccg tccccttctc catctccagc ctcggggctg ccgcaggggg
     1561 acggctgcct tcggggggga cggggcaggg cggggttcgg cttctggcgt gtgaccggcg
     1621 gctctagagc ctctgctaac catgttcatg ccttcttctt tttcctacag ctcctgggca
     1681 acgtgctggt tattgtgctg tctcatcatt ttggcaaaga attcagatcc gccgccacta
     1741 tgggatcaag atcgccaaaa aagaagagaa aggtgccgaa gaagcatgca gcaccaccaa
     1801 aaaaaaaacg aaaagtagaa gacccacgag gcaacaccag cggcgtgctg agcaccccca
     1861 aggccaagag ggccaagcac ccccccggca ccgagaagcc caggagcagg agccagagcg
     1921 agcagcccgc cacctgcccc atctgctacg ccgtgatcag gcagagcagg aacctgagga
     1981 ggcacctgga gctgaggcac ttcgccaagc ccggcgtgga tccaccggtc gccaccatgg
     2041 tgagcaaggg cgaggagctg ttcaccgggg tggtgcccat cctggtcgag ctggacggcg
     2101 acgtaaacgg ccacaagttc agcgtgtccg gctagggata acagggtaat acctacggca
     2161 agctgaccct gaagttcatc tgcaccaccg gcaagctgcc cgtgccctgg cccaccctcg
     2221 tgaccaccct gacctacggc gtgcagtgct tcagccgcta ccccgaccac atgaagcagc
     2281 acgacttctt caagtccgcc atgcccgaag gctacgtcca ggagcgcacc atcttcttca
     2341 aggacgacgg caactacaag acccgcgccg aggtgaagtt cgagggcgac accctggtga
     2401 accgcatcga gctgaagggc atcgacttca aggaggacgg caacatcctg gggcacaagc
     2461 tggagtacaa ctacaacagc cacaacgtct atatcatggc cgacaagcag aagaacggca
     2521 tcaaggtgaa cttcaagatc cgccacaaca tcgaggacgg cagcgtgcag ctcgccgacc
     2581 actaccagca gaacaccccc atcggcgacg gccccgtgct gctgcccgac aaccactacc
     2641 tgagcaccca gtccgccctg agcaaagacc ccaacgagaa gcgcgatcac atggtcctgc
     2701 tggagttcgt gaccgccgcc gggatcactc tcggcatgga cgagctgtac aagtaaagcg
     2761 gccgcgactc tagatcataa tcagccatac cacatttgta gaggttttac ttgctttaaa
     2821 aaacctccca cacctccccc tgaacctgaa acataaaatg aatgcaattg ttgttgttaa
     2881 cttgtttatt gcagcttata atggttacaa ataaagcaat agcatcacaa atttcacaaa
     2941 taaagcattt ttttcactgc attctagttg tggtttgtcc aaactcatca atgtatctta
     3001 aggcgtaaat tgtaagcgtt aatctgaagc ttctgatgga attagaactt ggcaaaacaa
     3061 tactgagaat gaagtgtatg tggaacagag gctgctgatc tcgttcttca ggctatgaaa
     3121 ctgacacatt tggaaaccac agtacttaga accacaaagt gggaatcaag agaaaaacaa
     3181 tgatcccacg agagatctat agatctatag atcatgagtg ggaggaatga gctggccctt
     3241 aatttggttt tgcttgttta aattatgata tccaactatg aaacattatc ataaagcaat
     3301 agtaaagagc cttcagtaaa gagcaggcat ttatctaatc ccaccccacc cccacccccg
     3361 tagctccaat ccttccattc aaaatgtagg tactctgttc tcacccttct taacaaagta
     3421 tgacaggaaa aacttccatt ttagtggaca tctttattgt ttaatagatc atcaatttcg
     3481 atccgctcct gggcaccgaa ctgcgccgcg tgttcagcag ggtcggcgtg ttcggtgtgt
     3541 cccccgcggt gggcctcggg ggcgggtgcg gggtcggcgg ggccgccccg ggtggcttcg
     3601 gtcggagcca tggggtcgtg cgctcctttc ggtcgggcgc tgcgggtcgt ggggcgggcg
     3661 tcaggcaccg ggcttgcggg tcatgcacca ggtgcgcggt ccttcgggca cctcgacgtc
     3721 ggcggtgacg gtgaagccga gccgctcgta gaaggggagg ttgcggggcg cggaggtctc
     3781 caggaaggcg ggcaccccgg cgcgctcggc cgcctccact ccggggagca cgacggcgct
     3841 gcccagaccc ttgccctggt ggtcgggcga gacgccgacg gtggccagga accacgcggg
     3901 ctccttgggc cggtgcggcg ccaggaggcc ttccatctgt tgctgcgcgg ccagccggga
     3961 accgctcaac tcggccatgc gcgggccgat ctcggcgaac accgcccccg cttcgacgct
     4021 ctccggcgtg gtccagaccg ccaccgcggc gccgtcgtcc gcgacccaca ccttgccgat
     4081 gtcgagcccg acgcgcgtga ggaagagttc ttgcagctcg gtgacccgct cgatgtggcg
     4141 gtccgggtcg acggtgtggc gcgtggcggg gtagtcggcg aacgcggcgg cgagggtgcg
     4201 tacggcccgg gggacgtcgt cgcgggtggc gaggcgcacc gtgggcttgt actcggtcat
     4261 ggtggcggct ggatcggtcg aaaggcccgg agatgaggaa gaggagaaca gcgcggcaga
     4321 cgtgcgcttt tgaagcgtgc agaatgccgg gcctccggag gaccttcggg cgcccgcccc
     4381 gcccctgagc ccgcccctga gcccgccccc ggacccaccc cttcccagcc tctgagccca
     4441 gaaagcgaag gagcaaagct gctattggcc gctgccccaa aggcctaccc gcttccattg
     4501 ctcagcggtg ctgtccatct gcacgagact agtgagacgt gctacttcca tttgtcacgt
     4561 cctgcacgac gcgagctgcg gggcgggggg gaacttcctg actaggggag gagtagaagg
     4621 tggcgcgaag gggccaccaa agaacggagc cggttggcgc ctaccggtgg atgtggaatg
     4681 tgtgaggcca gaggccactt gtgtagcgcc aagtgcccag cggggctgct aaagcgcatg
     4741 ctccagactg ccttgggaaa agcgcctccc ctacccggta gaattcactc ctcaggtgca
     4801 ggctgcctat cagaaggtgg tggctggtgt ggccaatgcc ctggctcaca aataccactg
     4861 agatcttttt ccctctgcca aaaattatgg ggacatcatg aagccccttg agcatctgac
     4921 ttctggctaa taaaggaaat ttattttcat tgcaatagtg tgttggaatt ttttgtgtct
     4981 ctcactcgga aggacatatg ggagggcaaa tcatttaaaa catcagaatg agtatttggt
     5041 ttagagtttg gcaacatatg ccatatgctg gctgccatga acaaaggtgg ctataaagag
     5101 gtcatcagta tatgaaacag ccccctgctg tccattcctt attccataga aaagccttga
     5161 cttgaggtta gatttttttt atattttgtt ttgtgttatt tttttcttta acatccctaa
     5221 aattttcctt acatgtttta ctagccagat ttttcctcct ctcctgacta ctcccagtca
     5281 tagctgtccc tcttctctta tgaagatccc tcgacctgca gcccaagctt tagggatcaa
     5341 gatcgccaaa aaagaagaga aaggtgccga agaagcatgc agcaccacca aaaaaaaaac
     5401 gaaaagtaga agacccacga ggcaacacca gcggcgtgct gagcaccccc aaggccaaga
     5461 gggccaagca cccccccggc accgagaagc ccaggagcag gagccagagc gagcagcccg
     5521 ccacctgccc catctgctac gccgtgatca ggcagagcag gaacctgagg aggcacctgg
     5581 agctgaggca cttcgccaag cccggcgtgg atccaccggt cgccaccatg gtgagcaagg
     5641 gcgaggagct gttcaccggg gtggtgccca tcctggtcga gctggacggc gacgtaaacg
     5701 gccacaagtt cagcgtgtcc ggcgagggcg agggcgatgc cacctacggc aagctgaccc
     5761 tgaagttcat ctgcaccacc ggcaagctgc ccgtgccctg gcccaccctc gtgaccaccc
     5821 tgacctacgg cgtgcagtgc ttcagccgct accccgacca catgaagcag cacgacttct
     5881 tcaagtccgc catgcccgaa ggctacgtcc aggagcgcac catcttcttc aaggacgacg
     5941 gcaactacaa gacccgcgcc gaggtgaagt tcgagggcga caccctggtg aaccgcatcg
     6001 agctgaaggg catcgacttc aaggaggacg gcaacatcct ggggcacaag ctggagtaca
     6061 actacaacag ccacaacgtc tatatcatgg ccgacaagca gaagaacggc atcaaggtga
     6121 acttcaataa aagcttggcg taatcatggt catagctgtt tcctgtgtga aattgttatc
     6181 cgctcacaat tccacacaac atacgagccg gaagcataaa gtgtaaagcc tggggtgcct
     6241 aatgagtgag ctaactcaca ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa
     6301 acctgtcgtg ccagcggatc cgcatctcaa ttagtcagca accatagtcc cgcccctaac
     6361 tccgcccatc ccgcccctaa ctccgcccag ttccgcccat tctccgcccc atggctgact
     6421 aatttttttt atttatgcag aggccgaggc cgcctcggcc tctgagctat tccagaagta
     6481 gtgaggaggc ttttttggag gcctaggctt ttgcaaaaag ctaacttgtt tattgcagct
     6541 tataatggtt acaaataaag caatagcatc acaaatttca caaataaagc atttttttca
     6601 ctgcattcta gttgtggttt gtccaaactc atcaatgtat cttatcatgt ctggatccgc
     6661 tgcattaatg aatcggccaa cgcgcgggga gaggcggttt gcgtattggg cgctcttccg
     6721 cttcctcgct cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc
     6781 actcaaaggc ggtaatacgg ttatccacag aatcagggga taacgcagga aagaacatgt
     6841 gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc
     6901 ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa
     6961 acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc gtgcgctctc
     7021 ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg ggaagcgtgg
     7081 cgctttctca aagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc
     7141 tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc ggtaactatc
     7201 gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc actggtaaca
     7261 ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg tggcctaact
     7321 acggctacac tagaagaaca gtatttggta tctgcgctct gctgaagcca gttaccttcg
     7381 gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc ggtggttttt
     7441 ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat cctttgatct
     7501 tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt ttggtcatga
     7561 gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt tttaaatcaa
     7621 tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc agtgaggcac
     7681 ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga
     7741 taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata ccgcgagacc
     7801 cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg gccgagcgca
     7861 gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc cgggaagcta
     7921 gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct acaggcatcg
     7981 tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa cgatcaaggc
     8041 gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg
     8101 ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca ctgcataatt
     8161 ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac tcaaccaagt
     8221 cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca atacgggata
     8281 ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt tcttcggggc
     8341 gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc actcgtgcac
     8401 ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca aaaacaggaa
     8461 ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata ctcatactct
     8521 tcctttttca atattattga agcatttatc agggttattg tctcatgagc ggatacatat
     8581 ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cacatttccc cgaaaagtgc
     8641 cacctg
//