LOCUS       pRK5-mEGFP-HMGB1        6010 bp DNA     circular SYN 16-NOV-2022
DEFINITION  .
ACCESSION   .
VERSION     .
KEYWORDS    pRK5-mEGFP-HMGB1-Patchless
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6010)
  TITLE     Spatial spread of Ras activity
REFERENCE   2  (bases 1 to 6010)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Nov 16, 2022 from SnapGene 6.1.0
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..6010
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     source          1659..2288
                     /mol_type="other DNA"
                     /note="color: #ffffff"
                     /organism="synthetic DNA construct"
     enhancer        14..393
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
                     /note="color: #ffffff"
     promoter        394..597
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
                     /note="color: #ffffff; direction: RIGHT"
     promoter        828..846
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
                     /note="color: #ffffff; direction: RIGHT"
     regulatory      933..941
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /note="color: #a6acb3"
                     /regulatory_class="other"
     CDS             939..1648
                     /codon_start=1
                     /product="enhanced GFP with monomerizing A206K mutation 
                     (Zacharias et al., 2002)"
                     /label=mEGFP
                     /note="mammalian codon-optimized"
                     /note="This feature has 3 segments:
                        1:  939 ..  941 / #05fd14
                        2:  942 ..  944 / #05fd14 / 1a
                        3:  945 .. 1648 / #05fd14"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTISFKDD
                     GTYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIK
                     ANFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDE"
     CDS             1659..2288
                     /codon_start=1
                     /label=HMGB1-mut-FL-del-tail
                     /note="color: #993366"
                     /translation="MGKGDPKKPRGKMSSYAFFVQTCREEHKKKHPDASVNFSEFSKKC
                     SERWKTMSAKEKGKFEDMAKADKARYEREMKTYIPPKGETKKKFKDPNAPKRPPSAFFL
                     FCSEYRPKIKGEHPGLSIGDVAKKLGEMWNNTAADDKQPYEKKAAKLKEKYEKDIAAYR
                     AKGKPDAAKKGVVKAEKSKKRKRRKMRKMKRMRRRRKMKKMKMKKK"
     misc_feature    2061..2285
                     /label=IDR
                     /note="color: #a6acb3; direction: RIGHT"
     polyA_signal    2322..2456
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
                     /note="color: #a6acb3"
     promoter        2525..2882
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
                     /note="color: #ffffff; direction: RIGHT"
     rep_origin      2733..2868
                     /label=SV40 ori
                     /note="SV40 origin of replication"
                     /note="color: #ffff00"
     primer_bind     complement(2902..2918)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
                     /note="color: #a020f0; direction: LEFT"
     rep_origin      3131..3586
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
                     /note="color: #ffff00"
     promoter        3868..3972
                     /gene="bla"
                     /label=AmpR promoter
                     /note="color: #ffffff; direction: RIGHT"
     CDS             3973..4833
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /note="This feature has 2 segments:
                        1: 3973 .. 4041 / #ccffcc / signal sequence
                        2: 4042 .. 4833 / #ccffcc
                     Cleavage site after base 4041"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRDDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     rep_origin      5004..5592
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
                     /note="color: #ffff00"
     promoter        5916..5946
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
                     /note="This forward directional feature has 3 segments:
                        1: 5916 .. 5921 / #ffffff / -35
                        2: 5922 .. 5939 / #ffffff
                        3: 5940 .. 5946 / #ffffff / -10"
     protein_bind    5954..5970
                     /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)."
                     /note="color: #31849b"
     primer_bind     5978..5994
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
                     /note="color: #a020f0; direction: RIGHT"
ORIGIN
        1 ttcgagctcg cccgacattg attattgact agttattaat agtaatcaat tacggggtca
       61 ttagttcata gcccatatat ggagttccgc gttacataac ttacggtaaa tggcccgcct
      121 ggctgaccgc ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt tcccatagta
      181 acgccaatag ggactttcca ttgacgtcaa tgggtggagt atttacggta aactgcccac
      241 ttggcagtac atcaagtgta tcatatgcca agtacgcccc ctattgacgt caatgacggt
      301 aaatggcccg cctggcatta tgcccagtac atgaccttat gggactttcc tacttggcag
      361 tacatctacg tattagtcat cgctattacc atggtgatgc ggttttggca gtacatcaat
      421 gggcgtggat agcggtttga ctcacgggga tttccaagtc tccaccccat tgacgtcaat
      481 gggagtttgt tttggcacca aaatcaacgg gactttccaa aatgtcgtaa caactccgcc
      541 ccattgacgc aaatgggcgg taggcgtgta cggtgggagg tctatataag cagagctcgt
      601 ttagtgaacc gtcagatcgc ctggagacgc catccacgct gttttgacct ccatagaaga
      661 caccgggacc gatccagcct ccgcggccgg gaacggtgca ttggaacgcg gattccccgt
      721 gccaagagtg acgtaagtac cgcctataga gtctataggc ccaccccctt ggcttcgtta
      781 gaacgcggct acaattaata cataacctta tgtatcatac acatacgatt taggtgacac
      841 tatagaataa catccacttt gcctttctct ccacaggtgt ccactcccag gtccaactgc
      901 acctcggttc tatcgattga attcgtaccg gtgccaccat ggtgagcaag ggcgaggagc
      961 tgttcaccgg ggtggtgccc atcctggtcg agctggacgg cgacgtaaac ggccacaagt
     1021 tcagcgtgcg cggcgagggc gagggcgatg ccaccaacgg caagctgacc ctgaagttca
     1081 tctgcaccac cggcaagctg cccgtgccct ggcccaccct cgtgaccacc ctgacctacg
     1141 gcgtgcagtg cttcagccgc taccccgacc acatgaagca gcacgacttc ttcaagtccg
     1201 ccatgcccga aggctacgtc caggagcgca ccatctcctt caaggacgac ggcacctaca
     1261 agacccgcgc cgaggtgaag ttcgagggcg acaccctggt gaaccgcatc gagctgaagg
     1321 gcatcgactt caaggaggac ggcaacatcc tggggcacaa gctggagtac aacttcaaca
     1381 gccacaacgt ctatatcacg gccgacaagc agaagaacgg catcaaggcg aacttcaaga
     1441 tccgccacaa cgtcgaggac ggcagcgtgc agctcgccga ccactaccag cagaacaccc
     1501 ccatcggcga cggccccgtg ctgctgcccg acaaccacta cctgagcacc cagtccaagc
     1561 tgagcaaaga ccccaacgag aagcgcgatc acatggtcct gctggagttc gtgaccgccg
     1621 ccgggatcac tctcggcatg gacgagctgt acacggcgat gggcaaagga gatcctaaga
     1681 agccgagagg caaaatgtca tcatatgcat tttttgtgca aacttgtcgg gaggagcata
     1741 agaagaagca cccagatgct tcagtcaact tctcagagtt ttctaagaag tgctcagaga
     1801 ggtggaagac catgtctgct aaagagaaag gaaaatttga agatatggca aaagcggaca
     1861 aggcccgtta tgaaagagaa atgaaaacct atatccctcc caaaggggag acaaaaaaga
     1921 agttcaagga tcccaatgca cccaagaggc ctccttcggc cttcttcctc ttctgctctg
     1981 agtatcgccc aaaaatcaaa ggagaacatc ctggcctgtc cattggtgat gttgcgaaga
     2041 aactgggaga gatgtggaat aacactgctg cagatgacaa gcagccttat gaaaagaagg
     2101 ctgcgaagct gaaggaaaaa tatgaaaagg atattgctgc atatcgagct aaaggaaagc
     2161 ctgatgcagc aaaaaaggga gttgtcaagg ctgaaaaaag caagaaaagg aagaggagga
     2221 agatgaggaa gatgaagagg atgaggagga ggaggaagat gaagaagatg aagatgaaga
     2281 agaagtaaag tcgacctgca gaagcttggc cgccatggcc caacttgttt attgcagctt
     2341 ataatggtta caaataaagc aatagcatca caaatttcac aaataaagca tttttttcac
     2401 tgcattctag ttgtggtttg tccaaactca tcaatgtatc ttatcatgtc tggatcggga
     2461 attaattcgg cgcagcacca tggcctgaaa taacctctga aagaggaact tggttaggta
     2521 ccttctgagg cggaaagaac cagctgtgga atgtgtgtca gttagggtgt ggaaagtccc
     2581 caggctcccc agcaggcaga agtatgcaaa gcatgcatct caattagtca gcaaccaggt
     2641 gtggaaagtc cccaggctcc ccagcaggca gaagtatgca aagcatgcat ctcaattagt
     2701 cagcaaccat agtcccgccc ctaactccgc ccatcccgcc cctaactccg cccagttccg
     2761 cccattctcc gccccatggc tgactaattt tttttattta tgcagaggcc gaggccgcct
     2821 cggcctctga gctattccag aagtagtgag gaggcttttt tggaggccta ggcttttgca
     2881 aaaagctgtt aacagcttgg cactggccgt cgttttacaa cgtcgtgact gggaaaaccc
     2941 tggcgttacc caacttaatc gccttgcagc acatcccccc ttcgccagct ggcgtaatag
     3001 cgaagaggcc cgcaccgatc gcccttccca acagttgcgt agcctgaatg gcgaatggcg
     3061 cctgatgcgg tattttctcc ttacgcatct gtgcggtatt tcacaccgca tacgtcaaag
     3121 caaccatagt acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc
     3181 agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc ctttcgcttt cttcccttcc
     3241 tttctcgcca cgttcgccgg ctttccccgt caagctctaa atcgggggct ccctttaggg
     3301 ttccgattta gtgctttacg gcacctcgac cccaaaaaac ttgatttggg tgatggttca
     3361 cgtagtgggc catcgccctg atagacggtt tttcgccctt tgacgttgga gtccacgttc
     3421 tttaatagtg gactcttgtt ccaaactgga acaacactca accctatctc gggctattct
     3481 tttgatttat aagggatttt gccgatttcg gcctattggt taaaaaatga gctgatttaa
     3541 caaaaattta acgcgaattt taacaaaata ttaacgttta caattttatg gtgcactctc
     3601 agtacaatct gctctgatgc cgcatagtta agccagcccc gacacccgcc aacacccgct
     3661 gacgcgccct gacgggcttg tctgctcccg gcatccgctt acagacaagc tgtgaccgtc
     3721 tccgggagct gcatgtgtca gaggttttca ccgtcatcac cgaaacgcgc gagacgaaag
     3781 ggcctcgtga tacgcctatt tttataggtt aatgtcatga taataatggt ttcttagacg
     3841 tcaggtggca cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt tttctaaata
     3901 cattcaaata tgtatccgct catgagacaa taaccctgat aaatgcttca ataatattga
     3961 aaaaggaaga gtatgagtat tcaacatttc cgtgtcgccc ttattccctt ttttgcggca
     4021 ttttgccttc ctgtttttgc tcacccagaa acgctggtga aagtaaaaga tgctgaagat
     4081 cagttgggtg cacgagtggg ttacatcgaa ctggatctca acagcggtaa gatccttgag
     4141 agttttcgcc ccgaagaacg ttttccaatg atgagcactt ttaaagttct gctatgtggc
     4201 gcggtattat cccgtgatga cgccgggcaa gagcaactcg gtcgccgcat acactattct
     4261 cagaatgact tggttgagta ctcaccagtc acagaaaagc atcttacgga tggcatgaca
     4321 gtaagagaat tatgcagtgc tgccataacc atgagtgata acactgcggc caacttactt
     4381 ctgacaacga tcggaggacc gaaggagcta accgcttttt tgcacaacat gggggatcat
     4441 gtaactcgcc ttgatcgttg ggaaccggag ctgaatgaag ccataccaaa cgacgagcgt
     4501 gacaccacga tgccagcagc aatggcaaca acgttgcgca aactattaac tggcgaacta
     4561 cttactctag cttcccggca acaattaata gactggatgg aggcggataa agttgcagga
     4621 ccacttctgc gctcggccct tccggctggc tggtttattg ctgataaatc tggagccggt
     4681 gagcgtgggt ctcgcggtat cattgcagca ctggggccag atggtaagcc ctcccgtatc
     4741 gtagttatct acacgacggg gagtcaggca actatggatg aacgaaatag acagatcgct
     4801 gagataggtg cctcactgat taagcattgg taactgtcag accaagttta ctcatatata
     4861 ctttagattg atttaaaact tcatttttaa tttaaaagga tctaggtgaa gatccttttt
     4921 gataatctca tgaccaaaat cccttaacgt gagttttcgt tccactgagc gtcagacccc
     4981 gtagaaaaga tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg
     5041 caaacaaaaa aaccaccgct accagcggtg gtttgtttgc cggatcaaga gctaccaact
     5101 ctttttccga aggtaactgg cttcagcaga gcgcagatac caaatactgt ccttctagtg
     5161 tagccgtagt taggccacca cttcaagaac tctgtagcac cgcctacata cctcgctctg
     5221 ctaatcctgt taccagtggc tgctgccagt ggcgataagt cgtgtcttac cgggttggac
     5281 tcaagacgat agttaccgga taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca
     5341 cagcccagct tggagcgaac gacctacacc gaactgagat acctacagcg tgagcattga
     5401 gaaagcgcca cgcttcccga agggagaaag gcggacaggt atccggtaag cggcagggtc
     5461 ggaacaggag agcgcacgag ggagcttcca gggggaaacg cctggtatct ttatagtcct
     5521 gtcgggtttc gccacctctg acttgagcgt cgatttttgt gatgctcgtc aggggggcgg
     5581 agcctatgga aaaacgccag caacgcggcc tttttacggt tcctggcctt ttgctggcct
     5641 tttgctcaca tgttctttcc tgcgttatcc cctgattctg tggataaccg tattaccgcc
     5701 tttgagtgag ctgataccgc tcgccgcagc cgaacgaccg agcgcagcga gtcagtgagc
     5761 gaggaagcgg aagagcgccc aatacgcaaa ccgcctctcc ccgcgcgttg gccgattcat
     5821 taatccagct ggcacgacag gtttcccgac tggaaagcgg gcagtgagcg caacgcaatt
     5881 aatgtgagtt acctcactca ttaggcaccc caggctttac actttatgct tccggctcgt
     5941 atgttgtgtg gaattgtgag cggataacaa tttcacacag gaaacagcta tgaccatgat
     6001 tacgaattaa
//
