LOCUS       pRK5-mEGFP-SQSTM        6703 bp DNA     circular SYN 16-NOV-2022
DEFINITION  .
ACCESSION   .
VERSION     .
KEYWORDS    pRK5-mEGFP-SQSTM1-WT
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6703)
  TITLE     Spatial spread of Ras activity
REFERENCE   2  (bases 1 to 6703)
  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..6703
                     /mol_type="other DNA"
                     /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..2981
                     /codon_start=1
                     /label=SQSTM1_WT
                     /note="color: #993366"
                     /translation="MASLTVKAYLLGKEDAAREIRRFSFCCSPEPEAEAEAAAGPGPCE
                     RLLSRVAALFPALRPGGFQAHYRDEDGDLVAFSSDEELTMAMSYVKDDIFRIYIKEKKE
                     CRRDHRPPCAQEAPRNMVHPNVICDGCNGPVVGTRYKCSVCPDYDLCSVCEGKGLHRGH
                     TKLAFPSPFGHLSEGFSHSRWLRKVKHGHFGWPGWEMGPPGNWSPRPPRAGEARPGPTA
                     ESASGPSEDPSVNFLKNVGESVAAALSPLGIEVDIDVEHGGKRSRLTPVSPESSSTEEK
                     SSSQPSSCCSDPSKPGGNVEGATQSLAEQMRKIALESEGRPEEQMESDNCSGGDDDWTH
                     LSSKEVDPSTGELQSLQMPESEGPSSLDPSQEGPTGLKEAALYPHLPPEADPRLIESLS
                     QMLSMGFSDEGGWLTRLLQTKNYDIGAALDTIQYSKHPPPL"
     polyA_signal    3015..3149
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
                     /note="color: #a6acb3"
     promoter        3218..3575
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
                     /note="color: #ffffff; direction: RIGHT"
     rep_origin      3426..3561
                     /label=SV40 ori
                     /note="SV40 origin of replication"
                     /note="color: #ffff00"
     primer_bind     complement(3595..3611)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
                     /note="color: #a020f0; direction: LEFT"
     rep_origin      3824..4279
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
                     /note="color: #ffff00"
     promoter        4561..4665
                     /gene="bla"
                     /label=AmpR promoter
                     /note="color: #ffffff; direction: RIGHT"
     CDS             4666..5526
                     /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: 4666 .. 4734 / #ccffcc / signal sequence
                        2: 4735 .. 5526 / #ccffcc
                     Cleavage site after base 4734"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRDDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     rep_origin      5697..6285
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
                     /note="color: #ffff00"
     promoter        6609..6639
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
                     /note="This forward directional feature has 3 segments:
                        1: 6609 .. 6614 / #ffffff / -35
                        2: 6615 .. 6632 / #ffffff
                        3: 6633 .. 6639 / #ffffff / -10"
     protein_bind    6647..6663
                     /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     6671..6687
                     /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 ggcgtcgctc accgtgaagg
     1681 cctaccttct gggcaaggag gacgcggcgc gcgagattcg ccgcttcagc ttctgctgca
     1741 gccccgagcc tgaggcggaa gccgaggctg cggcgggtcc gggaccctgc gagcggctgc
     1801 tgagccgggt ggccgccctg ttccccgcgc tgcggcctgg cggcttccag gcgcactacc
     1861 gcgatgagga cggggacttg gttgcctttt ccagtgacga ggaattgaca atggccatgt
     1921 cctacgtgaa ggatgacatc ttccgaatct acattaaaga gaaaaaagag tgccggcggg
     1981 accaccgccc accgtgtgct caggaggcgc cccgcaacat ggtgcacccc aatgtgatct
     2041 gcgatggctg caatgggcct gtggtaggaa cccgctacaa gtgcagcgtc tgcccagact
     2101 acgacttgtg tagcgtctgc gagggaaagg gcttgcaccg ggggcacacc aagctcgcat
     2161 tccccagccc cttcgggcac ctgtctgagg gcttctcgca cagccgctgg ctccggaagg
     2221 tgaaacacgg acacttcggg tggccaggat gggaaatggg tccaccagga aactggagcc
     2281 cacgtcctcc tcgtgcaggg gaggcccgcc ctggccccac ggcagaatca gcttctggtc
     2341 catcggagga tccgagtgtg aatttcctga agaacgttgg ggagagtgtg gcagctgccc
     2401 ttagccctct gggcattgaa gttgatatcg atgtggagca cggagggaaa agaagccgcc
     2461 tgacccccgt ctctccagag agttccagca cagaggagaa gagcagctca cagccaagca
     2521 gctgctgctc tgaccccagc aagccgggtg ggaatgttga gggcgccacg cagtctctgg
     2581 cggagcagat gaggaagatc gccttggagt ccgaggggcg ccctgaggaa cagatggagt
     2641 cggataactg ttcaggagga gatgatgact ggacccatct gtcttcaaaa gaagtggacc
     2701 cgtctacagg tgaactccag tccctacaga tgccagaatc cgaagggcca agctctctgg
     2761 acccctccca ggagggaccc acagggctga aggaagctgc cttgtaccca catctcccgc
     2821 cagaggctga cccgcggctg attgagtccc tctcccagat gctgtccatg ggcttctctg
     2881 atgaaggcgg ctggctcacc aggctcctgc agaccaagaa ctatgacatc ggagcggctc
     2941 tggacaccat ccagtattca aagcatcccc cgccgttgtg aagtcgacct gcagaagctt
     3001 ggccgccatg gcccaacttg tttattgcag cttataatgg ttacaaataa agcaatagca
     3061 tcacaaattt cacaaataaa gcattttttt cactgcattc tagttgtggt ttgtccaaac
     3121 tcatcaatgt atcttatcat gtctggatcg ggaattaatt cggcgcagca ccatggcctg
     3181 aaataacctc tgaaagagga acttggttag gtaccttctg aggcggaaag aaccagctgt
     3241 ggaatgtgtg tcagttaggg tgtggaaagt ccccaggctc cccagcaggc agaagtatgc
     3301 aaagcatgca tctcaattag tcagcaacca ggtgtggaaa gtccccaggc tccccagcag
     3361 gcagaagtat gcaaagcatg catctcaatt agtcagcaac catagtcccg cccctaactc
     3421 cgcccatccc gcccctaact ccgcccagtt ccgcccattc tccgccccat ggctgactaa
     3481 ttttttttat ttatgcagag gccgaggccg cctcggcctc tgagctattc cagaagtagt
     3541 gaggaggctt ttttggaggc ctaggctttt gcaaaaagct gttaacagct tggcactggc
     3601 cgtcgtttta caacgtcgtg actgggaaaa ccctggcgtt acccaactta atcgccttgc
     3661 agcacatccc cccttcgcca gctggcgtaa tagcgaagag gcccgcaccg atcgcccttc
     3721 ccaacagttg cgtagcctga atggcgaatg gcgcctgatg cggtattttc tccttacgca
     3781 tctgtgcggt atttcacacc gcatacgtca aagcaaccat agtacgcgcc ctgtagcggc
     3841 gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga ccgctacact tgccagcgcc
     3901 ctagcgcccg ctcctttcgc tttcttccct tcctttctcg ccacgttcgc cggctttccc
     3961 cgtcaagctc taaatcgggg gctcccttta gggttccgat ttagtgcttt acggcacctc
     4021 gaccccaaaa aacttgattt gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg
     4081 gtttttcgcc ctttgacgtt ggagtccacg ttctttaata gtggactctt gttccaaact
     4141 ggaacaacac tcaaccctat ctcgggctat tcttttgatt tataagggat tttgccgatt
     4201 tcggcctatt ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa ttttaacaaa
     4261 atattaacgt ttacaatttt atggtgcact ctcagtacaa tctgctctga tgccgcatag
     4321 ttaagccagc cccgacaccc gccaacaccc gctgacgcgc cctgacgggc ttgtctgctc
     4381 ccggcatccg cttacagaca agctgtgacc gtctccggga gctgcatgtg tcagaggttt
     4441 tcaccgtcat caccgaaacg cgcgagacga aagggcctcg tgatacgcct atttttatag
     4501 gttaatgtca tgataataat ggtttcttag acgtcaggtg gcacttttcg gggaaatgtg
     4561 cgcggaaccc ctatttgttt atttttctaa atacattcaa atatgtatcc gctcatgaga
     4621 caataaccct gataaatgct tcaataatat tgaaaaagga agagtatgag tattcaacat
     4681 ttccgtgtcg cccttattcc cttttttgcg gcattttgcc ttcctgtttt tgctcaccca
     4741 gaaacgctgg tgaaagtaaa agatgctgaa gatcagttgg gtgcacgagt gggttacatc
     4801 gaactggatc tcaacagcgg taagatcctt gagagttttc gccccgaaga acgttttcca
     4861 atgatgagca cttttaaagt tctgctatgt ggcgcggtat tatcccgtga tgacgccggg
     4921 caagagcaac tcggtcgccg catacactat tctcagaatg acttggttga gtactcacca
     4981 gtcacagaaa agcatcttac ggatggcatg acagtaagag aattatgcag tgctgccata
     5041 accatgagtg ataacactgc ggccaactta cttctgacaa cgatcggagg accgaaggag
     5101 ctaaccgctt ttttgcacaa catgggggat catgtaactc gccttgatcg ttgggaaccg
     5161 gagctgaatg aagccatacc aaacgacgag cgtgacacca cgatgccagc agcaatggca
     5221 acaacgttgc gcaaactatt aactggcgaa ctacttactc tagcttcccg gcaacaatta
     5281 atagactgga tggaggcgga taaagttgca ggaccacttc tgcgctcggc ccttccggct
     5341 ggctggttta ttgctgataa atctggagcc ggtgagcgtg ggtctcgcgg tatcattgca
     5401 gcactggggc cagatggtaa gccctcccgt atcgtagtta tctacacgac ggggagtcag
     5461 gcaactatgg atgaacgaaa tagacagatc gctgagatag gtgcctcact gattaagcat
     5521 tggtaactgt cagaccaagt ttactcatat atactttaga ttgatttaaa acttcatttt
     5581 taatttaaaa ggatctaggt gaagatcctt tttgataatc tcatgaccaa aatcccttaa
     5641 cgtgagtttt cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttcttga
     5701 gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg
     5761 gtggtttgtt tgccggatca agagctacca actctttttc cgaaggtaac tggcttcagc
     5821 agagcgcaga taccaaatac tgtccttcta gtgtagccgt agttaggcca ccacttcaag
     5881 aactctgtag caccgcctac atacctcgct ctgctaatcc tgttaccagt ggctgctgcc
     5941 agtggcgata agtcgtgtct taccgggttg gactcaagac gatagttacc ggataaggcg
     6001 cagcggtcgg gctgaacggg gggttcgtgc acacagccca gcttggagcg aacgacctac
     6061 accgaactga gatacctaca gcgtgagcat tgagaaagcg ccacgcttcc cgaagggaga
     6121 aaggcggaca ggtatccggt aagcggcagg gtcggaacag gagagcgcac gagggagctt
     6181 ccagggggaa acgcctggta tctttatagt cctgtcgggt ttcgccacct ctgacttgag
     6241 cgtcgatttt tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg
     6301 gcctttttac ggttcctggc cttttgctgg ccttttgctc acatgttctt tcctgcgtta
     6361 tcccctgatt ctgtggataa ccgtattacc gcctttgagt gagctgatac cgctcgccgc
     6421 agccgaacga ccgagcgcag cgagtcagtg agcgaggaag cggaagagcg cccaatacgc
     6481 aaaccgcctc tccccgcgcg ttggccgatt cattaatcca gctggcacga caggtttccc
     6541 gactggaaag cgggcagtga gcgcaacgca attaatgtga gttacctcac tcattaggca
     6601 ccccaggctt tacactttat gcttccggct cgtatgttgt gtggaattgt gagcggataa
     6661 caatttcaca caggaaacag ctatgaccat gattacgaat taa
//
