LOCUS       sequence-204244-        6719 bp    DNA     circular SYN 15-APR-2025
DEFINITION  To study altered downstream functions of phospholylation-site 
            mimetic kindlin2.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6719)
  AUTHORS   Artym VV, Swatkoski S, Matsumoto K, Campbell CB, Petrie RJ, 
            Dimitriadis EK, Li X, Mueller SC, Bugge TH, Gucek M, Yamada KM
  TITLE     Dense fibrillar collagen is a potent inducer of invadopodia via a 
            specific signaling network.
  JOURNAL   J Cell Biol. 2015 Feb 2;208(3):331-50. doi: 10.1083/jcb.201405099.
  PUBMED    25646088
REFERENCE   2  (bases 1 to 6719)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; doi: "10.1083/jcb"; 
            journalName: "J Cell Biol"; date: "2015-02-2- 2"; volume: "208"; 
            issue: "3"; pages: "331-50"
COMMENT     Sequence Label: pEGFP Kind2 S181E
FEATURES             Location/Qualifiers
     source          1..6719
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      11..599
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     500..519
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     enhancer        774..1077
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        1078..1281
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     1232..1252
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      1311..1320
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(3378..3380,3381..3383,3384..4097)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(3378..3380,3381..3383,3384..4094)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(3423..3444)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(3684..3703)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     4031..4052
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    4220..4341
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4257..4276)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4311..4330
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(4348..4803)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4485..4506)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4697..4716
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        4830..4934
                     /gene="bla"
                     /label=AmpR promoter
     promoter        4936..5293
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        4936..5127
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(4959..4979)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      5144..5279
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             5328..6122
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             5328..6122
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(5382..5401)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5992..6011
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(6310..6329)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    6354..6401
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
ORIGIN
        1 aaggatcttc ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac
       61 caccgctacc agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg
      121 taactggctt cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag
      181 gccaccactt caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac
      241 cagtggctgc tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt
      301 taccggataa ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg
      361 agcgaacgac ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc
      421 ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc
      481 gcacgaggga gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc
      541 acctctgact tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa
      601 acgccagcaa cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt
      661 tctttcctgc gttatcccct gattctgtgg ataaccgtat taccgccatg cattagttat
      721 taatagtaat caattacggg gtcattagtt catagcccat atatggagtt ccgcgttaca
      781 taacttacgg taaatggccc gcctggctga ccgcccaacg acccccgccc attgacgtca
      841 ataatgacgt atgttcccat agtaacgcca atagggactt tccattgacg tcaatgggtg
      901 gagtatttac ggtaaactgc ccacttggca gtacatcaag tgtatcatat gccaagtacg
      961 ccccctattg acgtcaatga cggtaaatgg cccgcctggc attatgccca gtacatgacc
     1021 ttatgggact ttcctacttg gcagtacatc tacgtattag tcatcgctat taccatggtg
     1081 atgcggtttt ggcagtacat caatgggcgt ggatagcggt ttgactcacg gggatttcca
     1141 agtctccacc ccattgacgt caatgggagt ttgttttggc accaaaatca acgggacttt
     1201 ccaaaatgtc gtaacaactc cgccccattg acgcaaatgg gcggtaggcg tgtacggtgg
     1261 gaggtctata taagcagagc tggtttagtg aaccgtcaga tccgctagcg gccaccatgg
     1321 ctctggacgg gataaggatg ccagatggct gctacgcgga cgggacgtgg gaactgagtg
     1381 tccatgtgac ggacctgaac cgcgatgtca ccctgagagt gaccggcgag gtgcacattg
     1441 gaggcgtgat gcttaagctg gtggagaaac tcgatgtaaa aaaagattgg tctgaccatg
     1501 ctctctggtg ggaaaagaag agaacttggc ttctgaagac acattggacc ttagataagt
     1561 atggtattca ggcagatgct aagcttcagt tcacccctca gcacaaactg ctccgcctgc
     1621 agcttcccaa catgaagtat gtgaaggtga aagtgaattt ctctgataga gtcttcaaag
     1681 ctgtttctga catctgtaag acttttaata tcagacaccc cgaagaactt tctctcttaa
     1741 agaaacccag agatccaaca aagaaaaaaa agaagaagct agatgaccag tctgaagatg
     1801 aggcacttga attagagggg cctcttatca ctcctggatc aggaagtata tattcagagc
     1861 caggactgta tagtaaaaca atgaccccca cttatgatgc tcatgatgga agccccttgt
     1921 caccaacttc tgcttggttt ggtgacagtg ctttgtcaga aggcaatcct ggtatacttg
     1981 ctgtcagtca accaatcacg tcaccagaaa tcttggcaaa aatgttcaag cctcaagctc
     2041 ttcttgataa agcaaaaatc aaccaaggat ggcttgattc ctcaagatct ctcatggaac
     2101 aagatgtgaa ggaaaatgag gccttgctgc tccgattcaa gtattacagc ttttttgatt
     2161 tgaatccaaa gtatgatgca atcagaatca atcagcttta tgagcaggcc aaatgggcca
     2221 ttctcctgga agagattgaa tgcacagaag aagaaatgat gatgtttgca gccctgcagt
     2281 atcatatcaa taagctgtca atcatgacat cagagaatca tttgaacaac agtgacaaag
     2341 aagttgatga agttgatgct gccctttcag acctggagat tactctggaa gggggtaaaa
     2401 cgtcaacaat tttgggtgac attacttcca ttcctgaact tgctgactac attaaagttt
     2461 tcaagccaaa aaagctgact ctgaaaggtt acaaacaata ttggtgcacc ttcaaagaca
     2521 catccatttc ttgttataag agcaaagaag aatccagtgg cacaccagct catcagatga
     2581 acctcagggg atgtgaagtt accccagatg taaacatttc aggccaaaaa tttaacatta
     2641 aactcctgat tccagttgca gaaggcatga atgaaatctg gcttcgttgt gacaatgaaa
     2701 aacagtatgc acactggatg gcagcctgca gattagcctc caaaggcaag accatggcgg
     2761 acagttctta caacttagaa gttcagaata ttctttcctt tctgaagatg cagcatttaa
     2821 acccagatcc tcagttaata ccagagcaga tcacgactga tataactcct gaatgtttgg
     2881 tgtctccccg ctatctaaaa aagtataaga acaagcagat aacagcgaga atcttggagg
     2941 cccatcagaa tgtagctcag atgagtctaa ttgaagccaa gatgagattt attcaagctt
     3001 ggcagtcact acctgaattt ggcatcactc acttcattgc aaggttccaa gggggcaaaa
     3061 aagaagaact tattggaatt gcatacaaca gactgattcg gatggatgcc agcactggag
     3121 atgcaattaa aacatggcgt ttcagcaaca tgaaacagtg gaatgtcaac tgggaaatca
     3181 aaatggtcac cgtagagttt gcagatgaag tacgattgtc cttcatttgt actgaagtag
     3241 attgcaaagt ggttcatgaa ttcattggtg gctacatatt tctctcaaca cgtgcaaaag
     3301 accaaaacga gagtttagat gaagagatgt tctacaaact taccagtggt tgggtgggta
     3361 actcagggga tccaggaatg gtgagcaagg gcgaggagct gttcaccggg gtggtgccca
     3421 tcctggtcga gctggacggc gacgtaaacg gccacaagtt cagcgtgtcc ggcgagggcg
     3481 agggcgatgc cacctacggc aagctgaccc tgaagttcat ctgcaccacc ggcaagctgc
     3541 ccgtgccctg gcccaccctc gtgaccaccc tgacctacgg cgtgcagtgc ttcagccgct
     3601 accccgacca catgaagcag cacgacttct tcaagtccgc catgcccgaa ggctacgtcc
     3661 aggagcgcac catcttcttc aaggacgacg gcaactacaa gacccgcgcc gaggtgaagt
     3721 tcgagggcga caccctggtg aaccgcatcg agctgaaggg catcgacttc aaggaggacg
     3781 gcaacatcct ggggcacaag ctggagtaca actacaacag ccacaacgtc tatatcatgg
     3841 ccgacaagca gaagaacggc atcaaggtga acttcaagat ccgccacaac atcgaggacg
     3901 gcagcgtgca gctcgccgac cactaccagc agaacacccc catcggcgac ggccccgtgc
     3961 tgctgcccga caaccactac ctgagcaccc agtccgccct gagcaaagac cccaacgaga
     4021 agcgcgatca catggtcctg ctggagttcg tgaccgccgc cgggatcact ctcggcatgg
     4081 acgagctgta caagtaaagc ggccgcgact ctagatcata atcagccata ccacatttgt
     4141 agaggtttta cttgctttaa aaaacctccc acacctcccc ctgaacctga aacataaaat
     4201 gaatgcaatt gttgttgtta acttgtttat tgcagcttat aatggttaca aataaagcaa
     4261 tagcatcaca aatttcacaa ataaagcatt tttttcactg cattctagtt gtggtttgtc
     4321 caaactcatc aatgtatctt aaggcgtaaa ttgtaagcgt taatattttg ttaaaattcg
     4381 cgttaaattt ttgttaaatc agctcatttt ttaaccaata ggccgaaatc ggcaaaatcc
     4441 cttataaatc aaaagaatag accgagatag ggttgagtgt tgttccagtt tggaacaaga
     4501 gtccactatt aaagaacgtg gactccaacg tcaaagggcg aaaaaccgtc tatcagggcg
     4561 atggcccact acgtgaacca tcaccctaat caagtttttt ggggtcgagg tgccgtaaag
     4621 cactaaatcg gaaccctaaa gggagccccc gatttagagc ttgacgggga aagccggcga
     4681 acgtggcgag aaaggaaggg aagaaagcga aaggagcggg cgctagggcg ctggcaagtg
     4741 tagcggtcac gctgcgcgta accaccacac ccgccgcgct taatgcgccg ctacagggcg
     4801 cgtcaggtgg cacttttcgg ggaaatgtgc gcggaacccc tatttgttta tttttctaaa
     4861 tacattcaaa tatgtatccg ctcatgagac aataaccctg ataaatgctt caataatatt
     4921 gaaaaaggaa gagtcctgag gcggaaagaa ccagctgtgg aatgtgtgtc agttagggtg
     4981 tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa agcatgcatc tcaattagtc
     5041 agcaaccagg tgtggaaagt ccccaggctc cccagcaggc agaagtatgc aaagcatgca
     5101 tctcaattag tcagcaacca tagtcccgcc cctaactccg cccatcccgc ccctaactcc
     5161 gcccagttcc gcccattctc cgccccatgg ctgactaatt ttttttattt atgcagaggc
     5221 cgaggccgcc tcggcctctg agctattcca gaagtagtga ggaggctttt ttggaggcct
     5281 aggcttttgc aaagatcgat caagagacag gatgaggatc gtttcgcatg attgaacaag
     5341 atggattgca cgcaggttct ccggccgctt gggtggagag gctattcggc tatgactggg
     5401 cacaacagac aatcggctgc tctgatgccg ccgtgttccg gctgtcagcg caggggcgcc
     5461 cggttctttt tgtcaagacc gacctgtccg gtgccctgaa tgaactgcaa gacgaggcag
     5521 cgcggctatc gtggctggcc acgacgggcg ttccttgcgc agctgtgctc gacgttgtca
     5581 ctgaagcggg aagggactgg ctgctattgg gcgaagtgcc ggggcaggat ctcctgtcat
     5641 ctcaccttgc tcctgccgag aaagtatcca tcatggctga tgcaatgcgg cggctgcata
     5701 cgcttgatcc ggctacctgc ccattcgacc accaagcgaa acatcgcatc gagcgagcac
     5761 gtactcggat ggaagccggt cttgtcgatc aggatgatct ggacgaagag catcaggggc
     5821 tcgcgccagc cgaactgttc gccaggctca aggcgagcat gcccgacggc gaggatctcg
     5881 tcgtgaccca tggcgatgcc tgcttgccga atatcatggt ggaaaatggc cgcttttctg
     5941 gattcatcga ctgtggccgg ctgggtgtgg cggaccgcta tcaggacata gcgttggcta
     6001 cccgtgatat tgctgaagag cttggcggcg aatgggctga ccgcttcctc gtgctttacg
     6061 gtatcgccgc tcccgattcg cagcgcatcg ccttctatcg ccttcttgac gagttcttct
     6121 gagcgggact ctggggttcg aaatgaccga ccaagcgacg cccaacctgc catcacgaga
     6181 tttcgattcc accgccgcct tctatgaaag gttgggcttc ggaatcgttt tccgggacgc
     6241 cggctggatg atcctccagc gcggggatct catgctggag ttcttcgccc accctagggg
     6301 gaggctaact gaaacacgga aggagacaat accggaagga acccgcgcta tgacggcaat
     6361 aaaaagacag aataaaacgc acggtgttgg gtcgtttgtt cataaacgcg gggttcggtc
     6421 ccagggctgg cactctgtcg ataccccacc gagaccccat tggggccaat acgcccgcgt
     6481 ttcttccttt tccccacccc accccccaag ttcgggtgaa ggcccagggc tcgcagccaa
     6541 cgtcggggcg gcaggccctg ccatagcctc aggttactca tatatacttt agattgattt
     6601 aaaacttcat ttttaattta aaaggatcta ggtgaagatc ctttttgata atctcatgac
     6661 caaaatccct taacgtgagt tttcgttcca ctgagcgtca gaccccgtag aaaagatca
//
