LOCUS       sequence-204232-        6719 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  To study altered downstream functions of phospholylation-site null 
            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 Dec 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 S181A
FEATURES             Location/Qualifiers
     source          1..6719
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      29..617
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     518..537
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     enhancer        792..1095
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        1096..1299
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     1250..1270
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      1329..1338
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(3396..3398,3399..3401,3402..4115)
                     /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(3396..3398,3399..3401,3402..4112)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(3441..3462)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(3702..3721)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     4049..4070
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    4238..4359
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4275..4294)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4329..4348
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(4366..4821)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4503..4524)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4715..4734
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        4848..4952
                     /gene="bla"
                     /label=AmpR promoter
     promoter        4954..5311
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        4954..5145
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(4977..4997)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      5162..5297
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             5346..6140
                     /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             5346..6140
                     /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(5400..5419)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     6010..6029
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(6328..6347)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    6372..6419
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
ORIGIN
        1 accccgtaga aaagatcaaa ggatcttctt gagatccttt ttttctgcgc gtaatctgct
       61 gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg tttgccggat caagagctac
      121 caactctttt tccgaaggta actggcttca gcagagcgca gataccaaat actgttcttc
      181 tagtgtagcc gtagttaggc caccacttca agaactctgt agcaccgcct acatacctcg
      241 ctctgctaat cctgttacca gtggctgctg ccagtggcga taagtcgtgt cttaccgggt
      301 tggactcaag acgatagtta ccggataagg cgcagcggtc gggctgaacg gggggttcgt
      361 gcacacagcc cagcttggag cgaacgacct acaccgaact gagataccta cagcgtgagc
      421 tatgagaaag cgccacgctt cccgaaggga gaaaggcgga caggtatccg gtaagcggca
      481 gggtcggaac aggagagcgc acgagggagc ttccaggggg aaacgcctgg tatctttata
      541 gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg
      601 ggcggagcct atggaaaaac gccagcaacg cggccttttt acggttcctg gccttttgct
      661 ggccttttgc tcacatgttc tttcctgcgt tatcccctga ttctgtggat aaccgtatta
      721 ccgccatgca ttagttatta atagtaatca attacggggt cattagttca tagcccatat
      781 atggagttcc gcgttacata acttacggta aatggcccgc ctggctgacc gcccaacgac
      841 ccccgcccat tgacgtcaat aatgacgtat gttcccatag taacgccaat agggactttc
      901 cattgacgtc aatgggtgga gtatttacgg taaactgccc acttggcagt acatcaagtg
      961 tatcatatgc caagtacgcc ccctattgac gtcaatgacg gtaaatggcc cgcctggcat
     1021 tatgcccagt acatgacctt atgggacttt cctacttggc agtacatcta cgtattagtc
     1081 atcgctatta ccatggtgat gcggttttgg cagtacatca atgggcgtgg atagcggttt
     1141 gactcacggg gatttccaag tctccacccc attgacgtca atgggagttt gttttggcac
     1201 caaaatcaac gggactttcc aaaatgtcgt aacaactccg ccccattgac gcaaatgggc
     1261 ggtaggcgtg tacggtggga ggtctatata agcagagctg gtttagtgaa ccgtcagatc
     1321 cgctagcggc caccatggct ctggacggga taaggatgcc agatggctgc tacgcggacg
     1381 ggacgtggga actgagtgtc catgtgacgg acctgaaccg cgatgtcacc ctgagagtga
     1441 ccggcgaggt gcacattgga ggcgtgatgc ttaagctggt ggagaaactc gatgtaaaaa
     1501 aagattggtc tgaccatgct ctctggtggg aaaagaagag aacttggctt ctgaagacac
     1561 attggacctt agataagtat ggtattcagg cagatgctaa gcttcagttc acccctcagc
     1621 acaaactgct ccgcctgcag cttcccaaca tgaagtatgt gaaggtgaaa gtgaatttct
     1681 ctgatagagt cttcaaagct gtttctgaca tctgtaagac ttttaatatc agacaccccg
     1741 aagaactttc tctcttaaag aaacccagag atccaacaaa gaaaaaaaag aagaagctag
     1801 atgaccagtc tgaagatgag gcacttgaat tagaggggcc tcttatcact cctggatcag
     1861 gaagtatata ttcagcgcca ggactgtata gtaaaacaat gacccccact tatgatgctc
     1921 atgatggaag ccccttgtca ccaacttctg cttggtttgg tgacagtgct ttgtcagaag
     1981 gcaatcctgg tatacttgct gtcagtcaac caatcacgtc accagaaatc ttggcaaaaa
     2041 tgttcaagcc tcaagctctt cttgataaag caaaaatcaa ccaaggatgg cttgattcct
     2101 caagatctct catggaacaa gatgtgaagg aaaatgaggc cttgctgctc cgattcaagt
     2161 attacagctt ttttgatttg aatccaaagt atgatgcaat cagaatcaat cagctttatg
     2221 agcaggccaa atgggccatt ctcctggaag agattgaatg cacagaagaa gaaatgatga
     2281 tgtttgcagc cctgcagtat catatcaata agctgtcaat catgacatca gagaatcatt
     2341 tgaacaacag tgacaaagaa gttgatgaag ttgatgctgc cctttcagac ctggagatta
     2401 ctctggaagg gggtaaaacg tcaacaattt tgggtgacat tacttccatt cctgaacttg
     2461 ctgactacat taaagttttc aagccaaaaa agctgactct gaaaggttac aaacaatatt
     2521 ggtgcacctt caaagacaca tccatttctt gttataagag caaagaagaa tccagtggca
     2581 caccagctca tcagatgaac ctcaggggat gtgaagttac cccagatgta aacatttcag
     2641 gccaaaaatt taacattaaa ctcctgattc cagttgcaga aggcatgaat gaaatctggc
     2701 ttcgttgtga caatgaaaaa cagtatgcac actggatggc agcctgcaga ttagcctcca
     2761 aaggcaagac catggcggac agttcttaca acttagaagt tcagaatatt ctttcctttc
     2821 tgaagatgca gcatttaaac ccagatcctc agttaatacc agagcagatc acgactgata
     2881 taactcctga atgtttggtg tctccccgct atctaaaaaa gtataagaac aagcagataa
     2941 cagcgagaat cttggaggcc catcagaatg tagctcagat gagtctaatt gaagccaaga
     3001 tgagatttat tcaagcttgg cagtcactac ctgaatttgg catcactcac ttcattgcaa
     3061 ggttccaagg gggcaaaaaa gaagaactta ttggaattgc atacaacaga ctgattcgga
     3121 tggatgccag cactggagat gcaattaaaa catggcgttt cagcaacatg aaacagtgga
     3181 atgtcaactg ggaaatcaaa atggtcaccg tagagtttgc agatgaagta cgattgtcct
     3241 tcatttgtac tgaagtagat tgcaaagtgg ttcatgaatt cattggtggc tacatatttc
     3301 tctcaacacg tgcaaaagac caaaacgaga gtttagatga agagatgttc tacaaactta
     3361 ccagtggttg ggtgggtaac tcaggggatc caggaatggt gagcaagggc gaggagctgt
     3421 tcaccggggt ggtgcccatc ctggtcgagc tggacggcga cgtaaacggc cacaagttca
     3481 gcgtgtccgg cgagggcgag ggcgatgcca cctacggcaa gctgaccctg aagttcatct
     3541 gcaccaccgg caagctgccc gtgccctggc ccaccctcgt gaccaccctg acctacggcg
     3601 tgcagtgctt cagccgctac cccgaccaca tgaagcagca cgacttcttc aagtccgcca
     3661 tgcccgaagg ctacgtccag gagcgcacca tcttcttcaa ggacgacggc aactacaaga
     3721 cccgcgccga ggtgaagttc gagggcgaca ccctggtgaa ccgcatcgag ctgaagggca
     3781 tcgacttcaa ggaggacggc aacatcctgg ggcacaagct ggagtacaac tacaacagcc
     3841 acaacgtcta tatcatggcc gacaagcaga agaacggcat caaggtgaac ttcaagatcc
     3901 gccacaacat cgaggacggc agcgtgcagc tcgccgacca ctaccagcag aacaccccca
     3961 tcggcgacgg ccccgtgctg ctgcccgaca accactacct gagcacccag tccgccctga
     4021 gcaaagaccc caacgagaag cgcgatcaca tggtcctgct ggagttcgtg accgccgccg
     4081 ggatcactct cggcatggac gagctgtaca agtaaagcgg ccgcgactct agatcataat
     4141 cagccatacc acatttgtag aggttttact tgctttaaaa aacctcccac acctccccct
     4201 gaacctgaaa cataaaatga atgcaattgt tgttgttaac ttgtttattg cagcttataa
     4261 tggttacaaa taaagcaata gcatcacaaa tttcacaaat aaagcatttt tttcactgca
     4321 ttctagttgt ggtttgtcca aactcatcaa tgtatcttaa ggcgtaaatt gtaagcgtta
     4381 atattttgtt aaaattcgcg ttaaattttt gttaaatcag ctcatttttt aaccaatagg
     4441 ccgaaatcgg caaaatccct tataaatcaa aagaatagac cgagataggg ttgagtgttg
     4501 ttccagtttg gaacaagagt ccactattaa agaacgtgga ctccaacgtc aaagggcgaa
     4561 aaaccgtcta tcagggcgat ggcccactac gtgaaccatc accctaatca agttttttgg
     4621 ggtcgaggtg ccgtaaagca ctaaatcgga accctaaagg gagcccccga tttagagctt
     4681 gacggggaaa gccggcgaac gtggcgagaa aggaagggaa gaaagcgaaa ggagcgggcg
     4741 ctagggcgct ggcaagtgta gcggtcacgc tgcgcgtaac caccacaccc gccgcgctta
     4801 atgcgccgct acagggcgcg tcaggtggca cttttcgggg aaatgtgcgc ggaaccccta
     4861 tttgtttatt tttctaaata cattcaaata tgtatccgct catgagacaa taaccctgat
     4921 aaatgcttca ataatattga aaaaggaaga gtcctgaggc ggaaagaacc agctgtggaa
     4981 tgtgtgtcag ttagggtgtg gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag
     5041 catgcatctc aattagtcag caaccaggtg tggaaagtcc ccaggctccc cagcaggcag
     5101 aagtatgcaa agcatgcatc tcaattagtc agcaaccata gtcccgcccc taactccgcc
     5161 catcccgccc ctaactccgc ccagttccgc ccattctccg ccccatggct gactaatttt
     5221 ttttatttat gcagaggccg aggccgcctc ggcctctgag ctattccaga agtagtgagg
     5281 aggctttttt ggaggcctag gcttttgcaa agatcgatca agagacagga tgaggatcgt
     5341 ttcgcatgat tgaacaagat ggattgcacg caggttctcc ggccgcttgg gtggagaggc
     5401 tattcggcta tgactgggca caacagacaa tcggctgctc tgatgccgcc gtgttccggc
     5461 tgtcagcgca ggggcgcccg gttctttttg tcaagaccga cctgtccggt gccctgaatg
     5521 aactgcaaga cgaggcagcg cggctatcgt ggctggccac gacgggcgtt ccttgcgcag
     5581 ctgtgctcga cgttgtcact gaagcgggaa gggactggct gctattgggc gaagtgccgg
     5641 ggcaggatct cctgtcatct caccttgctc ctgccgagaa agtatccatc atggctgatg
     5701 caatgcggcg gctgcatacg cttgatccgg ctacctgccc attcgaccac caagcgaaac
     5761 atcgcatcga gcgagcacgt actcggatgg aagccggtct tgtcgatcag gatgatctgg
     5821 acgaagagca tcaggggctc gcgccagccg aactgttcgc caggctcaag gcgagcatgc
     5881 ccgacggcga ggatctcgtc gtgacccatg gcgatgcctg cttgccgaat atcatggtgg
     5941 aaaatggccg cttttctgga ttcatcgact gtggccggct gggtgtggcg gaccgctatc
     6001 aggacatagc gttggctacc cgtgatattg ctgaagagct tggcggcgaa tgggctgacc
     6061 gcttcctcgt gctttacggt atcgccgctc ccgattcgca gcgcatcgcc ttctatcgcc
     6121 ttcttgacga gttcttctga gcgggactct ggggttcgaa atgaccgacc aagcgacgcc
     6181 caacctgcca tcacgagatt tcgattccac cgccgccttc tatgaaaggt tgggcttcgg
     6241 aatcgttttc cgggacgccg gctggatgat cctccagcgc ggggatctca tgctggagtt
     6301 cttcgcccac cctaggggga ggctaactga aacacggaag gagacaatac cggaaggaac
     6361 ccgcgctatg acggcaataa aaagacagaa taaaacgcac ggtgttgggt cgtttgttca
     6421 taaacgcggg gttcggtccc agggctggca ctctgtcgat accccaccga gaccccattg
     6481 gggccaatac gcccgcgttt cttccttttc cccaccccac cccccaagtt cgggtgaagg
     6541 cccagggctc gcagccaacg tcggggcggc aggccctgcc atagcctcag gttactcata
     6601 tatactttag attgatttaa aacttcattt ttaatttaaa aggatctagg tgaagatcct
     6661 ttttgataat ctcatgacca aaatccctta acgtgagttt tcgttccact gagcgtcag
//
