LOCUS       sequence-361173-        6903 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  expression of minitalin wt in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6903)
  AUTHORS   Azizi L, Varela L, Turkki P, Mykuliak VV, Korpela S, Ihalainen TO, 
            Church J, Hytonen VP, Goult BT
  TITLE     Talin variant P229S compromises integrin activation and associates 
            with multifaceted clinical symptoms.
  JOURNAL   Hum Mol Genet. 2022 Jul 21. pii: 6647920. doi: 10.1093/hmg/ddac163.
  PUBMED    35861643
REFERENCE   2  (bases 1 to 6903)
  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"; journalName: "Hum Mol 
            Genet. 2022 Jul 21. pii: 6647920. doi: 10.1093/hmg/ddac163."
COMMENT     Sequence Label: minitalin (headR13DD)
FEATURES             Location/Qualifiers
     source          1..6903
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        61..364
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        365..568
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     519..539
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      644..653
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(2831..2833,2834..2836,2837..3550)
                     /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(2831..2833,2834..2836,2837..3547)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(2876..2897)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(3137..3156)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3484..3505
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    3691..3812
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3728..3747)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3782..3801
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(3819..4274)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3956..3977)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4168..4187
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        4301..4405
                     /gene="bla"
                     /label=AmpR promoter
     promoter        4407..4764
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        4407..4598
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(4430..4450)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      4615..4750
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             4799..5593
                     /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             4799..5593
                     /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(4853..4872)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5463..5482
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(5781..5800)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    5825..5872
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      6201..6789
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6690..6709
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg
       61 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
      121 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      181 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      241 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      301 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      361 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
      421 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
      481 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
      541 acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatcc gctagcgcta
      601 ccggtattag gcaccccagg ctttacacga attcgcagaa gctgccacca tggttgcgct
      661 ttcgctgaag attagcattg ggaatgtggt gaagacgatg caatttgagc catctaccat
      721 ggtgtacgac gcctgccgca tgattcgtga gcggatccca gaggccctgg ctggccctcc
      781 caacgacttt gggctctttc tgtcagatga tgaccccaaa aaaggcatct ggctagaggc
      841 tgggaaagct ttggactact acatgctccg aaatggggac accatggagt acagaaaaaa
      901 gcagagaccc ctgaagatcc ggatgttaga cggaacggtg aagactatca tggtggatga
      961 ctccaagacc gtcacagaca tgctcatgac catttgtgcc cgaattggta tcaccaacca
     1021 tgatgagtat tcactggttc gagagctgat ggaagaaaag aaagatgagg ggacagggac
     1081 cctgagaaaa gacaagaccc tactgcgaga tgaaaagaag atggagaagc taaagcagaa
     1141 gttgcacaca gacgatgagt tgaactggct ggaccatggg cggacactga gggaacaggg
     1201 agtggaagag catgagacgt tgctgctgcg gagaaagttc ttctactcag accagaatgt
     1261 ggattcccga gaccctgtac agctgaacct tctctatgtg caggcacgag atgacatcct
     1321 gaatggctcc catcctgtct cctttgacaa ggcctgtgaa tttgcaggct tccagtgcca
     1381 gatccagttt ggacctcaca atgaacagaa gcacaaggct ggcttccttg acctgaagga
     1441 cttcctgccc aaggagtatg tgaagcagaa gggagagcgt aagatctttc aggcacacaa
     1501 gaattgtggg cagatgagtg aaattgaggc caaggtacgc tacgtgaagc tggcccgttc
     1561 cctcaagact tacggtgtct ccttcttcct agtcaaggaa aagatgaagg ggaagaataa
     1621 actggtgccc aggctgttgg gcatcactaa ggagtgtgtg atgcgtgtgg atgagaagac
     1681 caaggaggtg attcaggaat ggagcctcac caacatcaaa cgctgggctg cctctcccaa
     1741 gagctttact ctggactttg gagactatca ggatggctac tactcagtac agacgacaga
     1801 aggcgagcag atcgcacagc tcattgctgg ctacatagat atcatcctta agaagaaaaa
     1861 aagcaaggac cattttgggc tggagggaga tgaagagtct actatgttgg aggactcagt
     1921 gtctcccaaa aagtcaacag tccttcagca gcagtacaac cgagtgggga aagtggagca
     1981 cggctctgtg gctctgccag ccatcatgcg ctctggagcc tctggtcctg agaatttcca
     2041 ggtggggagc atgccacctg cccagcagca gatcaccagt ggccagatgc acgaccctac
     2101 tgtcattgct gagaatgagc tcttgggagc cgcagccgcc atcgaggctg cagccaagaa
     2161 gctggagcag ctgaagccca gggccaaacc caaggaggcg gatgagtcct tgaactttga
     2221 ggaacaaatc ctagaagctg ccaagtccat cgctgcagcc accagtgcac tggtaaaggc
     2281 tgcgtcagca gcccagaggg agctggtggc tcaaggaaag gtgggcgcca ttccagccaa
     2341 tgcactggat gatgggcagt ggtcgcaggg cctcatttcc gctgcccgta tggtggctgc
     2401 ggccaccaac aatctgtgtg aagcagctaa cgcagctgtc cagggccatg ctagccagga
     2461 gaaactcatt tcctcagcca agcaagtagc tgcctccaca gctcagctcc tggtagcttg
     2521 caaggtcaaa gcagatcagg actctgaggc aatgaaacgg ctccaggctg ctggcaatgc
     2581 agtgaagagg gcctcagata acctggtaaa ggcggcccag aaagctgcag ccttcgaaga
     2641 ccaggagaat gagacggtgg tggtgaagga gaaaatggtt gggggcattg cccagattat
     2701 cgcagcacag gaagagatgc ttcggaagga acgagagctg gaagaggctc ggaaaaagct
     2761 cgcccagatc cggcagcagc agtacaagtt cttgccttca gagcttcgag acgagcacgg
     2821 aggatccgga atggtgagca agggcgagga gctgttcacc ggggtggtgc ccatcctggt
     2881 cgagctggac ggcgacgtaa acggccacaa gttcagcgtg tccggcgagg gcgagggcga
     2941 tgccacctac ggcaagctga ccctgaagtt catctgcacc accggcaagc tgcccgtgcc
     3001 ctggcccacc ctcgtgacca ccctgaccta cggcgtgcag tgcttcagcc gctaccccga
     3061 ccacatgaag cagcacgact tcttcaagtc cgccatgccc gaaggctacg tccaggagcg
     3121 caccatcttc ttcaaggacg acggcaacta caagacccgc gccgaggtga agttcgaggg
     3181 cgacaccctg gtgaaccgca tcgagctgaa gggcatcgac ttcaaggagg acggcaacat
     3241 cctggggcac aagctggagt acaactacaa cagccacaac gtctatatca tggccgacaa
     3301 gcagaagaac ggcatcaagg tgaacttcaa gatccgccac aacatcgagg acggcagcgt
     3361 gcagctcgcc gaccactacc agcagaacac ccccatcggc gacggccccg tgctgctgcc
     3421 cgacaaccac tacctgagca cccagtccgc cctgagcaaa gaccccaacg agaagcgcga
     3481 tcacatggtc ctgctggagt tcgtgaccgc cgccgggatc actctcggca tggacgagct
     3541 gtacaagtaa ccgcgggccc gggatccacc ggatctagat aactgatcat aatcagccat
     3601 accacatttg tagaggtttt acttgcttta aaaaacctcc cacacctccc cctgaacctg
     3661 aaacataaaa tgaatgcaat tgttgttgtt aacttgttta ttgcagctta taatggttac
     3721 aaataaagca atagcatcac aaatttcaca aataaagcat ttttttcact gcattctagt
     3781 tgtggtttgt ccaaactcat caatgtatct taacgcgtaa attgtaagcg ttaatatttt
     3841 gttaaaattc gcgttaaatt tttgttaaat cagctcattt tttaaccaat aggccgaaat
     3901 cggcaaaatc ccttataaat caaaagaata gaccgagata gggttgagtg ttgttccagt
     3961 ttggaacaag agtccactat taaagaacgt ggactccaac gtcaaagggc gaaaaaccgt
     4021 ctatcagggc gatggcccac tacgtgaacc atcaccctaa tcaagttttt tggggtcgag
     4081 gtgccgtaaa gcactaaatc ggaaccctaa agggagcccc cgatttagag cttgacgggg
     4141 aaagccggcg aacgtggcga gaaaggaagg gaagaaagcg aaaggagcgg gcgctagggc
     4201 gctggcaagt gtagcggtca cgctgcgcgt aaccaccaca cccgccgcgc ttaatgcgcc
     4261 gctacagggc gcgtcaggtg gcacttttcg gggaaatgtg cgcggaaccc ctatttgttt
     4321 atttttctaa atacattcaa atatgtatcc gctcatgaga caataaccct gataaatgct
     4381 tcaataatat tgaaaaagga agagtcctga ggcggaaaga accagctgtg gaatgtgtgt
     4441 cagttagggt gtggaaagtc cccaggctcc ccagcaggca gaagtatgca aagcatgcat
     4501 ctcaattagt cagcaaccag gtgtggaaag tccccaggct ccccagcagg cagaagtatg
     4561 caaagcatgc atctcaatta gtcagcaacc atagtcccgc ccctaactcc gcccatcccg
     4621 cccctaactc cgcccagttc cgcccattct ccgccccatg gctgactaat tttttttatt
     4681 tatgcagagg ccgaggccgc ctcggcctct gagctattcc agaagtagtg aggaggcttt
     4741 tttggaggcc taggcttttg caaagatcga tcaagagaca ggatgaggat cgtttcgcat
     4801 gattgaacaa gatggattgc acgcaggttc tccggccgct tgggtggaga ggctattcgg
     4861 ctatgactgg gcacaacaga caatcggctg ctctgatgcc gccgtgttcc ggctgtcagc
     4921 gcaggggcgc ccggttcttt ttgtcaagac cgacctgtcc ggtgccctga atgaactgca
     4981 agacgaggca gcgcggctat cgtggctggc cacgacgggc gttccttgcg cagctgtgct
     5041 cgacgttgtc actgaagcgg gaagggactg gctgctattg ggcgaagtgc cggggcagga
     5101 tctcctgtca tctcaccttg ctcctgccga gaaagtatcc atcatggctg atgcaatgcg
     5161 gcggctgcat acgcttgatc cggctacctg cccattcgac caccaagcga aacatcgcat
     5221 cgagcgagca cgtactcgga tggaagccgg tcttgtcgat caggatgatc tggacgaaga
     5281 gcatcagggg ctcgcgccag ccgaactgtt cgccaggctc aaggcgagca tgcccgacgg
     5341 cgaggatctc gtcgtgaccc atggcgatgc ctgcttgccg aatatcatgg tggaaaatgg
     5401 ccgcttttct ggattcatcg actgtggccg gctgggtgtg gcggaccgct atcaggacat
     5461 agcgttggct acccgtgata ttgctgaaga gcttggcggc gaatgggctg accgcttcct
     5521 cgtgctttac ggtatcgccg ctcccgattc gcagcgcatc gccttctatc gccttcttga
     5581 cgagttcttc tgagcgggac tctggggttc gaaatgaccg accaagcgac gcccaacctg
     5641 ccatcacgag atttcgattc caccgccgcc ttctatgaaa ggttgggctt cggaatcgtt
     5701 ttccgggacg ccggctggat gatcctccag cgcggggatc tcatgctgga gttcttcgcc
     5761 caccctaggg ggaggctaac tgaaacacgg aaggagacaa taccggaagg aacccgcgct
     5821 atgacggcaa taaaaagaca gaataaaacg cacggtgttg ggtcgtttgt tcataaacgc
     5881 ggggttcggt cccagggctg gcactctgtc gataccccac cgagacccca ttggggccaa
     5941 tacgcccgcg tttcttcctt ttccccaccc caccccccaa gttcgggtga aggcccaggg
     6001 ctcgcagcca acgtcggggc ggcaggccct gccatagcct caggttactc atatatactt
     6061 tagattgatt taaaacttca tttttaattt aaaaggatct aggtgaagat cctttttgat
     6121 aatctcatga ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc agaccccgta
     6181 gaaaagatca aaggatcttc ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa
     6241 acaaaaaaac caccgctacc agcggtggtt tgtttgccgg atcaagagct accaactctt
     6301 tttccgaagg taactggctt cagcagagcg cagataccaa atactgttct tctagtgtag
     6361 ccgtagttag gccaccactt caagaactct gtagcaccgc ctacatacct cgctctgcta
     6421 atcctgttac cagtggctgc tgccagtggc gataagtcgt gtcttaccgg gttggactca
     6481 agacgatagt taccggataa ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag
     6541 cccagcttgg agcgaacgac ctacaccgaa ctgagatacc tacagcgtga gctatgagaa
     6601 agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga
     6661 acaggagagc gcacgaggga gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc
     6721 gggtttcgcc acctctgact tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc
     6781 ctatggaaaa acgccagcaa cgcggccttt ttacggttcc tggccttttg ctggcctttt
     6841 gctcacatgt tctttcctgc gttatcccct gattctgtgg ataaccgtat taccgccatg
     6901 cat
//
