LOCUS       sequence-365818-        6957 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  For expression of recombinant his10-beta1 integrin cytosolic domain 
            in bacteria.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6957)
  AUTHORS   Case LB, De Pasquale M, Henry L, Rosen MK
  TITLE     Synergistic phase separation of two pathways promotes integrin 
            clustering and nascent adhesion formation.
  JOURNAL   Elife. 2022 Jan 20;11. pii: 72588. doi: 10.7554/eLife.72588.
  PUBMED    35049497
REFERENCE   2  (bases 1 to 6957)
  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: "Elife."; 
            date: "2022-01-20"; volume: "11. pii: 72588. doi"; pages: " 
            10.7554/eLife.72588"
COMMENT     Sequence Label: His10-B1 Integrin
FEATURES             Location/Qualifiers
     source          1..6957
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      1..589
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     490..509
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     743..760
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     complement(866..885)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     1026..1048
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(1042..1233)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     promoter        1729..1806
                     /gene="lacI (mutant)"
                     /label=lacIq promoter
                     /note="In the lacIq allele, a single base change in the 
                     promoter boosts expression of the lacI gene about 10-fold."
     CDS             1807..2889
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /label=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)."
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     complement(1826..1845)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        join(3132..3137,3138..3153,3154..3160)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    3168..3184
                     /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)."
     primer_bind     3173..3195
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     CDS             3254..4354
                     /codon_start=1
                     /gene="malE (mutated)"
                     /product="maltose binding protein from E. coli"
                     /label=MBP
                     /note="This version of the gene does not encode a signal 
                     sequence, so MBP will remain in the cytosol."
                     /translation="MKTEEGKLVIWINGDKGYNGLAEVGKKFEKDTGIKVTVEHPDKLE
                     EKFPQVAATGDGPDIIFWAHDRFGGYAQSGLLAEITPDKAFQDKLYPFTWDAVRYNGKL
                     IAYPIAVEALSLIYNKDLLPNPPKTLEEIPALDKELKAKGKSALMFNLQEPYFTWPLIA
                     ADGGYAFKYENGKYDIKDVGVDNAGAKAGLTFLVDLIKNKHMNADTDYSIAEAAFNKGE
                     TAMTINGPWAWSNIDTSKVNYGVTVLPTFKGQPSKPFVGVLSAGINAASPNKELAKEFL
                     ENYLLTDEGLEAVNKDKPLGAVALKSYEEELAKDPRIAATMENAQKGEIMPNIPQMSAF
                     WYAVRTAVINAASGRQTVDEALKDAQT"
     regulatory      4211..4220
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     4328..4351
                     /label=MBP-F
                     /note="Maltose binding protein, forward primer"
     CDS             4403..4414
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     CDS             4427..4447
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             4460..4489
                     /codon_start=1
                     /product="10xHis affinity tag"
                     /label=10xHis
                     /translation="HHHHHHHHHH"
     CDS             4721..4738
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(4751..4768)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4751..4767)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4760..4782)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     complement(4946..4963)
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     terminator      5116..5202
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      5294..5321
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     promoter        5340..5431
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5432..5500,5501..6292)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(5650..5669)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      complement(6334..6847)
                     /direction=LEFT
                     /label=M13 ori
                     /note="M13 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6529..6550)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     6741..6760
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
       61 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
      121 cagcagagcg cagataccaa atactgtcct tctagtgtag ccgtagttag gccaccactt
      181 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
      241 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
      301 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
      361 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg
      421 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
      481 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
      541 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa
      601 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc
      661 gttatcccct gattctgtgg ataaccgtat taccgccttt gagtgagctg ataccgctcg
      721 ccgcagccga acgaccgagc gcagcgagtc agtgagcgag gaagcggaag agcgcctgat
      781 gcggtatttt ctccttacgc atctgtgcgg tatttcacac cgcatgtggt gcactctcag
      841 tacaccgcat gtggtgcact ctcagtacaa tctgctctga tgccgcatag ttaagccagt
      901 atacactccg ctatcgctac gtgactgggt catggctgcg ccccgacacc cgccaacacc
      961 cgctgacgcg ccctgacggg cttgtctgct cccggcatcc gcttacagac aagctgtgac
     1021 cgtctccggg agctgcatgt gtcagaggtt ttcaccgtca tcaccgaaac gcgcgaggca
     1081 gctgcggtaa agctcatcag cgtggtcgtg cagcgattca cagatgtctg cctgttcatc
     1141 cgcgtccagc tcgttgagtt tctccagaag cgttaatgtc tggcttctga taaagcgggc
     1201 catgttaagg gcggtttttt cctgtttggt cactgatgcc tccgtgtaag ggggatttct
     1261 gttcatgggg gtaatgatac cgatgaaacg agagaggatg ctcacgatac gggttactga
     1321 tgatgaacat gcccggttac tggaacgttg tgagggtaaa caactggcgg tatggatgcg
     1381 gcgggaccag agaaaaatca ctcagggtca atgccagcgc ttcgttaata cagatgtagg
     1441 tgttccacag ggtagccagc agcatcctgc gatgcagatc cggaacataa tggtgcaggg
     1501 cgctgacttc cgcgtttcca gactttacga aacacggaaa ccgaagacca ttcatgttgt
     1561 tgctcaggtc gcagacgttt tgcagcagca gtcgcttcac gttcgctcgc gtatcggtga
     1621 ttcattctgc taaccagtaa ggcaaccccg ccagcctagc cgggtcctca acgacaggag
     1681 cacgatcatg cgcacccgtg gccaggaccc aacgctgccc gaaattccga caccatcgaa
     1741 tggtgcaaaa cctttcgcgg tatggcatga tagcgcccgg aagagagtca attcagggtg
     1801 gtgaatgtga aaccagtaac gttatacgat gtcgcagagt atgccggtgt ctcttatcag
     1861 accgtttccc gcgtggtgaa ccaggccagc cacgtttctg cgaaaacgcg ggaaaaagtg
     1921 gaagcggcga tggcggagct gaattacatt cccaaccgcg tggcacaaca actggcgggc
     1981 aaacagtcgt tgctgattgg cgttgccacc tccagtctgg ccctgcacgc gccgtcgcaa
     2041 attgtcgcgg cgattaaatc tcgcgccgat caactgggtg ccagcgtggt ggtgtcgatg
     2101 gtagaacgaa gcggcgtcga agcctgtaaa gcggcggtgc acaatcttct cgcgcaacgc
     2161 gtcagtgggc tgatcattaa ctatccgctg gatgaccagg atgccattgc tgtggaagct
     2221 gcctgcacta atgttccggc gttatttctt gatgtctctg accagacacc catcaacagt
     2281 attattttct cccatgaaga cggtacgcga ctgggcgtgg agcatctggt cgcattgggt
     2341 caccagcaaa tcgcgctgtt agcgggccca ttaagttctg tctcggcgcg tctgcgtctg
     2401 gctggctggc ataaatatct cactcgcaat caaattcagc cgatagcgga acgggaaggc
     2461 gactggagtg ccatgtccgg ttttcaacaa accatgcaaa tgctgaatga gggcatcgtt
     2521 cccactgcga tgctggttgc caacgatcag atggcgctgg gcgcaatgcg cgccattacc
     2581 gagtccgggc tgcgcgttgg tgcggatatc tcggtagtgg gatacgacga taccgaagac
     2641 agctcatgtt atatcccgcc gttaaccacc atcaaacagg attttcgcct gctggggcaa
     2701 accagcgtgg accgcttgct gcaactctct cagggccagg cggtgaaggg caatcagctg
     2761 ttgcccgtct cactggtgaa aagaaaaacc accctggcgc ccaatacgca aaccgcctct
     2821 ccccgcgcgt tggccgattc attaatgcag ctggcacgac aggtttcccg actggaaagc
     2881 gggcagtgag cgcaacgcaa ttaatgtaag ttagctcact cattaggcac aattctcatg
     2941 tttgacagct tatcatcgac tgcacggtgc accaatgctt ctggcgtcag gcagccatcg
     3001 gaagctgtgg tatggctgtg caggtcgtaa atcactgcat aattcgtgtc gctcaaggcg
     3061 cactcccgtt ctggataatg ttttttgcgc cgacatcata acggttctgg caaatattct
     3121 gaaatgagct gttgacaatt aatcatcggc tcgtataatg tgtggaattg tgagcggata
     3181 acaatttcac acaggaaaca gccagtccgt ttaggtgttt tcacgagcac ttcaccaaca
     3241 aggaccatag attatgaaaa ctgaagaagg taaactggta atctggatta acggcgataa
     3301 aggctataac ggtctcgctg aagtcggtaa gaaattcgag aaagataccg gaattaaagt
     3361 caccgttgag catccggata aactggaaga gaaattccca caggttgcgg caactggcga
     3421 tggccctgac attatcttct gggcacacga ccgctttggt ggctacgctc aatctggcct
     3481 gttggctgaa atcaccccgg acaaagcgtt ccaggacaag ctgtatccgt ttacctggga
     3541 tgccgtacgt tacaacggca agctgattgc ttacccgatc gctgttgaag cgttatcgct
     3601 gatttataac aaagacctgc tgccgaaccc gccaaaaacc ttggaagaga tcccggcgct
     3661 ggataaagaa ctgaaagcga aaggtaagag cgcgctgatg ttcaacctgc aagaaccgta
     3721 cttcacctgg ccgctgattg ctgctgacgg gggttatgcg ttcaagtatg aaaacggcaa
     3781 gtacgacatt aaagacgtgg gcgtggataa cgctggcgcg aaagcgggtc tgaccttcct
     3841 ggttgacctg attaaaaaca aacacatgaa tgcagacacc gattactcca tcgcagaagc
     3901 tgcctttaat aaaggcgaaa cagcgatgac catcaacggc ccgtgggcat ggtccaacat
     3961 cgacaccagc aaagtgaatt atggtgtaac ggtactgccg accttcaagg gtcaaccatc
     4021 caaaccgttc gttggcgtgc tgagcgcagg tattaacgcc gccagtccga acaaagagct
     4081 ggcaaaagag ttcctcgaaa actatctgct gactgatgaa ggtctggaag cggttaataa
     4141 agacaaaccg ctgggtgccg tagcgctgaa gtcttacgag gaagagttgg cgaaagatcc
     4201 acgtattgcc gccaccatgg aaaacgccca gaaaggtgaa atcatgccga acatcccgca
     4261 gatgtccgct ttctggtatg ccgtgcgtac tgcggtgatc aacgccgcca gcggtcgtca
     4321 gactgtcgat gaagccctga aagacgcgca gactaattcg agctcgaaca acaacaacaa
     4381 taacaataac aacaacctcg ggatcgaggg aaggatttca gaatttgaaa acctgtattt
     4441 tcagggcggt tcgctcgagc accaccatca ccatcaccat caccatcatg gtggttcatg
     4501 cggggggtct ggtggttcag ggggcagtgg cggttctaaa ctgcttatga tcattcatga
     4561 tcgccgcgaa tttgccaaat ttgagaagga aaaaatgaac gctaaatggg atacgggcga
     4621 aaaccccatt tacaagtcag ctgtgacgac cgttgtaaac ccgaagtatg aggggaaata
     4681 gtgataagga tcctctagag tcgacctgca ggcaagcttg catcaccacc atcaccatta
     4741 atagcttggc actggccgtc gttttacaac gtcgtgactg ggaaaaccct ggcgttaccc
     4801 aacttaatcg ccttgcagca catccccctt tcgccagctg gcgtaatagc gaagaggccc
     4861 gcaccgatcg cccttcccaa cagttgcgca gcctgaatgg cgaatggcag cttggctgtt
     4921 ttggcggatg agataagatt ttcagcctga tacagattaa atcagaacgc agaagcggtc
     4981 tgataaaaca gaatttgcct ggcggcagta gcgcggtggt cccacctgac cccatgccga
     5041 actcagaagt gaaacgccgt agcgccgatg gtagtgtggg gtctccccat gcgagagtag
     5101 ggaactgcca ggcatcaaat aaaacgaaag gctcagtcga aagactgggc ctttcgtttt
     5161 atctgttgtt tgtcggtgaa cgctctcctg agtaggacaa atccgccggg agcggatttg
     5221 aacgttgcga agcaacggcc cggagggtgg cgggcaggac gcccgccata aactgccagg
     5281 catcaaatta agcagaaggc catcctgacg gatggccttt ttgcgtttct acaaactctt
     5341 ttgtttattt ttctaaatac attcaaatat gtatccgctc atgagacaat aaccctgata
     5401 aatgcttcaa taatattgaa aaaggaagag tatgagtatt caacatttcc gtgtcgccct
     5461 tattcccttt tttgcggcat tttgccttcc tgtttttgct cacccagaaa cgctggtgaa
     5521 agtaaaagat gctgaagatc agttgggtgc acgagtgggt tacatcgaac tggatctcaa
     5581 cagcggtaag atccttgaga gttttcgccc cgaagaacgt ttcccaatga tgagcacttt
     5641 taaagttctg ctatgtggcg cggtattatc ccgtgttgac gccgggcaag agcaactcgg
     5701 tcgccgcata cactattctc agaatgactt ggttgagtac tcaccagtca cagaaaagca
     5761 tcttacggat ggcatgacag taagagaatt atgcagtgct gccataacca tgagtgataa
     5821 cactgcggcc aacttacttc tgacaacgat cggaggaccg aaggagctaa ccgctttttt
     5881 gcacaacatg ggggatcatg taactcgcct tgatcgttgg gaaccggagc tgaatgaagc
     5941 cataccaaac gacgagcgtg acaccacgat gcctgtagca atggcaacaa cgttgcgcaa
     6001 actattaact ggcgaactac ttactctagc ttcccggcaa caattaatag actggatgga
     6061 ggcggataaa gttgcaggac cacttctgcg ctcggccctt ccggctggct ggtttattgc
     6121 tgataaatct ggagccggtg agcgtgggtc tcgcggtatc attgcagcac tggggccaga
     6181 tggtaagccc tcccgtatcg tagttatcta cacgacgggg agtcaggcaa ctatggatga
     6241 acgaaataga cagatcgctg agataggtgc ctcactgatt aagcattggt aactgtcaga
     6301 ccaagtttac tcatatatac tttagattga tttaccccgg ttgataatca gaaaagcccc
     6361 aaaaacagga agattgtata agcaaatatt taaattgtaa acgttaatat tttgttaaaa
     6421 ttcgcgttaa atttttgtta aatcagctca ttttttaacc aataggccga aatcggcaaa
     6481 atcccttata aatcaaaaga atagcccgag atagggttga gtgttgttcc agtttggaac
     6541 aagagtccac tattaaagaa cgtggactcc aacgtcaaag ggcgaaaaac cgtctatcag
     6601 ggcgatggcc cactacgtga accatcaccc aaatcaagtt ttttggggtc gaggtgccgt
     6661 aaagcactaa atcggaaccc taaagggagc ccccgattta gagcttgacg gggaaagccg
     6721 gcgaacgtgg cgagaaagga agggaagaaa gcgaaaggag cgggcgctag ggcgctggca
     6781 agtgtagcgg tcacgctgcg cgtaaccacc acacccgccg cgcttaatgc gccgctacag
     6841 ggcgcgtaaa aggatctagg tgaagatcct ttttgataat ctcatgacca aaatccctta
     6901 acgtgagttt tcgttccact gagcgtcaga ccccgtagaa aagatcaaag gatcttc
//
