LOCUS       sequence-194024-        5786 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Alive subunit for creating monovalent streptavidin with a single 
            femtomolar biotin binding site. Can also be used to create wild-type
            (tetravalent) streptavidin. .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5786)
  AUTHORS   Howarth M, Chinnapen DJ, Gerrow K, Dorrestein PC, Grandy MR, 
            Kelleher NL, El-Husseini A, Ting AY
  TITLE     A monovalent streptavidin with a single femtomolar biotin binding 
            site.
  JOURNAL   Nat Methods. 2006 Apr . 3(4):267-73.
  PUBMED    16554831
REFERENCE   2  (bases 1 to 5786)
  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: "Nat 
            Methods. 2006 Apr . 3(4):267-73."
COMMENT     Sequence Label: pET21a-Streptavidin-Alive
FEATURES             Location/Qualifiers
     source          1..5786
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             1241..1432
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     complement(1426..1448)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    1534..1673
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     1589..1608
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(1688..1705)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1859..2447)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1939..1958)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2618..3409,3410..3478))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     3241..3260
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3479..3583)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(3609..4064)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3746..3767)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3958..3977
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     terminator      complement(4101..4148)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     4144..4162
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     CDS             complement(4215..4232)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             complement(4260..4277)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     RBS             4669..4691
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     protein_bind    4706..4730
                     /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     complement(4730..4749)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(4731..4749)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(4833..4852)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     promoter        5058..5135
                     /gene="lacI"
                     /label=lacI promoter
     CDS             join(5136..5786,1..432)
                     /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(5155..5174)
                     /label=LacI-R
                     /note="LacI, reverse primer"
ORIGIN
        1 ggcgactgga gtgccatgtc cggttttcaa caaaccatgc aaatgctgaa tgagggcatc
       61 gttcccactg cgatgctggt tgccaacgat cagatggcgc tgggcgcaat gcgcgccatt
      121 accgagtccg ggctgcgcgt tggtgcggat atctcggtag tgggatacga cgataccgaa
      181 gacagctcat gttatatccc gccgttaacc accatcaaac aggattttcg cctgctgggg
      241 caaaccagcg tggaccgctt gctgcaactc tctcagggcc aggcggtgaa gggcaatcag
      301 ctgttgcccg tctcactggt gaaaagaaaa accaccctgg cgcccaatac gcaaaccgcc
      361 tctccccgcg cgttggccga ttcattaatg cagctggcac gacaggtttc ccgactggaa
      421 agcgggcagt gagcgcaacg caattaatgt aagttagctc actcattagg caccgggatc
      481 tcgaccgatg cccttgagag ccttcaaccc agtcagctcc ttccggtggg cgcggggcat
      541 gactatcgtc gccgcactta tgactgtctt ctttatcatg caactcgtag gacaggtgcc
      601 ggcagcgctc tgggtcattt tcggcgagga ccgctttcgc tggagcgcga cgatgatcgg
      661 cctgtcgctt gcggtattcg gaatcttgca cgccctcgct caagccttcg tcactggtcc
      721 cgccaccaaa cgtttcggcg agaagcaggc cattatcgcc ggcatggcgg ccccacgggt
      781 gcgcatgatc gtgctcctgt cgttgaggac ccggctaggc tggcggggtt gccttactgg
      841 ttagcagaat gaatcaccga tacgcgagcg aacgtgaagc gactgctgct gcaaaacgtc
      901 tgcgacctga gcaacaacat gaatggtctt cggtttccgt gtttcgtaaa gtctggaaac
      961 gcggaagtca gcgccctgca ccattatgtt ccggatctgc atcgcaggat gctgctggct
     1021 accctgtgga acacctacat ctgtattaac gaagcgctgg cattgaccct gagtgatttt
     1081 tctctggtcc cgccgcatcc ataccgccag ttgtttaccc tcacaacgtt ccagtaaccg
     1141 ggcatgttca tcatcagtaa cccgtatcgt gagcatcctc tctcgtttca tcggtatcat
     1201 tacccccatg aacagaaatc ccccttacac ggaggcatca gtgaccaaac aggaaaaaac
     1261 cgcccttaac atggcccgct ttatcagaag ccagacatta acgcttctgg agaaactcaa
     1321 cgagctggac gcggatgaac aggcagacat ctgtgaatcg cttcacgacc acgctgatga
     1381 gctttaccgc agctgcctcg cgcgtttcgg tgatgacggt gaaaacctct gacacatgca
     1441 gctcccggag acggtcacag cttgtctgta agcggatgcc gggagcagac aagcccgtca
     1501 gggcgcgtca gcgggtgttg gcgggtgtcg gggcgcagcc atgacccagt cacgtagcga
     1561 tagcggagtg tatactggct taactatgcg gcatcagagc agattgtact gagagtgcac
     1621 cattgcggtg tgaaataccg cacagatgcg taaggagaaa ataccgcatc aggcgctctt
     1681 ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag
     1741 ctcactcaaa ggcggtaata cggttatcca cagaatcagg ggataacgca ggaaagaaca
     1801 tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt
     1861 tccataggct ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc
     1921 gaaacccgac aggactataa agataccagg cgtttccccc tggaagctcc ctcgtgcgct
     1981 ctcctgttcc gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg
     2041 tggcgctttc tcatagctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca
     2101 agctgggctg tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact
     2161 atcgtcttga gtccaacccg gtaagacacg acttatcgcc actggcagca gccactggta
     2221 acaggattag cagagcgagg tatgtaggcg gtgctacaga gttcttgaag tggtggccta
     2281 actacggcta cactagaagg acagtatttg gtatctgcgc tctgctgaag ccagttacct
     2341 tcggaaaaag agttggtagc tcttgatccg gcaaacaaac caccgctggt agcggtggtt
     2401 tttttgtttg caagcagcag attacgcgca gaaaaaaagg atctcaagaa gatcctttga
     2461 tcttttctac ggggtctgac gctcagtgga acgaaaactc acgttaaggg attttggtca
     2521 tgagattatc aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga agttttaaat
     2581 caatctaaag tatatatgag taaacttggt ctgacagtta ccaatgctta atcagtgagg
     2641 cacctatctc agcgatctgt ctatttcgtt catccatagt tgcctgactc cccgtcgtgt
     2701 agataactac gatacgggag ggcttaccat ctggccccag tgctgcaatg ataccgcgag
     2761 acccacgctc accggctcca gatttatcag caataaacca gccagccgga agggccgagc
     2821 gcagaagtgg tcctgcaact ttatccgcct ccatccagtc tattaattgt tgccgggaag
     2881 ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt gctgcaggca
     2941 tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag ctccggttcc caacgatcaa
     3001 ggcgagttac atgatccccc atgttgtgca aaaaagcggt tagctccttc ggtcctccga
     3061 tcgttgtcag aagtaagttg gccgcagtgt tatcactcat ggttatggca gcactgcata
     3121 attctcttac tgtcatgcca tccgtaagat gcttttctgt gactggtgag tactcaacca
     3181 agtcattctg agaatagtgt atgcggcgac cgagttgctc ttgcccggcg tcaatacggg
     3241 ataataccgc gccacatagc agaactttaa aagtgctcat cattggaaaa cgttcttcgg
     3301 ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg
     3361 cacccaactg atcttcagca tcttttactt tcaccagcgt ttctgggtga gcaaaaacag
     3421 gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg gaaatgttga atactcatac
     3481 tcttcctttt tcaatattat tgaagcattt atcagggtta ttgtctcatg agcggataca
     3541 tatttgaatg tatttagaaa aataaacaaa taggggttcc gcgcacattt ccccgaaaag
     3601 tgccacctaa attgtaagcg ttaatatttt gttaaaattc gcgttaaatt tttgttaaat
     3661 cagctcattt tttaaccaat aggccgaaat cggcaaaatc ccttataaat caaaagaata
     3721 gaccgagata gggttgagtg ttgttccagt ttggaacaag agtccactat taaagaacgt
     3781 ggactccaac gtcaaagggc gaaaaaccgt ctatcagggc gatggcccac tacgtgaacc
     3841 atcaccctaa tcaagttttt tggggtcgag gtgccgtaaa gcactaaatc ggaaccctaa
     3901 agggagcccc cgatttagag cttgacgggg aaagccggcg aacgtggcga gaaaggaagg
     3961 gaagaaagcg aaaggagcgg gcgctagggc gctggcaagt gtagcggtca cgctgcgcgt
     4021 aaccaccaca cccgccgcgc ttaatgcgcc gctacagggc gcgtcccatt cgccaatccg
     4081 gatatagttc ctcctttcag caaaaaaccc ctcaagaccc gtttagaggc cccaaggggt
     4141 tatgctagtt attgctcagc ggtggcagca gccaactcag cttcctttcg ggctttgtta
     4201 gcagccggat ctcagtggtg gtggtggtgg tgctcgagtg cggccgcaag cttttattaa
     4261 tggtggtgat ggtgatggga agcagcggac ggtttaactt tggtgaaggt gtcgtgacca
     4321 accagggtgg atttccacgc gttggcttcg gtggtgccgg aggtcaacaa ccactgggtg
     4381 ttgatacgag cttcagcacc accaacgtac tggccagacc aggtggtagc ggagtgagcg
     4441 ttacggtagt tgtttttcca agcaacggtc caacccagag cggttccgga accgtcggta
     4501 gccggagcgg agtcgtaacg accggtcaga acgtatctag attcagcgtt accaacagcg
     4561 gattcgtagg taccggtcag agcaccgtca gcaccagcgg taacgatgaa ggtggatccc
     4621 agctggttgt accaggtgcc ggtgatacca gcttcagcca tatgtatatc tccttcttaa
     4681 agttaaacaa aattatttct agaggggaat tgttatccgc tcacaattcc cctatagtga
     4741 gtcgtattaa tttcgcggga tcgagatctc gatcctctac gccggacgca tcgtggccgg
     4801 catcaccggc gccacaggtg cggttgctgg cgcctatatc gccgacatca ccgatgggga
     4861 agatcgggct cgccacttcg ggctcatgag cgcttgtttc ggcgtgggta tggtggcagg
     4921 ccccgtggcc gggggactgt tgggcgccat ctccttgcat gcaccattcc ttgcggcggc
     4981 ggtgctcaac ggcctcaacc tactactggg ctgcttccta atgcaggagt cgcataaggg
     5041 agagcgtcga gatcccggac accatcgaat ggcgcaaaac ctttcgcggt atggcatgat
     5101 agcgcccgga agagagtcaa ttcagggtgg tgaatgtgaa accagtaacg ttatacgatg
     5161 tcgcagagta tgccggtgtc tcttatcaga ccgtttcccg cgtggtgaac caggccagcc
     5221 acgtttctgc gaaaacgcgg gaaaaagtgg aagcggcgat ggcggagctg aattacattc
     5281 ccaaccgcgt ggcacaacaa ctggcgggca aacagtcgtt gctgattggc gttgccacct
     5341 ccagtctggc cctgcacgcg ccgtcgcaaa ttgtcgcggc gattaaatct cgcgccgatc
     5401 aactgggtgc cagcgtggtg gtgtcgatgg tagaacgaag cggcgtcgaa gcctgtaaag
     5461 cggcggtgca caatcttctc gcgcaacgcg tcagtgggct gatcattaac tatccgctgg
     5521 atgaccagga tgccattgct gtggaagctg cctgcactaa tgttccggcg ttatttcttg
     5581 atgtctctga ccagacaccc atcaacagta ttattttctc ccatgaagac ggtacgcgac
     5641 tgggcgtgga gcatctggtc gcattgggtc accagcaaat cgcgctgtta gcgggcccat
     5701 taagttctgt ctcggcgcgt ctgcgtctgg ctggctggca taaatatctc actcgcaatc
     5761 aaattcagcc gatagcggaa cgggaa
//
