LOCUS       Exported                4730 bp DNA     circular SYN 21-AUG-2018
DEFINITION  .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4730)
  AUTHORS   Noboru Mizushima
  TITLE     Direct Submission
  JOURNAL   Exported Jun 21, 2022 from SnapGene Viewer 6.0.5
            https://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..4730
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    86..102
                     /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)."
                     /note="color: #31849b"
     primer_bind     210..228
                     /label=pCold Fw
                     /note="color: black; sequence: acgccatatcgccgaaagg"
     primer_bind     complement(245..272)
                     /label=Primer 2
                     /note="color: black; sequence: 
                     cactttgtgattcatggtgtattacctc"
     CDS             273..569
                     /codon_start=1
                     /label=L10K
                     /note="color: #993366"
                     /translation="MAQGQRKFQAHKPAKSKTAAAASEKNRGPRKGGRVIAPKKARVVQ
                     QQKLKKNLEVGIRKKIEHDVVMKASSSLPKKLALLKAPAKKKGAAAATSSKTPS"
     primer_bind     273..294
                     /label=Primer 1
                     /note="color: black; sequence: CACCatggcgcaggggcagcgcaagt"
     primer_bind     complement(541..569)
                     /label=Primer 2
                     /note="color: black; sequence: 
                     tcaggaaggtgtcttggaggaggtggcggcag"
     primer_bind     complement(546..569)
                     /label=Primer 3
                     /note="color: black; sequence: ggaaggtgtcttggaggaggtggc"
     primer_bind     570..631
                     /label=Primer 1
                     /note="color: black; sequence: 
                     GACTACAAAGACGATGACGACAAGcatcatcatcatcatcattaatcgaaggtaggcat
                     atg"
     CDS             570..593
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /note="color: #cc99b2"
                     /translation="DYKDDDDK"
     CDS             594..611
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /note="color: #cc99b2"
                     /translation="HHHHHH"
     CDS             614..625
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /note="color: #cc99b2
                     Cleavage site after base 625"
                     /translation="IEGR"
     primer_bind     complement(709..728)
                     /label=pCold Rv
                     /note="color: black; sequence: ggcagggatcttagattctg"
     rep_origin      complement(1036..1491)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
                     /note="color: #ffff00"
     promoter        1591..1695
                     /gene="bla"
                     /label=AmpR promoter
                     /note="color: #ffffff; direction: RIGHT"
     CDS             1696..2556
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /note="This feature has 2 segments:
                        1: 1696 .. 1764 / #ccffcc / signal sequence
                        2: 1765 .. 2556 / #ccffcc
                     Cleavage site after base 1764"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     rep_origin      2727..3315
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
                     /note="color: #ffff00"
     CDS             complement(3487..4569)
                     /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)."
                     /note="color: #993366"
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     promoter        complement(4570..4647)
                     /gene="lacI"
                     /label=lacI promoter
                     /note="color: #ffffff; direction: LEFT"
ORIGIN
        1 gcaaggaatg gtgtggccga ttaatcataa atatgaaaaa taattgttgc atcacccgcc
       61 aatgcgtggc ttaatgcaca tcaaattgtg agcggataac aatttgatgt gctagcgcat
      121 atccagtgta gtaaggcaag tcccttcaag agttatcgtt gatacccctc gtagtgcaca
      181 ttcctttaac gcttcaaaat ctgtaaagca cgccatatcg ccgaaaggca cacttaatta
      241 ttaagaggta atacaccatg aatcacaaag tgatggcgca ggggcagcgc aagtttcagg
      301 cgcacaaacc cgcaaagagt aagacggcag cggcagcctc tgaaaagaat cggggcccaa
      361 gaaaaggcgg tcgtgttatc gctcccaaga aggcgcgcgt cgtgcagcag caaaagctca
      421 agaagaacct agaagtcgga atccggaaga agatcgaaca tgacgtggtg atgaaagcca
      481 gcagcagcct gcccaagaag ctggcactgc tgaaggcccc agccaagaag aaaggggcag
      541 ctgccgccac ctcctccaag acaccttccg actacaaaga cgatgacgac aagcatcatc
      601 atcatcatca ttaatcgaag gtaggcatat ggagctcggt accctcgagg gatccgaatt
      661 caagcttgtc gacctgcagt ctagataggt aatctctgct taaaagcaca gaatctaaga
      721 tccctgccat ttggcgggga tttttttatt tgttttcagg aaataaataa tcgatcgcgt
      781 aataaaatct attattattt ttgtgaagaa taaatttggg tgcaatgaga atgcgcaggc
      841 cctttcgtct cgcgcgtttc ggtgatgacg gtgaaaacct ctgacacatg cagctcccgg
      901 agacggtcac agcttgtctg taagcggatg ccgggagcag acaagcccgt cagggcgcgt
      961 cagcgggtgt tggcgggtgt cggggctggc ttaactatgc ggcatcagag cagattgtac
     1021 tgagagtgca ccataaaatt gtaaacgtta atattttgtt aaaattcgcg ttaaattttt
     1081 gttaaatcag ctcatttttt aaccaatagg ccgaaatcgg caaaatccct tataaatcaa
     1141 aagaatagcc cgagataggg ttgagtgttg ttccagtttg gaacaagagt ccactattaa
     1201 agaacgtgga ctccaacgtc aaagggcgaa aaaccgtcta tcagggcgat ggcccactac
     1261 gtgaaccatc acccaaatca agttttttgg ggtcgaggtg ccgtaaagca ctaaatcgga
     1321 accctaaagg gagcccccga tttagagctt gacggggaaa gccggcgaac gtggcgagaa
     1381 aggaagggaa gaaagcgaaa ggagcgggcg ctagggcgct ggcaagtgta gcggtcacgc
     1441 tgcgcgtaac caccacaccc gccgcgctta atgcgccgct acagggcgcg tactatggtt
     1501 gctttgacgt atgcggtgtg aaataccgca cagatgcgta aggagaaaat accgcatcag
     1561 gcgtcaggtg gcacttttcg gggaaatgtg cgcggaaccc ctatttgttt atttttctaa
     1621 atacattcaa atatgtatcc gctcatgaga caataaccct gataaatgct tcaataatat
     1681 tgaaaaagga agagtatgag tattcaacat ttccgtgtcg cccttattcc cttttttgcg
     1741 gcattttgcc ttcctgtttt tgctcaccca gaaacgctgg tgaaagtaaa agatgctgaa
     1801 gatcagttgg gtgcacgagt gggttacatc gaactggatc tcaacagcgg taagatcctt
     1861 gagagttttc gccccgaaga acgttttcca atgatgagca cttttaaagt tctgctatgt
     1921 ggcgcggtat tatcccgtat tgacgccggg caagagcaac tcggtcgccg catacactat
     1981 tctcagaatg acttggttga gtactcacca gtcacagaaa agcatcttac ggatggcatg
     2041 acagtaagag aattatgcag tgctgccata accatgagtg ataacactgc ggccaactta
     2101 cttctgacaa cgatcggagg accgaaggag ctaaccgctt ttttgcacaa catgggggat
     2161 catgtaactc gccttgatcg ttgggaaccg gagctgaatg aagccatacc aaacgacgag
     2221 cgtgacacca cgatgcctgt agcaatggca acaacgttgc gcaaactatt aactggcgaa
     2281 ctacttactc tagcttcccg gcaacaatta atagactgga tggaggcgga taaagttgca
     2341 ggaccacttc tgcgctcggc ccttccggct ggctggttta ttgctgataa atctggagcc
     2401 ggtgagcgtg ggtctcgcgg tatcattgca gcactggggc cagatggtaa gccctcccgt
     2461 atcgtagtta tctacacgac ggggagtcag gcaactatgg atgaacgaaa tagacagatc
     2521 gctgagatag gtgcctcact gattaagcat tggtaactgt cagaccaagt ttactcatat
     2581 atactttaga ttgatttaaa acttcatttt taatttaaaa ggatctaggt gaagatcctt
     2641 tttgataatc tcatgaccaa aatcccttaa cgtgagtttt cgttccactg agcgtcagac
     2701 cccgtagaaa agatcaaagg atcttcttga gatccttttt ttctgcgcgt aatctgctgc
     2761 ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt tgccggatca agagctacca
     2821 actctttttc cgaaggtaac tggcttcagc agagcgcaga taccaaatac tgttcttcta
     2881 gtgtagccgt agttaggcca ccacttcaag aactctgtag caccgcctac atacctcgct
     2941 ctgctaatcc tgttaccagt ggctgctgcc agtggcgata agtcgtgtct taccgggttg
     3001 gactcaagac gatagttacc ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc
     3061 acacagccca gcttggagcg aacgacctac accgaactga gatacctaca gcgtgagcta
     3121 tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca ggtatccggt aagcggcagg
     3181 gtcggaacag gagagcgcac gagggagctt ccagggggaa acgcctggta tctttatagt
     3241 cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg
     3301 cggagcctat ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc cttttgctgg
     3361 ccttttgctc acatagtcat gccccgcgcc caccggaagg agctgactgg gttgaaggct
     3421 ctcaagggca tcggtcgaga tcccggtgcc taatgagtga gctaacttac attaattgcg
     3481 ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt gccagctgca ttaatgaatc
     3541 ggccaacgcg cggggagagg cggtttgcgt attgggcgcc agggtggttt ttcttttcac
     3601 cagtgagacg ggcaacagct gattgccctt caccgcctgg ccctgagaga gttgcagcaa
     3661 gcggtccacg ctggtttgcc ccagcaggcg aaaatcctgt ttgatggtgg ttaacggcgg
     3721 gatataacat gagctgtctt cggtatcgtc gtatcccact accgagatat ccgcaccaac
     3781 gcgcagcccg gactcggtaa tggcgcgcat tgcgcccagc gccatctgat cgttggcaac
     3841 cagcatcgca gtgggaacga tgccctcatt cagcatttgc atggtttgtt gaaaaccgga
     3901 catggcactc cagtcgcctt cccgttccgc tatcggctga atttgattgc gagtgagata
     3961 tttatgccag ccagccagac gcagacgcgc cgagacagaa cttaatgggc ccgctaacag
     4021 cgcgatttgc tggtgaccca atgcgaccag atgctccacg cccagtcgcg taccgtcttc
     4081 atgggagaaa ataatactgt tgatgggtgt ctggtcagag acatcaagaa ataacgccgg
     4141 aacattagtg caggcagctt ccacagcaat ggcatcctgg tcatccagcg gatagttaat
     4201 gatcagccca ctgacgcgtt gcgcgagaag attgtgcacc gccgctttac aggcttcgac
     4261 gccgcttcgt tctaccatcg acaccaccac gctggcaccc agttgatcgg cgcgagattt
     4321 aatcgccgcg acaatttgcg acggcgcgtg cagggccaga ctggaggtgg caacgccaat
     4381 cagcaacgac tgtttgcccg ccagttgttg tgccacgcgg ttgggaatgt aattcagctc
     4441 cgccatcgcc gcttccactt tttcccgcgt tttcgcagaa acgtggctgg cctggttcac
     4501 cacgcgggaa acggtctgat aagagacacc ggcatactct gcgacatcgt ataacgttac
     4561 tggtttcaca ttcaccaccc tgaattgact ctcttccggg cgctatcatg ccataccgcg
     4621 aaaggttttg cgccattcga tggtgtccgg gatctcgacg ctctccctta tgcgactcct
     4681 gcattaggaa gcagcccagt agtaggttga ggccgttgag caccgccgcc
//
