• 规格：20μl质粒
• 启动子：TEFin
• 复制子：pUC ori
• 终止子：cyc1t
• 质粒分类：大肠杆菌质粒；大肠编辑质粒；大肠Cas9质粒
• 质粒大小：11921bp
• 质粒标签：C-SV40 NLS
• 原核抗性：氨苄青霉素Amp
• 筛选标记：LEU2
• 克隆菌株：大肠杆菌Stbl3
• 表达宿主：解脂耶氏酵母细胞
• 5测序引物：M13R：CAGGAAACAGCTATGACC
• 3测序引物：M13F：TGTAAAACGACGGCCAGT
• 质粒图谱1： png/1-1F926153H0F1.png

质粒序列： 点击查看序列

LOCUS       Exported               11921 bp ds-DNA     circular SYN 29-AUG-2017

DEFINITION  synthetic circular DNA

SOURCE      synthetic DNA construct

  ORGANISM  synthetic DNA construct

REFERENCE   1  (bases 1 to 11921)

  TITLE     Direct Submission

  JOURNAL   Exported Aug 29, 2017 from MiaoLingPlasmid

            http://www.fenghbio.cn

FEATURES             Location/Qualifiers

     source          1..11921

                     /organism="synthetic DNA construct"

                     /mol_type="other DNA"

     misc_feature    414..643

                     /label=Cen1-1

     misc_feature    655..855

                     /label=cyc1t

     CDS             complement(859..879)

                     /codon_start=1

                     /product="nuclear localization signal of SV40 large T 

                     antigen"

                     /label=SV40 NLS

                     /translation="PKKKRKV"

     CDS             complement(892..4992)

                     /codon_start=1

                     /product="Cas9 (Csn1) endonuclease from the Streptococcus 

                     pyogenes Type II CRISPR/Cas system"

                     /label=Cas9

                     /note="generates RNA-guided double strand breaks in DNA"

                     /translation="DKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKN

                     LIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESF

                     LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKF

                     RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE

                     NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI

                     GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ

                     QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR

                     KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN

                     SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF

                     TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS

                     VEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERL

                     KTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQ

                     LIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRH

                     KPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYL

                     YYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVPS

                     EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV

                     AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN

                     AVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTE

                     ITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE

                     SILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT

                     IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNEL

                     ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA

                     NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLD

                     ATLIHQSITGLYETRIDLSQLGGD"

     misc_feature    complement(4993..5523)

                     /label=TEFin

     misc_feature    5530..6030

                     /label=mig1t

     misc_feature    6031..6098

                     /label=HDV

     misc_RNA        complement(6099..6178)

                     /label=gRNA scaffold

                     /note="guide RNA scaffold for the Streptococcus pyogenes 

                     CRISPR/Cas9 system"

     misc_feature    6179..6198

                     /label=trp1.gRNA

     misc_feature    6199..6235

                     /label=HH

     misc_feature    complement(6242..6772)

                     /label=TEFin

     rep_origin      complement(7205..7793)

                     /direction=LEFT

                     /label=ori

                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 

                     replication"

     CDS             complement(7964..8824)

                     /codon_start=1

                     /gene="bla"

                     /product="beta-lactamase"

                     /label=AmpR

                     /note="confers resistance to ampicillin, carbenicillin, and

                     related antibiotics"

                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI

                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS

                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW

                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA

                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS

                     LIKHW"

     promoter        complement(8825..8929)

                     /gene="bla"

                     /label=AmpR promoter

     misc_feature    9165..11903

                     /label=LEU2

ORIGIN

        1 tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat gcagctcccg gagacggtca

       61 cagcttgtct gtaagcggat gccgggagca gacaagcccg tcagggcgcg tcagcgggtg

      121 ttggcgggtg tcggggctgg cttaactatg cggcatcaga gcagattgta ctgagagtgc

      181 accatatgcg gtgtgaaata ccgcacagat gcgtaaggag aaaataccgc atcaggcgcc

      241 attcgccatt caggctgcgc aactgttggg aagggcgatc ggtgcgggcc tcttcgctat

      301 tacgccagct ggcgaaaggg ggatgtgctg caaggcgatt aagttgggta acgccagggt

      361 tttcccagtc acgacgttgt aaaacgacgg ccagtgaatt cgagctcggt acccgctttt

      421 cgtagataat ggaatacaaa tggatatcca gagtatacac atggatagta tacactgaca

      481 cgacaattct gtatctcttt atgttaacta ctgtgaggcg ttaaatagag cttgatatat

      541 aaaatgttac atttcacagt ctgaactttt gcagattacc taatttggta agatattaat

      601 tatgaactga aagttgatgg catccctaaa tttgatgaaa gggggatcac gcgtctgtac

      661 agaaaaaaaa gaaaaatttg aaatataaat aacgttctta atactaacat aactataaaa

      721 aaataaatag ggacctagac ttcaggttgt ctaactcctt ccttttcggt tagagcggat

      781 gtggggggag ggcgtgaatg taagcgtgac ataactaatt acatgactcg aggtccaatg

      841 actcgaggtc caaaatcaca ccttcctctt cttcttgggg tcagccctgc tgtctccacc

      901 gagctgagag aggtcgattc ttgtttcata gagccccgta attgactgat gaatcagtgt

      961 ggcgtccagg acctcctttg tagaggtgta ccgctttctg tctatggtgg tgtcgaagta

     1021 cttgaaggct gcaggcgcgc ccaagttggt cagagtaaac aagtggataa tgttttctgc

     1081 ctgctccctg atgggcttat ccctgtgctt attgtaagca gaaagcacct tatcgaggtt

     1141 agcgtcggcg aggatcactc ttttggagaa ttcgcttatt tgctcgatga tctcatcaag

     1201 gtagtgtttg tgttgttcca cgaacagctg cttctgctca ttatcttcgg gagacccttt

     1261 gagcttttca tagtggctgg ccagatacaa gaaattaacg tatttagagg gcagtgccag

     1321 ctcgttacct ttctgcagct cgcccgcact agcgagcatt cgtttccggc cgttttcaag

     1381 ctcaaagaga gagtacttgg gaagcttaat gatgaggtct tttttgacct ctttatatcc

     1441 tttcgcctcg agaaagtcga tggggttttt ttcgaagctt gatcgctcca tgattgtgat

     1501 gcccagcagt tccttgacgc ttttgagttt tttagacttc cctttctcca ctttggccac

     1561 aaccagtaca ctgtaagcga ctgtaggaga atcgaatccg ccgtatttct tggggtccca

     1621 atcttttttg cgtgcgatca gcttgtcgct gttccttttc gggaggatac tttccttgga

     1681 gaagcctccg gtctgtactt cggtcttttt aacgatgttc acctgcggca tggacaggac

     1741 cttccggact gtcgcgaaat ccctaccctt gtcccacacg atttctcctg tttctccgtt

     1801 tgtttcgata agtggtcgct tccgaatctc tccattggcc agtgtaatct cggtcttgaa

     1861 aaaattcata atattgctgt aaaagaagta cttagcggtg gccttgccta tttcctgctc

     1921 agactttgcg atcattttcc taacatcgta cactttatag tctccgtaaa caaattcaga

     1981 ttcaagcttg ggatattttt tgataagtgc agtgcctacc actgcattca ggtaggcatc

     2041 atgcgcatgg tggtaattgt tgatctctct caccttataa aactgaaagt cctttctgaa

     2101 atctgagacc agcttagact tcagagtaat aactttcacc tctcgaatca gtttgtcatt

     2161 ttcatcgtac ttggtgttca tgcgtgaatc gagaatttgg gccacgtgct tggtgatctg

     2221 gcgtgtctca acaagctgcc ttttgatgaa gccggcttta tccaactcag acaggccacc

     2281 tcgttcagcc ttagtcagat tatcgaactt ccgttgtgtg atcagtttgg cgttcagcag

     2341 ctgccgccaa taatttttca ttttcttgac aacttcttct gaggggacgt tatcactctt

     2401 ccctctattt ttatcggatc ttgtcaacac tttattatca atagaatcat ctttgagaaa

     2461 agactggggc acgatatgat ccacgtcgta gtcggagagc cgattgatgt ccagttcctg

     2521 atccacgtac atgtccctgc cgttctgcag gtagtacagg tagagcttct cattctgaag

     2581 ctgggtgttt tcaactgggt gttccttaag gatttgggac cccagttctt ttataccctc

     2641 ttcaatcctc ttcatccttt ccctactgtt cttctgtccc ttctgggtag tttggttctc

     2701 tcgggccatc tcgataacga tattctcggg cttatgcctt cccattactt tgacgagttc

     2761 atccacgacc ttaacggtct gcagtattcc ctttttgata gctgggctac ctgcaagatt

     2821 agcgatgtgc tcgtgaagac tgtccccctg gccagaaact tgtgctttct ggatgtcctc

     2881 cttaaaggtg agagagtcat catggatcaa ctgcatgaag ttccggttgg caaatccatc

     2941 ggacttaaga aaatccagga ttgtctttcc actctgcttg tctcggatcc cattgatcag

     3001 ttttcttgac agccgccccc atcctgtata tcggcgcctc ttgagctgtt tcatgacttt

     3061 gtcgtcgaag agatgagcgt aagttttcaa gcgttcttca atcatctccc tatcttcaaa

     3121 caacgtaagg gtgaggacaa tgtcctcaag aatgtcctcg ttctcctcat tgtccaggaa

     3181 gtccttgtct ttaatgattt tcaggagatc gtgatacgtt cccagggatg cgttgaagcg

     3241 atcctccact ccgctgattt caacagagtc gaaacattca atctttttga aatagtcttc

     3301 tttgagctgt ttcacggtaa ctttccggtt cgtcttgaag aggaggtcca cgatagcttt

     3361 cttctgctct ccagacagga atgctggctt tctcatccct tctgtgacgt atttgacctt

     3421 ggtgagctcg ttataaactg tgaagtactc gtacagcaga gagtgtttag gaagcacctt

     3481 ttcgttaggc agatttttat caaagttagt catcctttcg atgaaggact gggcagaggc

     3541 ccccttatcc acgacttcct cgaagttcca gggagtgatg gtctcttctg atttgcgagt

     3601 catccacgcg aatctggaat ttccccgggc gagggggcct acatagtagg gtatccgaaa

     3661 tgtgaggatt ttctcaatct tttccctgtt atctttcaaa aaggggtaga aatcctcttg