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- 详细信息
- 文献和实验
- 技术资料
- 保存条件：
-20
- 保质期：
2年
- 英文名：
Plasmid#63142
- 库存：
100
- 供应商：
上海烜雅生物科技有限公司
- 规格：
干粉/液体
基本信息
名称：pRGEB32植物编辑质粒
别称: Plasmid#63142
质粒属性
质粒简介
pRGEB32质粒由上海烜雅生物科技有限公司提供
质粒图谱
质粒序列
LOCUS Exported 15888 bp ds-DNA circular SYN 24-SEP-2017
DEFINITION synthetic circular DNA
ACCESSION .
VERSION .
KEYWORDS .
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 15888)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported Sep 24, 2017 from MiaoLingPlasmid
http://www.miaolingbio.com
FEATURES Location/Qualifiers
source 1..15888
/organism="synthetic DNA construct"
/mol_type="other DNA"
terminator 71..323
/label=NOS terminator
/note="nopaline synthase terminator and poly(A) signal"
misc_feature 551..575
/label=RB T-DNA repeat
/note="right border repeat from nopaline C58 T-DNA"
CDS 1876..2505
/codon_start=1
/product="stability protein from the Pseudomonas plasmid
pVS1 (Heeb et al., 2000)"
/label=pVS1 StaA
/translation="MKVIAVLNQKGGSGKTTIATHLARALQLAGADVLLVDSDPQGSAR
DWAAVREDQPLTVVGIDRPTIDRDVKAIGRRDFVVIDGAPQAADLAVSAIKAADFVLIP
VQPSPYDIWATADLVELVKQRIEVTDGRLQAAFVVSRAIKGTRIGGEVAEALAGYELPI
LESRITQRVSYPGTAAAGTTVLESEPEGDAAREVQALAAEIKSKLI"
CDS 2934..4007
/codon_start=1
/product="replication protein from the Pseudomonas plasmid
pVS1 (Heeb et al., 2000)"
/label=pVS1 RepA
/translation="MSGRKPSGPVQIGAALGDDLVEKLKAAQAAQRQRIEAEARPGESW
QAAADRIRKESRQPPAAGAPSIRKPPKGDEQPDFFVPMLYDVGTRDSRSIMDVAVFRLS
KRDRRAGEVIRYELPDGHVEVSAGPAGMASVWDYDLVLMAVSHLTESMNRYREGKGDKP
GRVFRPHVADVLKFCRRADGGKQKDDLVETCIRLNTTHVAMQRTKKAKNGRLVTVSEGE
ALISRYKIVKSETGRPEYIEIELADWMYREITEGKNPDVLTVHPDYFLIDPGIGRFLYR
LARRAAGKAEARWLFKTIYERSGSAGEFKKFCFTVRKLIGSNDLPEYDLKEEAGQAGPI
LVMRYRNLIEGEASAGS"
rep_origin 4073..4267
/label=pVS1 oriV
/note="origin of replication for the Pseudomonas plasmid
pVS1 (Heeb et al., 2000)"
misc_feature 4611..4751
/label=bom
/note="basis of mobility region from pBR322"
rep_origin complement(4937..5525)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
CDS complement(5612..6406)
/codon_start=1
/gene="aphA-3"
/product="aminoglycoside phosphotransferase"
/label=KanR
/note="confers resistance to kanamycin"
/translation="MAKMRISPELKKLIEKYRCVKDTEGMSPAKVYKLVGENENLYLKM
TDSRYKGTTYDVEREKDMMLWLEGKLPVPKVLHFERHDGWSNLLMSEADGVLCSEEYED
EQSPEKIIELYAECIRLFHSIDISDCPYTNSLDSRLAELDYLLNNDLADVDCENWEEDT
PFKDPRELYDFLKTEKPEEELVFSHGDLGDSNIFVKDGKVSGFIDLGRSGRADKWYDIA
FCVRSIREDIGEEQYVELFFDLLGIKPDWEKIKYYILLDELF"
misc_feature 6831..6855
/label=LB T-DNA repeat
/note="left border repeat from nopaline C58 T-DNA"
polyA_signal 6933..7107
/label=CaMV poly(A) signal
/note="cauliflower mosaic virus polyadenylation signal"
CDS complement(7148..8173)
/codon_start=1
/gene="aph(4)-Ia"
/product="aminoglycoside phosphotransferase from E. coli"
/label=HygR
/note="confers resistance to hygromycin"
/translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRSQGVTLQDLP
ETELPAVLQPVAEAM-DAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
K"
promoter complement(8240..8916)
/label=CaMV 35S promoter (enhanced)
/note="cauliflower mosaic virus 35S promoter with a
duplicated enhancer region"
promoter 9239..9619
/label=OsU3 promoter
/note="Oryza sativa (rice) snRNA U3 promoter"
misc_RNA 9636..9711
/label=gRNA scaffold
/note="guide RNA scaffold for the Streptococcus pyogenes
CRISPR/Cas9 system"
promoter 9792..11501
/gene="RUBQ2"
/label=Ubi promoter
/note="promoter for the rice (Oryza sativa) polyubiquitin
gene RUBQ2"
CDS 11581..11646
/codon_start=1
/product="three tandem FLAG(R) epitope tags, followed by an
enterokinase cleavage site"
/label=3xFLAG
/translation="DYKDHDGDYKDHDIDYKDDDDK"
CDS 11653..11673
/codon_start=1
/product="nuclear localization signal of SV40 (simian virus
40) large T antigen"
/label=SV40 NLS
/translation="PKKKRKV"
CDS 11698..15798
/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
GDQYADLFLAAKNLS-DAILL-SDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ
QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR
KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN
SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF
TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS
VEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERL
KTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQ
LIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRH
KPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYL
YYLQNGRDMYVDQELDINRLS-DYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVPS
EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV
AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN
AVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTE
ITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE
SILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT
IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNEL
ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA
NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLD
ATLIHQSITGLYETRIDLSQLGGD"
CDS 15799..15846
/codon_start=1
/product="bipartite nuclear localization signal from
nucleoplasmin"
/label=nucleoplasmin NLS
/translation="KRPAATKKAGQAKKKK"
ORIGIN
1 cttgtacaaa gtggttgata acagcgacta caaggatgac gatgacaagg cttagagctc
61 gaatttcccc gatcgttcaa acatttggca ataaagtttc ttaagattga atcctgttgc
121 cggtcttgcg atgattatca tataatttct gttgaattac gttaagcatg taataattaa
181 catgtaatgc atgacgttat ttatgagatg ggtttttatg attagagtcc cgcaattata
241 catttaatac gcgatagaaa acaaaatata gcgcgcaaac taggataaat tatcgcgcgc
301 ggtgtcatct atgttactag atcgggaatt cactggccgt cgttttacaa cgtcgtgact
361 gggaaaaccc tggcgttacc caacttaatc gccttgcagc acatccccct ttcgccagct
421 ggcgtaatag cgaagaggcc cgcaccgatc gcccttccca acagttgcgc agcctgaatg
481 gcgaatgcta gagcagcttg agcttggatc agattgtcgt ttcccgcctt cagtttaaac
541 tatcagtgtt tgacaggata tattggcggg taaacctaag agaaaagagc gtttattaga
601 ataatcggat atttaaaagg gcgtgaaaag gtttatccgt tcgtccattt gtatgtgcat
661 gccaaccaca gggttcccct cgggatcaaa gtactttgat ccaacccctc cgctgctata
721 gtgcagtcgg cttctgacgt tcagtgcagc cgtcttctga aaacgacatg tcgcacaagt
781 cctaagttac gcgacaggct gccgccctgc ccttttcctg gcgttttctt gtcgcgtgtt
841 ttagtcgcat aaagtagaat acttgcgact agaaccggag acattacgcc atgaacaaga
901 gcgccgccgc tggcctgctg ggctatgccc gcgtcagcac cgacgaccag gacttgacca
961 accaacgggc cgaactgcac gcggccggct gcaccaagct gttttccgag aagatcaccg
1021 gcaccaggcg cgaccgcccg gagctggcca ggatgcttga ccacctacgc cctggcgacg
1081 ttgtgacagt gaccaggcta gaccgcctgg cccgcagcac ccgcgaccta ctggacattg
1141 ccgagcgcat ccaggaggcc ggcgcgggcc tgcgtagcct ggcagagccg tgggccgaca
1201 ccaccacgcc ggccggccgc atggtgttga ccgtgttcgc cggcattgcc gagttcgagc
1261 gttccctaat catcgaccgc acccggagcg ggcgcgaggc cgccaaggcc cgaggcgtga
1321 agtttggccc ccgccctacc ctcaccccgg cacagatcgc gcacgcccgc gagctgatcg
1381 accaggaagg ccgcaccgtg aaagaggcgg ctgcactgct tggcgtgcat cgctcgaccc
1441 tgtaccgcgc acttgagcgc agcgaggaag tgacgcccac cgaggccagg cggcgcggtg
1501 ccttccgtga ggacgcattg accgaggccg acgccctggc ggccgccgag aatgaacgcc
1561 aagaggaaca agcatgaaac cgcaccagga cggccaggac gaaccgtttt tcattaccga
1621 agagatcgag gcggagatga tcgcggccgg gtacgtgttc gagccgcccg cgcacgtctc
1681 aaccgtgcgg ctgcatgaaa tcctggccgg tttgtctgat gccaagctgg cggcctggcc
1741 ggccagcttg gccgctgaag aaaccgagcg ccgccgtcta aaaaggtgat gtgtatttga
1801 gtaaaacagc ttgcgtcatg cggtcgctgc gtatatgatg cgatgagtaa ataaacaaat
1861 acgcaagggg aacgcatgaa ggttatcgct gtacttaacc agaaaggcgg gtcaggcaag
1921 acgaccatcg caacccatct agcccgcgcc ctgcaactcg ccggggccga tgttctgtta
1981 gtcgattccg atccccaggg cagtgcccgc gattgggcgg ccgtgcggga agatcaaccg
2041 ctaaccgttg tcggcatcga ccgcccgacg attgaccgcg acgtgaaggc catcggccgg
2101 cgcgacttcg tagtgatcga cggagcgccc caggcggcgg acttggctgt gtccgcgatc
2161 aaggcagccg acttcgtgct gattccggtg cagccaagcc cttacgacat atgggccacc
2221 gccgacctgg tggagctggt taagcagcgc attgaggtca cggatggaag gctacaagcg
2281 gcctttgtcg tgtcgcgggc gatcaaaggc acgcgcatcg gcggtgaggt tgccgaggcg
2341 ctggccgggt acgagctgcc cattcttgag tcccgtatca cgcagcgcgt gagctaccca
2401 ggcactgccg ccgccggcac aaccgttctt gaatcagaac ccgagggcga cgctgcccgc
2461 gaggtccagg cgctggccgc tgaaattaaa tcaaaactca tttgagttaa tgaggtaaag
2521 agaaaatgag caaaagcaca aacacgctaa gtgccggccg tccgagcgca cgcagcagca
2581 aggctgcaac gttggccagc ctggcagaca cgccagccat gaagcgggtc aactttcagt
2641 tgccggcgga ggatcacacc aagctgaaga tgtacgcggt acgccaaggc aagaccatta
2701 ccgagctgct atctgaatac atcgcgcagc taccagagta aatgagcaaa tgaataaatg
2761 agtagatgaa ttttagcggc taaaggaggc ggcatggaaa atcaagaaca accaggcacc
2821 gacgccgtgg aatgccccat gtgttgagga acgggcggtt ggccaggcgt aagcggctgg
2881 gttgcctgcc ggccctgcaa tggcactgga acccccaagc ccgaggaatc ggcgtgagcg
2941 gtcgcaaacc atccggcccg gtacaaatcg gcgcggcgct gggtgatgac ctggtggaga
3001 agttgaaggc cgcgcaggcc gcccagcggc aacgcatcga ggcagaagca cgccccggtg
3061 aatcgtggca agcggccgct gatcgaatcc gcaaagaatc ccggcaaccg ccggcagccg
3121 gtgcgccgtc gattaggaag ccgcccaagg gcgacgagca accagatttt ttcgttccga
3181 tgctctatga cgtgggcacc cgcgatagtc gcagcatcat ggacgtggcc gttttccgtc
3241 tgtcgaagcg tgaccgacga gctggcgagg tgatccgcta cgagcttcca gacgggcacg
3301 tagaggtttc cgcagggccg gccggcatgg ccagtgtgtg ggattacgac ctggtactga
3361 tggcggtttc ccatctaacc gaatccatga accgataccg ggaagggaag ggagacaagc
3421 ccggccgcgt gttccgtcca cacgttgcgg acgtactcaa gttctgccgg cgagccgatg
3481 gcggaaagca gaaagacgac ctggtagaaa cctgcattcg gttaaacacc acgcacgttg
3541 ccatgcagcg tacgaagaag gccaagaacg gccgcctggt gacggtatcc gagggtgaag
3601 ccttgattag ccgctacaag atcgtaaaga gcgaaaccgg gcggccggag tacatcgaga
3661 tcgagctagc tgattggatg taccgcgaga tcacagaagg caagaacccg gacgtgctga
3721 cggttcaccc cgattacttt ttgatcgatc ccggcatcgg ccgttttctc taccgcctgg
3781 cacgccgcgc cgcaggcaag gcagaagcca gatggttgtt caagacgatc tacgaacgca
3841 gtggcagcgc cggagagttc aagaagttct gtttcaccgt gcgcaagctg atcgggtcaa
3901 atgacctgcc ggagtacgat ttgaaggagg aggcggggca ggctggcccg atcctagtca
3961 tgcgctaccg caacctgatc gagggcgaag catccgccgg ttcctaatgt acggagcaga
4021 tgctagggca aattgcccta gcaggggaaa aaggtcgaaa agcactcttt cctgtgaata
4081 gcacgtacat tgggaaccca aagccgtaca ttgggaaccg gaacccgtac attgggaacc
4141 caaagccgta cattgggaac cggtcacaca tgtaagtgac tgatataaaa gagaaaaaag
4201 gcgatttttc cgcctaaaac tctttaaaac ttattaaaac tcttaaaacc cgcctggcct
4261 gtgcataact gtctggccag cgcacagccg aagagctgca aaaagcgcct acccttcggt
4321 cgctgcgctc cctacgcccc gccgcttcgc gtcggcctat cgcggccgct ggccgctcaa
4381 aaatggctgg cctacggcca ggcaatctac cagggcgcgg acaagccgcg ccgtcgccac
4441 tcgaccgccg gcgcccacat caaggcaccc tgcctcgcgc gtttcggtga tgacggtgaa
4501 aacctctgac acatgcagct cccggagacg gtcacagctt gtctgtaagc ggatgccggg
4561 agcagacaag cccgtcaggg cgcgtcagcg ggtgttggcg ggtgtcgggg cgcagccatg
4621 acccagtcac gtagcgatag cggagtgtat actggcttaa ctatgcggca tcagagcaga
4681 ttgtactgag agtgcaccat atgcggtgtg aaataccgca cagatgcgta aggagaaaat
4741 accgcatcag gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc
4801 tgcggcgagc ggtatcagct cactcaaagg cggtaatacg gttatccaca gaatcagggg
4861 ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg
4921 ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac aaaaatcgac
4981 gctcaagtca gaggtggcga aacccgacag gactataaag ataccaggcg tttccccctg
5041 gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac ctgtccgcct
5101 ttctcccttc gggaagcgtg gcgctttctc atagctcacg ctgtaggtat ctcagttcgg
5161 tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct
5221 gcgccttatc cggtaactat cgtcttgagt ccaacccggt aagacacgac ttatcgccac
5281 tggcagcagc cactggtaac aggattagca gagcgaggta tgtaggcggt gctacagagt
5341 tcttgaagtg gtggcctaac tacggctaca ctagaaggac agtatttggt atctgcgctc
5401 tgctgaagcc agttaccttc ggaaaaagag ttggtagctc ttgatccggc aaacaaacca
5461 ccgctggtag cggtggtttt tttgtttgca agcagcagat tacgcgcaga aaaaaaggat
5521 ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc tcagtggaac gaaaactcac
5581 gttaagggat tttggtcatg cattctaggt actaaaacaa ttcatccagt aaaatataat
5641 attttatttt ctcccaatca ggcttgatcc ccagtaagtc aaaaaatagc tcgacatact
5701 gttcttcccc gatatcctcc ctgatcgacc ggacgcagaa ggcaatgtca taccacttgt
5761 ccgccctgcc gcttctccca agatcaataa agccacttac tttgccatct ttcacaaaga
5821 tgttgctgtc tcccaggtcg ccgtgggaaa agacaagttc ctcttcgggc ttttccgtct
5881 ttaaaaaatc atacagctcg cgcggatctt taaatggagt gtcttcttcc cagttttcgc
5941 aatccacatc ggccagatcg ttattcagta agtaatccaa ttcggctaag cggctgtcta
6001 agctattcgt atagggacaa tccgatatgt cgatggagtg aaagagcctg atgcactccg
6061 catacagctc gataatcttt tcagggcttt gttcatcttc atactcttcc gagcaaagga
6121 cgccatcggc ctcactcatg agcagattgc tccagccatc atgccgttca aagtgcagga
6181 cctttggaac aggcagcttt ccttccagcc atagcatcat gtccttttcc cgttccacat
6241 cataggtggt ccctttatac cggctgtccg tcatttttaa atataggttt tcattttctc
6301 ccaccagctt atatacctta gcaggagaca ttccttccgt atcttttacg cagcggtatt
6361 tttcgatcag ttttttcaat tccggtgata ttctcatttt agccatttat tatttccttc
6421 ctcttttcta cagtatttaa agatacccca agaagctaat tataacaaga cgaactccaa
6481 ttcactgttc cttgcattct aaaaccttaa ataccagaaa acagcttttt caaagttgtt
6541 ttcaaagttg gcgtataaca tagtatcgac ggagccgatt ttgaaaccgc ggtgatcaca
6601 ggcagcaacg ctctgtcatc gttacaatca acatgctacc ctccgcgaga tcatccgtgt
6661 ttcaaacccg gcagcttagt tgccgttctt ccgaatagca tcggtaacat gagcaaagtc
6721 tgccgcctta caacggctct cccgctgacg ccgtcccgga ctgatgggct gcctgtatcg
6781 agtggtgatt ttgtgccgag ctgccggtcg gggagctgtt ggctggctgg tggcaggata
6841 tattgtggtg taaacaaatt gacgcttaga caacttaata acacattgcg gacgttttta
6901 atgtactgaa ttaacgccga attaattcgg gggatctgga ttttagtact ggattttggt
6961 tttaggaatt agaaatttta ttgatagaag tattttacaa atacaaatac atactaaggg
7021 tttcttatat gctcaacaca tgagcgaaac cctataggaa ccctaattcc cttatctggg
7081 aactactcac acattattat ggagaaactc gagcttgtcg atcgacagat cccggtcggc
7141 atctactcta tttctttgcc ctcggacgag tgctggggcg tcggtttcca ctatcggcga
7201 gtacttctac acagccatcg gtccagacgg ccgcgcttct gcgggcgatt tgtgtacgcc
7261 cgacagtccc ggctccggat cggacgattg cgtcgcatcg accctgcgcc caagctgcat
7321 catcgaaatt gccgtcaacc aagctctgat agagttggtc aagaccaatg cggagcatat
7381 acgcccggag tcgtggcgat cctgcaagct ccggatgcct ccgctcgaag tagcgcgtct
7441 gctgctccat acaagccaac cacggcctcc agaagaagat gttggcgacc tcgtattggg
7501 aatccccgaa catcgcctcg ctccagtcaa tgaccgctgt tatgcggcca ttgtccgtca
7561 ggacattgtt ggagccgaaa tccgcgtgca cgaggtgccg gacttcgggg cagtcctcgg
7621 cccaaagcat cagctcatcg agagcctgcg cgacggacgc actgacggtg tcgtccatca
7681 cagtttgcca gtgatacaca tggggatcag caatcgcgca tatgaaatca cgccatgtag
7741 tgtattgacc gattccttgc ggtccgaatg ggccgaaccc gctcgtctgg ctaagatcgg
7801 ccgcagcgat cgcatccata gcctccgcga ccggttgtag aacagcgggc agttcggttt
7861 caggcaggtc ttgcaacgtg acaccctgtg aacggcggga gatgcaatag gtcaggctct
7921 cgctaaactc cccaatgtca agcacttccg gaatcgggag cgcggccgat gcaaagtgcc
7981 gataaacata acgatctttg tagaaaccat cggcgcagct atttacccgc aggacatatc
8041 cacgccctcc tacatcgaag ctgaaagcac gagattcttc gccctccgag agctgcatca
8101 ggtcggagac gctgtcgaac ttttcgatca gaaacttctc gacagacgtc gcggtgagtt
8161 caggcttttt catatctcat tgccccccgg atctgcgaaa gctcgagaga gatagatttg
8221 tagagagaga ctggtgattt cagcgtgtcc tctccaaatg aaatgaactt ccttatatag
8281 aggaagggtc ttgcgaagga tagtgggatt gtgcgtcatc ccttacgtca gtggagatat
8341 cacatcaatc cacttgcttt gaagacgtgg ttggaacgtc ttctttttcc acgatgctcc
8401 tcgtgggtgg gggtccatct ttgggaccac tgtcggcaga ggcatcttga acgatagcct
8461 ttcctttatc gcaatgatgg catttgtagg tgccaccttc cttttctact gtccttttga
8521 tgaagtgaca gatagctggg caatggaatc cgaggaggtt tcccgatatt accctttgtt
8581 gaaaagtctc aatagccctt tggtcttctg agactgtatc tttgatattc ttggagtaga
8641 cgagagtgtc gtgctccacc atgttcacat caatccactt gctttgaaga cgtggttgga
8701 acgtcttctt tttccacgat gctcctcgtg ggtgggggtc catctttggg accactgtcg
8761 gcagaggcat cttgaacgat agcctttcct ttatcgcaat gatggcattt gtaggtgcca
8821 ccttcctttt ctactgtcct tttgatgaag tgacagatag ctgggcaatg gaatccgagg
8881 aggtttcccg atattaccct ttgttgaaaa gtctcaatag ccctttggtc ttctgagact
8941 gtatctttga tattcttgga gtagacgaga gtgtcgtgct ccaccatgtt ggcaagctgc
9001 tctagccaat acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagctggc
9061 acgacaggtt tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat gtgagttagc
9121 tcactcatta ggcaccccag gctttacact ttatgcttcc ggctcgtatg ttgtgtggaa
9181 ttgtgagcgg ataacaattt cacacaggaa acagctatga catgattacg ccaagcttaa
9241 ggaatcttta aacatacgaa cagatcactt aaagttcttc tgaagcaact taaagttatc
9301 aggcatgcat ggatcttgga ggaatcagat gtgcagtcag ggaccatagc acaagacagg
9361 cgtcttctac tggtgctacc agcaaatgct ggaagccggg aacactgggt acgttggaaa
9421 ccacgtgatg tgaagaagta agataaactg taggagaaaa gcatttcgta gtgggccatg
9481 aagcctttca ggacatgtat tgcagtatgg gccggcccat tacgcaattg gacgacaaca
9541 aagactagta ttagtaccac ctcggctatc cacatagatc aaagctgatt taaaagagtt
9601 gtgcagatga tccgtggcag gagaccgagg tctcggtttt agagctagaa atagcaagtt
9661 aaaataaggc tagtccgtta tcaacttgaa aaagtggcac cgagtcggtg cttttttgtt
9721 ttagagctag aaatagcaag ttaaaataag gctagtccgt ttttagcgcg tgcatgcctg
9781 caggtccaca aattcgggtc aaggcggaag ccagcgcgcc accccacgtc agcaaatacg
9841 gaggcgcggg gttgacggcg tcacccggtc ctaacggcga ccaacaaacc agccagaaga
9901 aattacagta aaaaaaaagt aaattgcact ttgatccacc ttttattacc taagtctcaa
9961 tttggatcac ccttaaacct atcttttcaa tttgggccgg gttgtggttt ggactaccat
10021 gaacaacttt tcgtcatgtc taacttccct ttcagcaaac atatgaacca tatatagagg
10081 agatcggccg tatactagag ctgatgtgtt taaggtcgtt gattgcacga gaaaaaaaaa
10141 tccaaatcgc aacaatagca aatttatctg gttcaaagtg aaaagatatg tttaaaggta
10201 gtccaaagta aaacttatag ataataaaat gtggtccaaa gcgtaattca ctcaaaaaaa
10261 atcaacgaga cgtgtaccaa acggagacaa acggcatctt ctcgaaattt cccaaccgct
10321 cgctcgcccg cctcgtcttc ccggaaaccg cggtggtttc agcgtggcgg attctccaag
10381 cagacggaga cgtcacggca cgggactcct cccaccaccc aaccgccata aataccagcc
10441 ccctcatctc ctctcctcgc atcagctcca cccccgaaaa atttctcccc aatctcgcga
10501 ggctctcgtc gtcgaatcga atcctctcgc gtcctcaagg tacgctgctt ctcctctcct
10561 cgcttcgttt cgattcgatt tcggacgggt gaggttgttt tgttgctaga tccgattggt
10621 ggttagggtt gtcgatgtga ttatcgtgag atgtttaggg gttgtagatc tgatggttgt
10681 gatttgggca cggttggttc gataggtgga atcgtggtta ggttttggga ttggatgttg
10741 gttctgatga ttggggggaa tttttacggt tagatgaatt gttggatgat tcgattgggg
10801 aaatcggtgt agatctgttg gggaattgtg gaactagtca tgcctgagtg attggtgcga
10861 tttgtagcgt gttccatctt gtaggccttg ttgcgagcat gttcagatct actgttccgc
10921 tcttgattga gttattggtg ccatgggttg gtgcaaacac aggctttaat atgttatatc
10981 tgttttgtgt ttgatgtaga tctgtagggt agttcttctt agacatggtt caattatgta
11041 gcttgtgcgt ttcgatttga tttcatatgt tcacagatta gataatgatg aactctttta
11101 attaattgtc aatggtaaat aggaagtctt gtcgctatat ctgtcataat gatctcatgt
11161 tactatctgc cagtaattta tgctaagaac tatattagaa tatcatgtta caatctgtag
11221 taatatcatg ttacaatctg tagttcatct atataatcta ttgtggtaat ttctttttac
11281 tatctgtgtg aagattattg ccactagttc attctactta tttctgaagt tcaggatacg
11341 tgtgctgtta ctacctatct gaatacatgt gtgatgtgcc tgttactatc tttttgaata
11401 catgtatgtt ctgttggaat atgtttgctg tttgatccgt tgttgtgtcc ttaatcttgt
11461 gctagttctt accctatctg tttggtgatt atttcttgca gatagttatc aacaagtttg
11521 tacaaaaaag caggcttcga aggagataga accaattctc taaggaaata cttaaccatg
11581 gactataagg accacgacgg agactacaag gatcatgata ttgattacaa agacgatgac
11641 gataagatgg ccccaaagaa gaagcggaag gtcggtatcc acggagtccc agcagccgac
11701 aagaagtaca gcatcggcct ggacatcggc accaactctg tgggctgggc cgtgatcacc
11761 gacgagtaca aggtgcccag caagaaattc aaggtgctgg gcaacaccga ccggcacagc
11821 atcaagaaga acctgatcgg agccctgctg ttcgacagcg gcgaaacagc cgaggccacc
11881 cggctgaaga gaaccgccag aagaagatac accagacgga agaaccggat ctgctatctg
11941 caagagatct tcagcaacga gatggccaag gtggacgaca gcttcttcca cagactggaa
12001 gagtccttcc tggtggaaga ggataagaag cacgagcggc accccatctt cggcaacatc
12061 gtggacgagg tggcctacca cgagaagtac cccaccatct accacctgag aaagaaactg
12121 gtggacagca ccgacaaggc cgacctgcgg ctgatctatc tggccctggc ccacatgatc
12181 aagttccggg gccacttcct gatcgagggc gacctgaacc ccgacaacag cgacgtggac
12241 aagctgttca tccagctggt gcagacctac aaccagctgt tcgaggaaaa ccccatcaac
12301 gccagcggcg tggacgccaa ggccatcctg tctgccagac tgagcaagag cagacggctg
12361 gaaaatctga tcgcccagct gcccggcgag aagaagaatg gcctgttcgg aaacctgatt
12421 gccctgagcc tgggcctgac ccccaacttc aagagcaact tcgacctggc cgaggatgcc
12481 aaactgcagc tgagcaagga cacctacgac gacgacctgg acaacctgct ggcccagatc
12541 ggcgaccagt acgccgacct gtttctggcc gccaagaacc tgtccgacgc catcctgctg
12601 agcgacatcc tgagagtgaa caccgagatc accaaggccc ccctgagcgc ctctatgatc
12661 aagagatacg acgagcacca ccaggacctg accctgctga aagctctcgt gcggcagcag
12721 ctgcctgaga agtacaaaga gattttcttc gaccagagca agaacggcta cgccggctac
12781 attgacggcg gagccagcca ggaagagttc tacaagttca tcaagcccat cctggaaaag
12841 atggacggca ccgaggaact gctcgtgaag ctgaacagag aggacctgct gcggaagcag
12901 cggaccttcg acaacggcag catcccccac cagatccacc tgggagagct gcacgccatt
12961 ctgcggcggc aggaagattt ttacccattc ctgaaggaca accgggaaaa gatcgagaag
13021 atcctgacct tccgcatccc ctactacgtg ggccctctgg ccaggggaaa cagcagattc
13081 gcctggatga ccagaaagag cgaggaaacc atcaccccct ggaacttcga ggaagtggtg
13141 gacaagggcg cttccgccca gagcttcatc gagcggatga ccaacttcga taagaacctg
13201 cccaacgaga aggtgctgcc caagcacagc ctgctgtacg agtacttcac cgtgtataac
13261 gagctgacca aagtgaaata cgtgaccgag ggaatgagaa agcccgcctt cctgagcggc
13321 gagcagaaaa aggccatcgt ggacctgctg ttcaagacca accggaaagt gaccgtgaag
13381 cagctgaaag aggactactt caagaaaatc gagtgcttcg actccgtgga aatctccggc
13441 gtggaagatc ggttcaacgc ctccctgggc acataccacg atctgctgaa aattatcaag
13501 gacaaggact tcctggacaa tgaggaaaac gaggacattc tggaagatat cgtgctgacc
13561 ctgacactgt ttgaggacag agagatgatc gaggaacggc tgaaaaccta tgcccacctg
13621 ttcgacgaca aagtgatgaa gcagctgaag cggcggagat acaccggctg gggcaggctg
13681 agccggaagc tgatcaacgg catccgggac aagcagtccg gcaagacaat cctggatttc
13741 ctgaagtccg acggcttcgc caacagaaac ttcatgcagc tgatccacga cgacagcctg
13801 acctttaaag aggacatcca gaaagcccag gtgtccggcc agggcgatag cctgcacgag
13861 cacattgcca atctggccgg cagccccgcc attaagaagg gcatcctgca gacagtgaag
13921 gtggtggacg agctcgtgaa agtgatgggc cggcacaagc ccgagaacat cgtgatcgaa
13981 atggccagag agaaccagac cacccagaag ggacagaaga acagccgcga gagaatgaag
14041 cggatcgaag agggcatcaa agagctgggc agccagatcc tgaaagaaca ccccgtggaa
14101 aacacccagc tgcagaacga gaagctgtac ctgtactacc tgcagaatgg gcgggatatg
14161 tacgtggacc aggaactgga catcaaccgg ctgtccgact acgatgtgga ccatatcgtg
14221 cctcagagct ttctgaagga cgactccatc gacaacaagg tgctgaccag aagcgacaag
14281 aaccggggca agagcgacaa cgtgccctcc gaagaggtcg tgaagaagat gaagaactac
14341 tggcggcagc tgctgaacgc caagctgatt acccagagaa agttcgacaa tctgaccaag
14401 gccgagagag gcggcctgag cgaactggat aaggccggct tcatcaagag acagctggtg
14461 gaaacccggc agatcacaaa gcacgtggca cagatcctgg actcccggat gaacactaag
14521 tacgacgaga atgacaagct gatccgggaa gtgaaagtga tcaccctgaa gtccaagctg
14581 gtgtccgatt tccggaagga tttccagttt tacaaagtgc gcgagatcaa caactaccac
14641 cacgcccacg acgcctacct gaacgccgtc gtgggaaccg ccctgatcaa aaagtaccct
14701 aagctggaaa gcgagttcgt gtacggcgac tacaaggtgt acgacgtgcg gaagatgatc
14761 gccaagagcg agcaggaaat cggcaaggct accgccaagt acttcttcta cagcaacatc
14821 atgaactttt tcaagaccga gattaccctg gccaacggcg agatccggaa gcggcctctg
14881 atcgagacaa acggcgaaac cggggagatc gtgtgggata agggccggga ttttgccacc
14941 gtgcggaaag tgctgagcat gccccaagtg aatatcgtga aaaagaccga ggtgcagaca
15001 ggcggcttca gcaaagagtc tatcctgccc aagaggaaca gcgataagct gatcgccaga
15061 aagaaggact gggaccctaa gaagtacggc ggcttcgaca gccccaccgt ggcctattct
15121 gtgctggtgg tggccaaagt ggaaaagggc aagtccaaga aactgaagag tgtgaaagag
15181 ctgctgggga tcaccatcat ggaaagaagc agcttcgaga agaatcccat cgactttctg
15241 gaagccaagg gctacaaaga agtgaaaaag gacctgatca tcaagctgcc taagtactcc
15301 ctgttcgagc tggaaaacgg ccggaagaga atgctggcct ctgccggcga actgcagaag
15361 ggaaacgaac tggccctgcc ctccaaatat gtgaacttcc tgtacctggc cagccactat
15421 gagaagctga agggctcccc cgaggataat gagcagaaac agctgtttgt ggaacagcac
15481 aagcactacc tggacgagat catcgagcag atcagcgagt tctccaagag agtgatcctg
15541 gccgacgcta atctggacaa agtgctgtcc gcctacaaca agcaccggga taagcccatc
15601 agagagcagg ccgagaatat catccacctg tttaccctga ccaatctggg agcccctgcc
15661 gccttcaagt actttgacac caccatcgac cggaagaggt acaccagcac caaagaggtg
15721 ctggacgcca ccctgatcca ccagagcatc accggcctgt acgagacacg gatcgacctg
15781 tctcagctgg gaggcgacaa aaggccggcg gccacgaaaa aggccggcca ggcaaaaaag
15841 aaaaagtaag aattcgcggc cgcactcgag atatctagac ccagcttt
//
质粒菌株产品操作说明书
一、扩增流程
收到产品后，请先根据产品管壁标签来判断产品形式，并在扩增前准确查找该质粒菌株的抗性、感受态和培养温度。
1、质粒干粉（常温运输，存于-20度，90天保质期，请务必转化挑单克隆培养，不要直接使用和测序）
①收到质粒干粉后请先5000rpm离心1min，再加入20μl ddH2O去离子水溶解质粒；
②取1支100μl 感受态于冰上解冻10min，加入2μl质粒，再冰浴30min后，42℃热激60s，不要搅动，再冰浴2min；（从第二步开始均要在超净工作台中无菌操作）
③加入900μl无抗的LB液体培养基，180rpm震荡37℃培养45min；
④6000rpm离心5min，仅留100μl上清液重悬细菌沉淀，并涂布至目标质粒抗性的LB平板上；（可使用本平台的平板涂布专用玻璃珠进行涂布，可以比传统涂布方法获得更多转化子）
⑤将平板正向培养1h，再倒置37℃培养14h。如果要求是30度则培养20h；
（菌落过多则将质粒稀释后再转化。没有菌落则加入10μl质粒转化。另不要直接转表达感受态，要先转克隆感受态，重提质粒后再导入表达感受态）
⑥挑取单菌落至LB液体培养基中，加入对应抗生素，220rpm震荡培养14h，根据实验需要和质粒提取试剂盒说明书提取质粒。
2、甘油菌种（冰袋运输，存于-80℃，保质期90天，请务必划线挑单克隆培养）
四区划线后挑单菌落培养，酵母菌需要先液体复苏再四区划线，再挑单菌落液体培养。
3、穿刺菌种（冰袋运输，存于4℃，保质期7天）
穿刺接种，液体培养后四区划线，再挑单菌落液体培养。
4、菌落平板（冰袋运输，存于4℃，保质期7天）
直接挑取单菌落至液体培养基中。
5、液体质粒（冰袋运输，存于-20℃，保质期90天）
单独提取的液体质粒收到后可直接使用。
6、滤纸质粒（常温运输，存于-20度，90天保质期，请务必转化挑单克隆培养，不要直接使用和测序）
收到货后将滤纸画圈部分剪下放入EP管中，加100ul无菌水将滤纸浸湿并浸泡5min，吸取5ul质粒转化，离心全涂。
二、转化图片
名称：pRGEB32植物编辑质粒
别称: Plasmid#63142
| 启动子: | Ubi,Osu3 |
|---|---|
| 复制子: | pVS1 |
| 原核抗性: | Kan |
| 筛选标记: | Hyg |
| 克隆菌株: | Stbl3 |
| 培养条件: | 37度 |
质粒属性
| 载体宿主： | 植物 |
|---|---|
| 载体用途： | 基因敲除 |
| 基因种属： | 空载体 |
| 基因类型： | CRISPR |
| 原核抗性： | Kan |
| 筛选标记： | Hyg |
| 荧光蛋白： |
质粒简介
pRGEB32质粒由上海烜雅生物科技有限公司提供
质粒图谱
质粒序列
LOCUS Exported 15888 bp ds-DNA circular SYN 24-SEP-2017
DEFINITION synthetic circular DNA
ACCESSION .
VERSION .
KEYWORDS .
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 15888)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported Sep 24, 2017 from MiaoLingPlasmid
http://www.miaolingbio.com
FEATURES Location/Qualifiers
source 1..15888
/organism="synthetic DNA construct"
/mol_type="other DNA"
terminator 71..323
/label=NOS terminator
/note="nopaline synthase terminator and poly(A) signal"
misc_feature 551..575
/label=RB T-DNA repeat
/note="right border repeat from nopaline C58 T-DNA"
CDS 1876..2505
/codon_start=1
/product="stability protein from the Pseudomonas plasmid
pVS1 (Heeb et al., 2000)"
/label=pVS1 StaA
/translation="MKVIAVLNQKGGSGKTTIATHLARALQLAGADVLLVDSDPQGSAR
DWAAVREDQPLTVVGIDRPTIDRDVKAIGRRDFVVIDGAPQAADLAVSAIKAADFVLIP
VQPSPYDIWATADLVELVKQRIEVTDGRLQAAFVVSRAIKGTRIGGEVAEALAGYELPI
LESRITQRVSYPGTAAAGTTVLESEPEGDAAREVQALAAEIKSKLI"
CDS 2934..4007
/codon_start=1
/product="replication protein from the Pseudomonas plasmid
pVS1 (Heeb et al., 2000)"
/label=pVS1 RepA
/translation="MSGRKPSGPVQIGAALGDDLVEKLKAAQAAQRQRIEAEARPGESW
QAAADRIRKESRQPPAAGAPSIRKPPKGDEQPDFFVPMLYDVGTRDSRSIMDVAVFRLS
KRDRRAGEVIRYELPDGHVEVSAGPAGMASVWDYDLVLMAVSHLTESMNRYREGKGDKP
GRVFRPHVADVLKFCRRADGGKQKDDLVETCIRLNTTHVAMQRTKKAKNGRLVTVSEGE
ALISRYKIVKSETGRPEYIEIELADWMYREITEGKNPDVLTVHPDYFLIDPGIGRFLYR
LARRAAGKAEARWLFKTIYERSGSAGEFKKFCFTVRKLIGSNDLPEYDLKEEAGQAGPI
LVMRYRNLIEGEASAGS"
rep_origin 4073..4267
/label=pVS1 oriV
/note="origin of replication for the Pseudomonas plasmid
pVS1 (Heeb et al., 2000)"
misc_feature 4611..4751
/label=bom
/note="basis of mobility region from pBR322"
rep_origin complement(4937..5525)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
CDS complement(5612..6406)
/codon_start=1
/gene="aphA-3"
/product="aminoglycoside phosphotransferase"
/label=KanR
/note="confers resistance to kanamycin"
/translation="MAKMRISPELKKLIEKYRCVKDTEGMSPAKVYKLVGENENLYLKM
TDSRYKGTTYDVEREKDMMLWLEGKLPVPKVLHFERHDGWSNLLMSEADGVLCSEEYED
EQSPEKIIELYAECIRLFHSIDISDCPYTNSLDSRLAELDYLLNNDLADVDCENWEEDT
PFKDPRELYDFLKTEKPEEELVFSHGDLGDSNIFVKDGKVSGFIDLGRSGRADKWYDIA
FCVRSIREDIGEEQYVELFFDLLGIKPDWEKIKYYILLDELF"
misc_feature 6831..6855
/label=LB T-DNA repeat
/note="left border repeat from nopaline C58 T-DNA"
polyA_signal 6933..7107
/label=CaMV poly(A) signal
/note="cauliflower mosaic virus polyadenylation signal"
CDS complement(7148..8173)
/codon_start=1
/gene="aph(4)-Ia"
/product="aminoglycoside phosphotransferase from E. coli"
/label=HygR
/note="confers resistance to hygromycin"
/translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRSQGVTLQDLP
ETELPAVLQPVAEAM-DAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
K"
promoter complement(8240..8916)
/label=CaMV 35S promoter (enhanced)
/note="cauliflower mosaic virus 35S promoter with a
duplicated enhancer region"
promoter 9239..9619
/label=OsU3 promoter
/note="Oryza sativa (rice) snRNA U3 promoter"
misc_RNA 9636..9711
/label=gRNA scaffold
/note="guide RNA scaffold for the Streptococcus pyogenes
CRISPR/Cas9 system"
promoter 9792..11501
/gene="RUBQ2"
/label=Ubi promoter
/note="promoter for the rice (Oryza sativa) polyubiquitin
gene RUBQ2"
CDS 11581..11646
/codon_start=1
/product="three tandem FLAG(R) epitope tags, followed by an
enterokinase cleavage site"
/label=3xFLAG
/translation="DYKDHDGDYKDHDIDYKDDDDK"
CDS 11653..11673
/codon_start=1
/product="nuclear localization signal of SV40 (simian virus
40) large T antigen"
/label=SV40 NLS
/translation="PKKKRKV"
CDS 11698..15798
/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
GDQYADLFLAAKNLS-DAILL-SDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ
QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR
KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN
SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF
TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS
VEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERL
KTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQ
LIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRH
KPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYL
YYLQNGRDMYVDQELDINRLS-DYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVPS
EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV
AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN
AVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTE
ITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE
SILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT
IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNEL
ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA
NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLD
ATLIHQSITGLYETRIDLSQLGGD"
CDS 15799..15846
/codon_start=1
/product="bipartite nuclear localization signal from
nucleoplasmin"
/label=nucleoplasmin NLS
/translation="KRPAATKKAGQAKKKK"
ORIGIN
1 cttgtacaaa gtggttgata acagcgacta caaggatgac gatgacaagg cttagagctc
61 gaatttcccc gatcgttcaa acatttggca ataaagtttc ttaagattga atcctgttgc
121 cggtcttgcg atgattatca tataatttct gttgaattac gttaagcatg taataattaa
181 catgtaatgc atgacgttat ttatgagatg ggtttttatg attagagtcc cgcaattata
241 catttaatac gcgatagaaa acaaaatata gcgcgcaaac taggataaat tatcgcgcgc
301 ggtgtcatct atgttactag atcgggaatt cactggccgt cgttttacaa cgtcgtgact
361 gggaaaaccc tggcgttacc caacttaatc gccttgcagc acatccccct ttcgccagct
421 ggcgtaatag cgaagaggcc cgcaccgatc gcccttccca acagttgcgc agcctgaatg
481 gcgaatgcta gagcagcttg agcttggatc agattgtcgt ttcccgcctt cagtttaaac
541 tatcagtgtt tgacaggata tattggcggg taaacctaag agaaaagagc gtttattaga
601 ataatcggat atttaaaagg gcgtgaaaag gtttatccgt tcgtccattt gtatgtgcat
661 gccaaccaca gggttcccct cgggatcaaa gtactttgat ccaacccctc cgctgctata
721 gtgcagtcgg cttctgacgt tcagtgcagc cgtcttctga aaacgacatg tcgcacaagt
781 cctaagttac gcgacaggct gccgccctgc ccttttcctg gcgttttctt gtcgcgtgtt
841 ttagtcgcat aaagtagaat acttgcgact agaaccggag acattacgcc atgaacaaga
901 gcgccgccgc tggcctgctg ggctatgccc gcgtcagcac cgacgaccag gacttgacca
961 accaacgggc cgaactgcac gcggccggct gcaccaagct gttttccgag aagatcaccg
1021 gcaccaggcg cgaccgcccg gagctggcca ggatgcttga ccacctacgc cctggcgacg
1081 ttgtgacagt gaccaggcta gaccgcctgg cccgcagcac ccgcgaccta ctggacattg
1141 ccgagcgcat ccaggaggcc ggcgcgggcc tgcgtagcct ggcagagccg tgggccgaca
1201 ccaccacgcc ggccggccgc atggtgttga ccgtgttcgc cggcattgcc gagttcgagc
1261 gttccctaat catcgaccgc acccggagcg ggcgcgaggc cgccaaggcc cgaggcgtga
1321 agtttggccc ccgccctacc ctcaccccgg cacagatcgc gcacgcccgc gagctgatcg
1381 accaggaagg ccgcaccgtg aaagaggcgg ctgcactgct tggcgtgcat cgctcgaccc
1441 tgtaccgcgc acttgagcgc agcgaggaag tgacgcccac cgaggccagg cggcgcggtg
1501 ccttccgtga ggacgcattg accgaggccg acgccctggc ggccgccgag aatgaacgcc
1561 aagaggaaca agcatgaaac cgcaccagga cggccaggac gaaccgtttt tcattaccga
1621 agagatcgag gcggagatga tcgcggccgg gtacgtgttc gagccgcccg cgcacgtctc
1681 aaccgtgcgg ctgcatgaaa tcctggccgg tttgtctgat gccaagctgg cggcctggcc
1741 ggccagcttg gccgctgaag aaaccgagcg ccgccgtcta aaaaggtgat gtgtatttga
1801 gtaaaacagc ttgcgtcatg cggtcgctgc gtatatgatg cgatgagtaa ataaacaaat
1861 acgcaagggg aacgcatgaa ggttatcgct gtacttaacc agaaaggcgg gtcaggcaag
1921 acgaccatcg caacccatct agcccgcgcc ctgcaactcg ccggggccga tgttctgtta
1981 gtcgattccg atccccaggg cagtgcccgc gattgggcgg ccgtgcggga agatcaaccg
2041 ctaaccgttg tcggcatcga ccgcccgacg attgaccgcg acgtgaaggc catcggccgg
2101 cgcgacttcg tagtgatcga cggagcgccc caggcggcgg acttggctgt gtccgcgatc
2161 aaggcagccg acttcgtgct gattccggtg cagccaagcc cttacgacat atgggccacc
2221 gccgacctgg tggagctggt taagcagcgc attgaggtca cggatggaag gctacaagcg
2281 gcctttgtcg tgtcgcgggc gatcaaaggc acgcgcatcg gcggtgaggt tgccgaggcg
2341 ctggccgggt acgagctgcc cattcttgag tcccgtatca cgcagcgcgt gagctaccca
2401 ggcactgccg ccgccggcac aaccgttctt gaatcagaac ccgagggcga cgctgcccgc
2461 gaggtccagg cgctggccgc tgaaattaaa tcaaaactca tttgagttaa tgaggtaaag
2521 agaaaatgag caaaagcaca aacacgctaa gtgccggccg tccgagcgca cgcagcagca
2581 aggctgcaac gttggccagc ctggcagaca cgccagccat gaagcgggtc aactttcagt
2641 tgccggcgga ggatcacacc aagctgaaga tgtacgcggt acgccaaggc aagaccatta
2701 ccgagctgct atctgaatac atcgcgcagc taccagagta aatgagcaaa tgaataaatg
2761 agtagatgaa ttttagcggc taaaggaggc ggcatggaaa atcaagaaca accaggcacc
2821 gacgccgtgg aatgccccat gtgttgagga acgggcggtt ggccaggcgt aagcggctgg
2881 gttgcctgcc ggccctgcaa tggcactgga acccccaagc ccgaggaatc ggcgtgagcg
2941 gtcgcaaacc atccggcccg gtacaaatcg gcgcggcgct gggtgatgac ctggtggaga
3001 agttgaaggc cgcgcaggcc gcccagcggc aacgcatcga ggcagaagca cgccccggtg
3061 aatcgtggca agcggccgct gatcgaatcc gcaaagaatc ccggcaaccg ccggcagccg
3121 gtgcgccgtc gattaggaag ccgcccaagg gcgacgagca accagatttt ttcgttccga
3181 tgctctatga cgtgggcacc cgcgatagtc gcagcatcat ggacgtggcc gttttccgtc
3241 tgtcgaagcg tgaccgacga gctggcgagg tgatccgcta cgagcttcca gacgggcacg
3301 tagaggtttc cgcagggccg gccggcatgg ccagtgtgtg ggattacgac ctggtactga
3361 tggcggtttc ccatctaacc gaatccatga accgataccg ggaagggaag ggagacaagc
3421 ccggccgcgt gttccgtcca cacgttgcgg acgtactcaa gttctgccgg cgagccgatg
3481 gcggaaagca gaaagacgac ctggtagaaa cctgcattcg gttaaacacc acgcacgttg
3541 ccatgcagcg tacgaagaag gccaagaacg gccgcctggt gacggtatcc gagggtgaag
3601 ccttgattag ccgctacaag atcgtaaaga gcgaaaccgg gcggccggag tacatcgaga
3661 tcgagctagc tgattggatg taccgcgaga tcacagaagg caagaacccg gacgtgctga
3721 cggttcaccc cgattacttt ttgatcgatc ccggcatcgg ccgttttctc taccgcctgg
3781 cacgccgcgc cgcaggcaag gcagaagcca gatggttgtt caagacgatc tacgaacgca
3841 gtggcagcgc cggagagttc aagaagttct gtttcaccgt gcgcaagctg atcgggtcaa
3901 atgacctgcc ggagtacgat ttgaaggagg aggcggggca ggctggcccg atcctagtca
3961 tgcgctaccg caacctgatc gagggcgaag catccgccgg ttcctaatgt acggagcaga
4021 tgctagggca aattgcccta gcaggggaaa aaggtcgaaa agcactcttt cctgtgaata
4081 gcacgtacat tgggaaccca aagccgtaca ttgggaaccg gaacccgtac attgggaacc
4141 caaagccgta cattgggaac cggtcacaca tgtaagtgac tgatataaaa gagaaaaaag
4201 gcgatttttc cgcctaaaac tctttaaaac ttattaaaac tcttaaaacc cgcctggcct
4261 gtgcataact gtctggccag cgcacagccg aagagctgca aaaagcgcct acccttcggt
4321 cgctgcgctc cctacgcccc gccgcttcgc gtcggcctat cgcggccgct ggccgctcaa
4381 aaatggctgg cctacggcca ggcaatctac cagggcgcgg acaagccgcg ccgtcgccac
4441 tcgaccgccg gcgcccacat caaggcaccc tgcctcgcgc gtttcggtga tgacggtgaa
4501 aacctctgac acatgcagct cccggagacg gtcacagctt gtctgtaagc ggatgccggg
4561 agcagacaag cccgtcaggg cgcgtcagcg ggtgttggcg ggtgtcgggg cgcagccatg
4621 acccagtcac gtagcgatag cggagtgtat actggcttaa ctatgcggca tcagagcaga
4681 ttgtactgag agtgcaccat atgcggtgtg aaataccgca cagatgcgta aggagaaaat
4741 accgcatcag gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc
4801 tgcggcgagc ggtatcagct cactcaaagg cggtaatacg gttatccaca gaatcagggg
4861 ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg
4921 ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac aaaaatcgac
4981 gctcaagtca gaggtggcga aacccgacag gactataaag ataccaggcg tttccccctg
5041 gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac ctgtccgcct
5101 ttctcccttc gggaagcgtg gcgctttctc atagctcacg ctgtaggtat ctcagttcgg
5161 tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct
5221 gcgccttatc cggtaactat cgtcttgagt ccaacccggt aagacacgac ttatcgccac
5281 tggcagcagc cactggtaac aggattagca gagcgaggta tgtaggcggt gctacagagt
5341 tcttgaagtg gtggcctaac tacggctaca ctagaaggac agtatttggt atctgcgctc
5401 tgctgaagcc agttaccttc ggaaaaagag ttggtagctc ttgatccggc aaacaaacca
5461 ccgctggtag cggtggtttt tttgtttgca agcagcagat tacgcgcaga aaaaaaggat
5521 ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc tcagtggaac gaaaactcac
5581 gttaagggat tttggtcatg cattctaggt actaaaacaa ttcatccagt aaaatataat
5641 attttatttt ctcccaatca ggcttgatcc ccagtaagtc aaaaaatagc tcgacatact
5701 gttcttcccc gatatcctcc ctgatcgacc ggacgcagaa ggcaatgtca taccacttgt
5761 ccgccctgcc gcttctccca agatcaataa agccacttac tttgccatct ttcacaaaga
5821 tgttgctgtc tcccaggtcg ccgtgggaaa agacaagttc ctcttcgggc ttttccgtct
5881 ttaaaaaatc atacagctcg cgcggatctt taaatggagt gtcttcttcc cagttttcgc
5941 aatccacatc ggccagatcg ttattcagta agtaatccaa ttcggctaag cggctgtcta
6001 agctattcgt atagggacaa tccgatatgt cgatggagtg aaagagcctg atgcactccg
6061 catacagctc gataatcttt tcagggcttt gttcatcttc atactcttcc gagcaaagga
6121 cgccatcggc ctcactcatg agcagattgc tccagccatc atgccgttca aagtgcagga
6181 cctttggaac aggcagcttt ccttccagcc atagcatcat gtccttttcc cgttccacat
6241 cataggtggt ccctttatac cggctgtccg tcatttttaa atataggttt tcattttctc
6301 ccaccagctt atatacctta gcaggagaca ttccttccgt atcttttacg cagcggtatt
6361 tttcgatcag ttttttcaat tccggtgata ttctcatttt agccatttat tatttccttc
6421 ctcttttcta cagtatttaa agatacccca agaagctaat tataacaaga cgaactccaa
6481 ttcactgttc cttgcattct aaaaccttaa ataccagaaa acagcttttt caaagttgtt
6541 ttcaaagttg gcgtataaca tagtatcgac ggagccgatt ttgaaaccgc ggtgatcaca
6601 ggcagcaacg ctctgtcatc gttacaatca acatgctacc ctccgcgaga tcatccgtgt
6661 ttcaaacccg gcagcttagt tgccgttctt ccgaatagca tcggtaacat gagcaaagtc
6721 tgccgcctta caacggctct cccgctgacg ccgtcccgga ctgatgggct gcctgtatcg
6781 agtggtgatt ttgtgccgag ctgccggtcg gggagctgtt ggctggctgg tggcaggata
6841 tattgtggtg taaacaaatt gacgcttaga caacttaata acacattgcg gacgttttta
6901 atgtactgaa ttaacgccga attaattcgg gggatctgga ttttagtact ggattttggt
6961 tttaggaatt agaaatttta ttgatagaag tattttacaa atacaaatac atactaaggg
7021 tttcttatat gctcaacaca tgagcgaaac cctataggaa ccctaattcc cttatctggg
7081 aactactcac acattattat ggagaaactc gagcttgtcg atcgacagat cccggtcggc
7141 atctactcta tttctttgcc ctcggacgag tgctggggcg tcggtttcca ctatcggcga
7201 gtacttctac acagccatcg gtccagacgg ccgcgcttct gcgggcgatt tgtgtacgcc
7261 cgacagtccc ggctccggat cggacgattg cgtcgcatcg accctgcgcc caagctgcat
7321 catcgaaatt gccgtcaacc aagctctgat agagttggtc aagaccaatg cggagcatat
7381 acgcccggag tcgtggcgat cctgcaagct ccggatgcct ccgctcgaag tagcgcgtct
7441 gctgctccat acaagccaac cacggcctcc agaagaagat gttggcgacc tcgtattggg
7501 aatccccgaa catcgcctcg ctccagtcaa tgaccgctgt tatgcggcca ttgtccgtca
7561 ggacattgtt ggagccgaaa tccgcgtgca cgaggtgccg gacttcgggg cagtcctcgg
7621 cccaaagcat cagctcatcg agagcctgcg cgacggacgc actgacggtg tcgtccatca
7681 cagtttgcca gtgatacaca tggggatcag caatcgcgca tatgaaatca cgccatgtag
7741 tgtattgacc gattccttgc ggtccgaatg ggccgaaccc gctcgtctgg ctaagatcgg
7801 ccgcagcgat cgcatccata gcctccgcga ccggttgtag aacagcgggc agttcggttt
7861 caggcaggtc ttgcaacgtg acaccctgtg aacggcggga gatgcaatag gtcaggctct
7921 cgctaaactc cccaatgtca agcacttccg gaatcgggag cgcggccgat gcaaagtgcc
7981 gataaacata acgatctttg tagaaaccat cggcgcagct atttacccgc aggacatatc
8041 cacgccctcc tacatcgaag ctgaaagcac gagattcttc gccctccgag agctgcatca
8101 ggtcggagac gctgtcgaac ttttcgatca gaaacttctc gacagacgtc gcggtgagtt
8161 caggcttttt catatctcat tgccccccgg atctgcgaaa gctcgagaga gatagatttg
8221 tagagagaga ctggtgattt cagcgtgtcc tctccaaatg aaatgaactt ccttatatag
8281 aggaagggtc ttgcgaagga tagtgggatt gtgcgtcatc ccttacgtca gtggagatat
8341 cacatcaatc cacttgcttt gaagacgtgg ttggaacgtc ttctttttcc acgatgctcc
8401 tcgtgggtgg gggtccatct ttgggaccac tgtcggcaga ggcatcttga acgatagcct
8461 ttcctttatc gcaatgatgg catttgtagg tgccaccttc cttttctact gtccttttga
8521 tgaagtgaca gatagctggg caatggaatc cgaggaggtt tcccgatatt accctttgtt
8581 gaaaagtctc aatagccctt tggtcttctg agactgtatc tttgatattc ttggagtaga
8641 cgagagtgtc gtgctccacc atgttcacat caatccactt gctttgaaga cgtggttgga
8701 acgtcttctt tttccacgat gctcctcgtg ggtgggggtc catctttggg accactgtcg
8761 gcagaggcat cttgaacgat agcctttcct ttatcgcaat gatggcattt gtaggtgcca
8821 ccttcctttt ctactgtcct tttgatgaag tgacagatag ctgggcaatg gaatccgagg
8881 aggtttcccg atattaccct ttgttgaaaa gtctcaatag ccctttggtc ttctgagact
8941 gtatctttga tattcttgga gtagacgaga gtgtcgtgct ccaccatgtt ggcaagctgc
9001 tctagccaat acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagctggc
9061 acgacaggtt tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat gtgagttagc
9121 tcactcatta ggcaccccag gctttacact ttatgcttcc ggctcgtatg ttgtgtggaa
9181 ttgtgagcgg ataacaattt cacacaggaa acagctatga catgattacg ccaagcttaa
9241 ggaatcttta aacatacgaa cagatcactt aaagttcttc tgaagcaact taaagttatc
9301 aggcatgcat ggatcttgga ggaatcagat gtgcagtcag ggaccatagc acaagacagg
9361 cgtcttctac tggtgctacc agcaaatgct ggaagccggg aacactgggt acgttggaaa
9421 ccacgtgatg tgaagaagta agataaactg taggagaaaa gcatttcgta gtgggccatg
9481 aagcctttca ggacatgtat tgcagtatgg gccggcccat tacgcaattg gacgacaaca
9541 aagactagta ttagtaccac ctcggctatc cacatagatc aaagctgatt taaaagagtt
9601 gtgcagatga tccgtggcag gagaccgagg tctcggtttt agagctagaa atagcaagtt
9661 aaaataaggc tagtccgtta tcaacttgaa aaagtggcac cgagtcggtg cttttttgtt
9721 ttagagctag aaatagcaag ttaaaataag gctagtccgt ttttagcgcg tgcatgcctg
9781 caggtccaca aattcgggtc aaggcggaag ccagcgcgcc accccacgtc agcaaatacg
9841 gaggcgcggg gttgacggcg tcacccggtc ctaacggcga ccaacaaacc agccagaaga
9901 aattacagta aaaaaaaagt aaattgcact ttgatccacc ttttattacc taagtctcaa
9961 tttggatcac ccttaaacct atcttttcaa tttgggccgg gttgtggttt ggactaccat
10021 gaacaacttt tcgtcatgtc taacttccct ttcagcaaac atatgaacca tatatagagg
10081 agatcggccg tatactagag ctgatgtgtt taaggtcgtt gattgcacga gaaaaaaaaa
10141 tccaaatcgc aacaatagca aatttatctg gttcaaagtg aaaagatatg tttaaaggta
10201 gtccaaagta aaacttatag ataataaaat gtggtccaaa gcgtaattca ctcaaaaaaa
10261 atcaacgaga cgtgtaccaa acggagacaa acggcatctt ctcgaaattt cccaaccgct
10321 cgctcgcccg cctcgtcttc ccggaaaccg cggtggtttc agcgtggcgg attctccaag
10381 cagacggaga cgtcacggca cgggactcct cccaccaccc aaccgccata aataccagcc
10441 ccctcatctc ctctcctcgc atcagctcca cccccgaaaa atttctcccc aatctcgcga
10501 ggctctcgtc gtcgaatcga atcctctcgc gtcctcaagg tacgctgctt ctcctctcct
10561 cgcttcgttt cgattcgatt tcggacgggt gaggttgttt tgttgctaga tccgattggt
10621 ggttagggtt gtcgatgtga ttatcgtgag atgtttaggg gttgtagatc tgatggttgt
10681 gatttgggca cggttggttc gataggtgga atcgtggtta ggttttggga ttggatgttg
10741 gttctgatga ttggggggaa tttttacggt tagatgaatt gttggatgat tcgattgggg
10801 aaatcggtgt agatctgttg gggaattgtg gaactagtca tgcctgagtg attggtgcga
10861 tttgtagcgt gttccatctt gtaggccttg ttgcgagcat gttcagatct actgttccgc
10921 tcttgattga gttattggtg ccatgggttg gtgcaaacac aggctttaat atgttatatc
10981 tgttttgtgt ttgatgtaga tctgtagggt agttcttctt agacatggtt caattatgta
11041 gcttgtgcgt ttcgatttga tttcatatgt tcacagatta gataatgatg aactctttta
11101 attaattgtc aatggtaaat aggaagtctt gtcgctatat ctgtcataat gatctcatgt
11161 tactatctgc cagtaattta tgctaagaac tatattagaa tatcatgtta caatctgtag
11221 taatatcatg ttacaatctg tagttcatct atataatcta ttgtggtaat ttctttttac
11281 tatctgtgtg aagattattg ccactagttc attctactta tttctgaagt tcaggatacg
11341 tgtgctgtta ctacctatct gaatacatgt gtgatgtgcc tgttactatc tttttgaata
11401 catgtatgtt ctgttggaat atgtttgctg tttgatccgt tgttgtgtcc ttaatcttgt
11461 gctagttctt accctatctg tttggtgatt atttcttgca gatagttatc aacaagtttg
11521 tacaaaaaag caggcttcga aggagataga accaattctc taaggaaata cttaaccatg
11581 gactataagg accacgacgg agactacaag gatcatgata ttgattacaa agacgatgac
11641 gataagatgg ccccaaagaa gaagcggaag gtcggtatcc acggagtccc agcagccgac
11701 aagaagtaca gcatcggcct ggacatcggc accaactctg tgggctgggc cgtgatcacc
11761 gacgagtaca aggtgcccag caagaaattc aaggtgctgg gcaacaccga ccggcacagc
11821 atcaagaaga acctgatcgg agccctgctg ttcgacagcg gcgaaacagc cgaggccacc
11881 cggctgaaga gaaccgccag aagaagatac accagacgga agaaccggat ctgctatctg
11941 caagagatct tcagcaacga gatggccaag gtggacgaca gcttcttcca cagactggaa
12001 gagtccttcc tggtggaaga ggataagaag cacgagcggc accccatctt cggcaacatc
12061 gtggacgagg tggcctacca cgagaagtac cccaccatct accacctgag aaagaaactg
12121 gtggacagca ccgacaaggc cgacctgcgg ctgatctatc tggccctggc ccacatgatc
12181 aagttccggg gccacttcct gatcgagggc gacctgaacc ccgacaacag cgacgtggac
12241 aagctgttca tccagctggt gcagacctac aaccagctgt tcgaggaaaa ccccatcaac
12301 gccagcggcg tggacgccaa ggccatcctg tctgccagac tgagcaagag cagacggctg
12361 gaaaatctga tcgcccagct gcccggcgag aagaagaatg gcctgttcgg aaacctgatt
12421 gccctgagcc tgggcctgac ccccaacttc aagagcaact tcgacctggc cgaggatgcc
12481 aaactgcagc tgagcaagga cacctacgac gacgacctgg acaacctgct ggcccagatc
12541 ggcgaccagt acgccgacct gtttctggcc gccaagaacc tgtccgacgc catcctgctg
12601 agcgacatcc tgagagtgaa caccgagatc accaaggccc ccctgagcgc ctctatgatc
12661 aagagatacg acgagcacca ccaggacctg accctgctga aagctctcgt gcggcagcag
12721 ctgcctgaga agtacaaaga gattttcttc gaccagagca agaacggcta cgccggctac
12781 attgacggcg gagccagcca ggaagagttc tacaagttca tcaagcccat cctggaaaag
12841 atggacggca ccgaggaact gctcgtgaag ctgaacagag aggacctgct gcggaagcag
12901 cggaccttcg acaacggcag catcccccac cagatccacc tgggagagct gcacgccatt
12961 ctgcggcggc aggaagattt ttacccattc ctgaaggaca accgggaaaa gatcgagaag
13021 atcctgacct tccgcatccc ctactacgtg ggccctctgg ccaggggaaa cagcagattc
13081 gcctggatga ccagaaagag cgaggaaacc atcaccccct ggaacttcga ggaagtggtg
13141 gacaagggcg cttccgccca gagcttcatc gagcggatga ccaacttcga taagaacctg
13201 cccaacgaga aggtgctgcc caagcacagc ctgctgtacg agtacttcac cgtgtataac
13261 gagctgacca aagtgaaata cgtgaccgag ggaatgagaa agcccgcctt cctgagcggc
13321 gagcagaaaa aggccatcgt ggacctgctg ttcaagacca accggaaagt gaccgtgaag
13381 cagctgaaag aggactactt caagaaaatc gagtgcttcg actccgtgga aatctccggc
13441 gtggaagatc ggttcaacgc ctccctgggc acataccacg atctgctgaa aattatcaag
13501 gacaaggact tcctggacaa tgaggaaaac gaggacattc tggaagatat cgtgctgacc
13561 ctgacactgt ttgaggacag agagatgatc gaggaacggc tgaaaaccta tgcccacctg
13621 ttcgacgaca aagtgatgaa gcagctgaag cggcggagat acaccggctg gggcaggctg
13681 agccggaagc tgatcaacgg catccgggac aagcagtccg gcaagacaat cctggatttc
13741 ctgaagtccg acggcttcgc caacagaaac ttcatgcagc tgatccacga cgacagcctg
13801 acctttaaag aggacatcca gaaagcccag gtgtccggcc agggcgatag cctgcacgag
13861 cacattgcca atctggccgg cagccccgcc attaagaagg gcatcctgca gacagtgaag
13921 gtggtggacg agctcgtgaa agtgatgggc cggcacaagc ccgagaacat cgtgatcgaa
13981 atggccagag agaaccagac cacccagaag ggacagaaga acagccgcga gagaatgaag
14041 cggatcgaag agggcatcaa agagctgggc agccagatcc tgaaagaaca ccccgtggaa
14101 aacacccagc tgcagaacga gaagctgtac ctgtactacc tgcagaatgg gcgggatatg
14161 tacgtggacc aggaactgga catcaaccgg ctgtccgact acgatgtgga ccatatcgtg
14221 cctcagagct ttctgaagga cgactccatc gacaacaagg tgctgaccag aagcgacaag
14281 aaccggggca agagcgacaa cgtgccctcc gaagaggtcg tgaagaagat gaagaactac
14341 tggcggcagc tgctgaacgc caagctgatt acccagagaa agttcgacaa tctgaccaag
14401 gccgagagag gcggcctgag cgaactggat aaggccggct tcatcaagag acagctggtg
14461 gaaacccggc agatcacaaa gcacgtggca cagatcctgg actcccggat gaacactaag
14521 tacgacgaga atgacaagct gatccgggaa gtgaaagtga tcaccctgaa gtccaagctg
14581 gtgtccgatt tccggaagga tttccagttt tacaaagtgc gcgagatcaa caactaccac
14641 cacgcccacg acgcctacct gaacgccgtc gtgggaaccg ccctgatcaa aaagtaccct
14701 aagctggaaa gcgagttcgt gtacggcgac tacaaggtgt acgacgtgcg gaagatgatc
14761 gccaagagcg agcaggaaat cggcaaggct accgccaagt acttcttcta cagcaacatc
14821 atgaactttt tcaagaccga gattaccctg gccaacggcg agatccggaa gcggcctctg
14881 atcgagacaa acggcgaaac cggggagatc gtgtgggata agggccggga ttttgccacc
14941 gtgcggaaag tgctgagcat gccccaagtg aatatcgtga aaaagaccga ggtgcagaca
15001 ggcggcttca gcaaagagtc tatcctgccc aagaggaaca gcgataagct gatcgccaga
15061 aagaaggact gggaccctaa gaagtacggc ggcttcgaca gccccaccgt ggcctattct
15121 gtgctggtgg tggccaaagt ggaaaagggc aagtccaaga aactgaagag tgtgaaagag
15181 ctgctgggga tcaccatcat ggaaagaagc agcttcgaga agaatcccat cgactttctg
15241 gaagccaagg gctacaaaga agtgaaaaag gacctgatca tcaagctgcc taagtactcc
15301 ctgttcgagc tggaaaacgg ccggaagaga atgctggcct ctgccggcga actgcagaag
15361 ggaaacgaac tggccctgcc ctccaaatat gtgaacttcc tgtacctggc cagccactat
15421 gagaagctga agggctcccc cgaggataat gagcagaaac agctgtttgt ggaacagcac
15481 aagcactacc tggacgagat catcgagcag atcagcgagt tctccaagag agtgatcctg
15541 gccgacgcta atctggacaa agtgctgtcc gcctacaaca agcaccggga taagcccatc
15601 agagagcagg ccgagaatat catccacctg tttaccctga ccaatctggg agcccctgcc
15661 gccttcaagt actttgacac caccatcgac cggaagaggt acaccagcac caaagaggtg
15721 ctggacgcca ccctgatcca ccagagcatc accggcctgt acgagacacg gatcgacctg
15781 tctcagctgg gaggcgacaa aaggccggcg gccacgaaaa aggccggcca ggcaaaaaag
15841 aaaaagtaag aattcgcggc cgcactcgag atatctagac ccagcttt
//
质粒菌株产品操作说明书
一、扩增流程
收到产品后，请先根据产品管壁标签来判断产品形式，并在扩增前准确查找该质粒菌株的抗性、感受态和培养温度。
1、质粒干粉（常温运输，存于-20度，90天保质期，请务必转化挑单克隆培养，不要直接使用和测序）
①收到质粒干粉后请先5000rpm离心1min，再加入20μl ddH2O去离子水溶解质粒；
②取1支100μl 感受态于冰上解冻10min，加入2μl质粒，再冰浴30min后，42℃热激60s，不要搅动，再冰浴2min；（从第二步开始均要在超净工作台中无菌操作）
③加入900μl无抗的LB液体培养基，180rpm震荡37℃培养45min；
④6000rpm离心5min，仅留100μl上清液重悬细菌沉淀，并涂布至目标质粒抗性的LB平板上；（可使用本平台的平板涂布专用玻璃珠进行涂布，可以比传统涂布方法获得更多转化子）
⑤将平板正向培养1h，再倒置37℃培养14h。如果要求是30度则培养20h；
（菌落过多则将质粒稀释后再转化。没有菌落则加入10μl质粒转化。另不要直接转表达感受态，要先转克隆感受态，重提质粒后再导入表达感受态）
⑥挑取单菌落至LB液体培养基中，加入对应抗生素，220rpm震荡培养14h，根据实验需要和质粒提取试剂盒说明书提取质粒。
2、甘油菌种（冰袋运输，存于-80℃，保质期90天，请务必划线挑单克隆培养）
四区划线后挑单菌落培养，酵母菌需要先液体复苏再四区划线，再挑单菌落液体培养。
3、穿刺菌种（冰袋运输，存于4℃，保质期7天）
穿刺接种，液体培养后四区划线，再挑单菌落液体培养。
4、菌落平板（冰袋运输，存于4℃，保质期7天）
直接挑取单菌落至液体培养基中。
5、液体质粒（冰袋运输，存于-20℃，保质期90天）
单独提取的液体质粒收到后可直接使用。
6、滤纸质粒（常温运输，存于-20度，90天保质期，请务必转化挑单克隆培养，不要直接使用和测序）
收到货后将滤纸画圈部分剪下放入EP管中，加100ul无菌水将滤纸浸湿并浸泡5min，吸取5ul质粒转化，离心全涂。
二、转化图片
| P1019/pCR3.1哺乳表达质粒 |
| P1027/pBK-CMV哺乳表达质粒 |
| P1101/pJG-ALPHA-MHC哺乳表达质粒 |
| P1149/pRC-CMV哺乳表达质粒 |
| P1750/pRK5-HA哺乳表达质粒 |
| P1300/pRK5-2×HA哺乳表达质粒 |
| P0133/pEGFP-N1哺乳荧光质粒 |
| P0153/pEGFP-N2哺乳荧光质粒 |
| P0452/pEGFP-N3哺乳荧光质粒 |
| P0134/pEGFP-C1哺乳荧光质粒 |
| P0135/pEGFP-C2哺乳荧光质粒 |
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pRGEB32植物编辑质粒
￥100 - 1000
