pRed-FLp线虫同源重组编辑质粒

基本信息
名称：pRed-FLp线虫同源重组编辑质粒
别称: pRed-FLp

复制子:	pSC101
原核抗性:	Amp
克隆菌株:	DH5a
培养条件:	30度

质粒属性

载体宿主：	昆虫细胞
载体用途：
基因种属：
基因类型：
原核抗性：	Amp
真核抗性：
荧光蛋白：

质粒简介
 pRed-FLp是一个线虫同源重组基因编辑质粒。

质粒图谱


质粒序列
LOCUS       Exported               11160 bp ds-DNA     circular SYN 09-JAN-2018
DEFINITION  synthetic circular DNA
FEATURES             Location/Qualifiers
     source          1..11160
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     CDS             1..861
                     /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"
     CDS             868..1494
                     /codon_start=1
                     /gene="tetR"
                     /product="tetracycline repressor TetR"
                     /label=TetR
                     /note="TetR binds to the tetracycline operator tetO to 
                     inhibit transcription. This inhibition can be relieved by 
                     adding tetracycline or doxycycline."
                     /translation="MMSRLDKSKVINSALELLNEVGIEGLTTRKLAQKLGVEQPTLYWH
                     VKNKRALLDALAIEMLDRHHTHFCPLEGESWQDFLRNNAKSFRCALLSHRDGAKVHLGT
                     RPTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEDQEHQVAKEERETPT
                     TDSMPPLLRQAIELFDHQGAEPAFLFGLELIICGLEKQLKCESGS"
     CDS             complement(1536..2486)
                     /codon_start=1
                     /gene="rep101"
                     /product="RepA protein needed for replication with the 
                     pSC101 origin"
                     /label=Rep101
                     /translation="MSELVVFKANELAISRYDLTEHETKLILCCVALLNPTIENPTRKE
                     RTVSFTYNQYAQMMNISRENAYGVLAKATRELMTRTVEIRNPLVKGFEIFQWTNYAKFS
                     SEKLELVFSEEILPYLFQLKKFIKYNLEHVKSFENKYSMRIYEWLLKELTQKKTHKANI
                     EISLDEFKFMLMLENNYHEFKRLNQWVLKPISKDLNTYSNMKLVVDKRGRPTDTLIFQV
                     ELDRQMDLVTELENNQIKMNGDKIPTTITSDSYLRNGLRKTLHDALTAKIQLTSFEAKF
                     L-SDMQSKHDLNGSFSWLTQKQRTTLENILAKYGRI"
     rep_origin      2534..2756
                     /label=pSC101 ori
                     /note="low-copy replication origin that requires the Rep101
                     protein"
     promoter        3300..3355
                     /gene="tetR"
                     /label=tetR/tetA promoters
                     /note="overlapping promoters for bacterial tetR and tetA"
     protein_bind    3306..3324
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O1 for the tetR and tetA genes"
     protein_bind    3336..3354
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     CDS             3403..4674
                     /codon_start=1
                     /product="site-specific recombinase"
                     /label=FLP
                     /note="FLP is a site-specific recombinase from 
                     Saccharomyces cerevisiae. Recombination occurs at FRT 
                     sequences."
                     /translation="MPQFDILCKTPPKVLVRQFVERFERPSGEKIALCAAELTYLCWMI
                     THNGTAIKRATFMSYNTIISNSLSLDIVNKSLQFKYKTQKATILEASLKKLIPAWEFTI
                     IPYYGQKHQSDITDIVSSLQLQFESSEEADKGNSHSKKMLKALLSEGESIWEITEKILN
                     SFEYTSRFTKTKTLYQFLFLATFINCGRFSDIKNVDPKSFKLVQNKYLGVIIQCLVTET
                     KTSVSRHIYFFSARGRIDPLVYLDEFLRNSEPVLKRVNRTGNSSSNKQEYQLLKDNLVR
                     SYNKALKKNAPYSIFAIKNGPKSHIGRHLMTSFLSMKGLTELTNVVGNWSDKRASAVAR
                     TTYTHQITAIPDHYFALVSRYYAYDPISKEMIALKDETNPIEEWQHIEQLKGSAEGSIR
                     YPAWNGIISQEVLDYLSSYINRRI"
     CDS             complement(4836..5072)
                     /codon_start=1
                     /gene="dhfrIIb from plasmid R388"
                     /product="E. coli plasmid-associated dihydrofolate 
                     reductase"
                     /label=TpR
                     /note="confers resistance to trimethoprim"
                     /translation="MGQSSDEANAPVAGQFALPLSATFGLGDRVRKKSGAAWQGQVVGW
                     YCTKLTPEGYAVESESHPGSVQIYPVAALERVA"
     promoter        complement(5399..5427)
                     /gene="intI1 (promoter lies within the coding sequence)"
                     /label=Pc promoter
                     /note="class 1 integron promoter"
     CDS             complement(5458..6306)
                     /codon_start=1
                     /gene="E. coli rhaR"
                     /product="transcriptional activator of rhaR and rhaS"
                     /label=rhaR
                     /note="needed for rhamnose-dependent activation of the rhaB
                     promoter"
                     /translation="MAHQLKLLKDDFFASDQQAVAVADRYPQDVFAEHTHDFCELVIVW
                     RGNGLHVLNDRPYRITRGDLFYIHADDKHSYASVNDLVLQNIIYCPERLKLNLDWQGAI
                     PGFNASAGQPHWRLGSMGMAQARQVIGQLEHESSQHVPFANEMAELLFGQLVMLLNRHR
                     YTSDSLPPTSSETLLDKLITRLAASLKSPFALDKFCDEASCSERVLRQQFRQQTGMTIN
                     QYLRQVRVCHAQYLLQHSRLLISDISTECGFEDSNYFSVVFTRETGMTPSQWRHLNSQK
                     D"
     CDS             complement(6380..7216)
                     /codon_start=1
                     /gene="E. coli rhaS"
                     /product="positive regulator of the rhaB promoter"
                     /label=rhaS
                     /note="activated by L-rhamnose"
                     /translation="MTVLHSVDFFPSGNASVAIEPRLPQADFPEHHHDFHEIVIVEHGT
                     GIHVFNGQPYTITGGTVCFVRDHDRHLYEHTDNLCLTNVLYRSPDRFQFLAGLNQLLPQ
                     ELDGQYPSHWRVNHSVLQQVRQLVAQMEQQEGENDLPSTASREILFMQLLLLLRKSSLQ
                     ENLENSASRLNLLLAWLEDHFADEVNWDAVADQFSLSLRTLHRQLKQQTGLTPQRYLNR
                     LRLMKARHLLRHSEASVTDIAYRCGFSDSNHFSTLFRREFNWSPRDIRQGRDGFLQ"
     promoter        7362..7480
                     /label=rhaB promoter
                     /note="promoter of the E. coli rhaBAD operon, conferring 
                     tight induction with L-rhamnose and repression with 
                     D-glucose in the presence of RhaR and RhaS (Giacalone et 
                     al., 2006)"
     CDS             7504..7920
                     /codon_start=1
                     /gene="gam"
                     /product="inhibitor of the host RecBCD nuclease in the 
                     lambda Red system"
                     /label=Gam
                     /note="protects the double-stranded DNA substrate for 
                     recombination"
                     /translation="MDINTETEIKQKHSLTPFPVFLISPAFRGRYFHSYFRSSAMNAYY
                     IQDRLEAQSWARHYQQLAREEKEAELADDMEKGLPQHLFESLCIDHLQRHGASKKSITR
                     AFDDDVEFQERMAEHIRYMVETIAHHQVDIDSEV"
     CDS             7926..8711
                     /codon_start=1
                     /gene="bet"
                     /product="single-stranded DNA binding recombinase in the 
                     lambda Red system"
                     /label=Beta
                     /note="mediates pairing of a single-stranded DNA with a 
                     complementary target"
                     /translation="MSTALATLAGKLAERVGMDSVDPQELITTLRQTAFKGDASDAQFI
                     ALLIVANQYGLNPWTKEIYAFPDKQNGIVPVVGVDGWSRIINENQQFDGMDFEQDNESC
                     TCRIYRKDRNHPICVTEWMDECRREPFKTREGREITGPWQSHPKRMLRHKAMIQCARLA
                     FGFAGIYDKDEAERIVENTAYTAERQPERDITPVNDETMQEINTLLIALDKTWDDDLLP
                     LCSQIFRRDIRASSELTQAEAVKALGFLKQKAAEQKVAA"
     CDS             8708..9388
                     /codon_start=1
                     /gene="exo"
                     /product="5' to 3' double-stranded DNA exonuclease in the 
                     lambda Red system"
                     /label=Exo
                     /note="creates 3' single-stranded DNA tails"
                     /translation="MTPDIILQRTGIDVRAVEQGDDAWHKLRLGVITASEVHNVIAKPR
                     SGKKWPDMKMSYFHTLLAEVCTGVAPEVNAKALAWGKQYENDARTLFEFTSGVNVTESP
                     IIYRDESMRTACSPDGLCSDGNGLELKCPFTSRDFMKFRLGGFEAIKSAYMAQVQYSMW
                     VTRKNAWYFANYDPRMKREGLHYVVIERDEKYMASFDEIVPEFIEKMDEALAEIGFVFG
                     EQWR"
     CDS             9412..10473
                     /codon_start=1
                     /gene="recA"
                     /product="DNA repair protein from E. coli"
                     /label=RecA
                     /translation="MAIDENKQKALAAALGQIEKQFGKGSIMRLGEDRSMDVETISTGS
                     LSLDIALGAGGLPMGRIVEIYGPESSGKTTLTLQVIAAAQREGKTCAFIDAEHALDPIY
                     ARKLGVDIDNLLCSQPDTGEQALEICDALARSGAVDVIVVDSVAALTPKAEIEGEIGDS
                     HMGLAARMMSQAMRKLAGNLKQSNTLLIFINQIRMKIGVMFGNPETTTGGNALKFYASV
                     RLDIRRIGAVKEGENVVGSETRVKVVKNKIAAPFKQAEFQILYGEGINFYGELVDLGVK
                     EKLIEKAGAWYSYKGEKIGQGKANATAWLKDNPETAKEIEKKVRELLLSNPNSTPDFSV
                     DDSEGVAETNEDF"
     rep_origin      10574..11029
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     promoter        11056..11160
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 atgagtattc aacatttccg tgtcgccctt attccctttt ttgcggcatt ttgccttcct
       61 gtttttgctc acccagaaac gctggtgaaa gtaaaagatg ctgaagatca gttgggtgca
      121 cgagtgggtt acatcgaact ggatctcaac agcggtaaga tccttgagag ttttcgcccc
      181 gaagaacgtt ttccaatgat gagcactttt aaagttctgc tatgtggcgc ggtattatcc
      241 cgtattgacg ccgggcaaga gcaactcggt cgccgcatac actattctca gaatgacttg
      301 gttgagtact caccagtcac agaaaagcat cttacggatg gcatgacagt aagagaatta
      361 tgcagtgctg ccataaccat gagtgataac actgcggcca acttacttct gacaacgatc
      421 ggaggaccga aggagctaac cgcttttttg cacaacatgg gggatcatgt aactcgcctt
      481 gatcgttggg aaccggagct gaatgaagcc ataccaaacg acgagcgtga caccacgatg
      541 cctgtagcaa tggcaacaac gttgcgcaaa ctattaactg gcgaactact tactctagct
      601 tcccggcaac aattgataga ctggatggag gcggataaag ttgcaggacc acttctgcgc
      661 tcggcccttc cggctggctg gtttattgct gataaatctg gagccggtga gcgtggctct
      721 cgcggtatca ttgcagcact ggggccagat ggtaagccct cccgtatcgt agttatctac
      781 acgacgggga gtcaggcaac tatggatgaa cgaaatagac agatcgctga gataggtgcc
      841 tcactgatta agcattggta ggaattaatg atgtctcgtt tagataaaag taaagtgatt
      901 aacagcgcat tagagctgct taatgaggtc ggaatcgaag gtttaacaac ccgtaaactc
      961 gcccagaagc taggtgtaga gcagcctaca ttgtattggc atgtaaaaaa taagcgggct
     1021 ttgctcgacg ccttagccat tgagatgtta gataggcacc atactcactt ttgcccttta
     1081 gaaggggaaa gctggcaaga ttttttacgt aataacgcta aaagttttag atgtgcttta
     1141 ctaagtcatc gcgatggagc aaaagtacat ttaggtacac ggcctacaga aaaacagtat
     1201 gaaactctcg aaaatcaatt agccttttta tgccaacaag gtttttcact agagaatgca
     1261 ttatatgcac tcagcgcagt ggggcatttt actttaggtt gcgtattgga agatcaagag
     1321 catcaagtcg ctaaagaaga aagggaaaca cctactactg atagtatgcc gccattatta
     1381 cgacaagcta tcgaattatt tgatcaccaa ggtgcagagc cagccttctt attcggcctt
     1441 gaattgatca tatgcggatt agaaaaacaa cttaaatgtg aaagtgggtc ttaaaagcag
     1501 cataaccttt ttccgtgatg gtaacttcag ctagctcaga tccttccgta tttagccagt
     1561 atgttctcta gtgtggttcg ttgtttttgc gtgagccatg agaacgaacc attgagatca
     1621 tgcttacttt gcatgtcact caaaaatttt gcctcaaaac tggtgagctg aatttttgca
     1681 gttaaagcat cgtgtagtgt ttttcttagt ccgttacgta ggtaggaatc tgatgtaatg
     1741 gttgttggta ttttgtcacc attcattttt atctggttgt tctcaagttc ggttacgaga
     1801 tccatttgtc tatctagttc aacttggaaa atcaacgtat cagtcgggcg gcctcgctta
     1861 tcaaccacca atttcatatt gctgtaagtg tttaaatctt tacttattgg tttcaaaacc
     1921 cattggttaa gccttttaaa ctcatggtag ttattttcaa gcattaacat gaacttaaat
     1981 tcatcaaggc taatctctat atttgccttg tgagttttct tttgtgttag ttcttttaat
     2041 aaccactcat aaatcctcat agagtatttg ttttcaaaag acttaacatg ttccagatta
     2101 tattttatga atttttttaa ctggaaaaga taaggcaata tctcttcact aaaaactaat
     2161 tctaattttt cgcttgagaa cttggcatag tttgtccact ggaaaatctc aaagccttta
     2221 accaaaggat tcctgatttc cacagttctc gtcatcagct ctctggttgc tttagctaat
     2281 acaccataag cattttccct actgatgttc atcatctgag cgtattggtt ataagtgaac
     2341 gataccgtcc gttctttcct tgtagggttt tcaatcgtgg ggttgagtag tgccacacag
     2401 cataaaatta gcttggtttc atgctccgtt aagtcatagc gactaatcgc tagttcattt
     2461 gctttgaaaa caactaattc agacatacat ctcaattggt ctaggtgatt ttaatcacta
     2521 taccaattga gatgggctag tcaatgataa ttactagtcc ttttcctttg agttgtgggt
     2581 atctgtaaat tctgctagac ctttgctgga aaacttgtaa attctgctag accctctgta
     2641 aattccgcta gacctttgtg tgtttttttt gtttatattc aagtggttat aatttataga
     2701 ataaagaaag aataaaaaaa gataaaaaga atagatccca gccctgtgta taactcacta
     2761 ctttagtcag ttccgcagta ttacaaaagg atgtcgcaaa cgctgtttgc tcctctacaa
     2821 aacagacctt aaaaccctaa aggcttaagt agcaccctcg caagctcggg caaatcgctg
     2881 aatattcctt ttgtctccga ccatcaggca cctgagtcgc tgtctttttc gtgacattca
     2941 gttcgctgcg ctcacggctc tggcagtgaa tgggggtaaa tggcactaca ggcgcctttt
     3001 atggattcat gcaaggaaac tacccataat acaagaaaag cccgtcacgg gcttctcagg
     3061 gcgttttatg gcgggtctgc tatgtggtgc tatctgactt tttgctgttc agcagttcct
     3121 gccctctgat tttccagtct gaccacttcg gattatcccg tgacaggtca ttcagactgg
     3181 ctaatgcacc cagtaaggca gcggtatcat caacaggctt acccgtctta ctgtctagac
     3241 tcgaggaatt gatatcaccc gacaccatcg aatggccaga tgattaattc ctaatttttg
     3301 ttgacactct atcattgata gagttatttt accactccct atcagtgata gagaaaagtg
     3361 aaatgaatag ttcgacaaaa atctagataa cgagggcaaa aaatgccaca atttgatata
     3421 ttatgtaaaa caccacctaa ggtgcttgtt cgtcagtttg tggaaaggtt tgaaagacct
     3481 tcaggtgaga aaatagcatt atgtgctgct gaactaacct atttatgttg gatgattaca
     3541 cataacggaa cagcaatcaa gagagccaca ttcatgagct ataatactat cataagcaat
     3601 tcgctgagtt tggatattgt caacaagtca ctgcagttta aatacaagac gcaaaaagca
     3661 acaattctgg aagcctcatt aaagaaattg attcctgctt gggaatttac aattattcct
     3721 tactatggac aaaaacatca atctgatatc actgatattg taagtagttt gcaattacag
     3781 ttcgaatcat cggaagaagc agataaggga aatagccaca gtaaaaaaat gcttaaagca
     3841 cttctaagtg agggtgaaag catctgggag atcactgaga aaatactaaa ttcgtttgag
     3901 tatacttcga gatttacaaa aacaaaaact ttataccaat tcctcttcct agctactttc
     3961 atcaattgtg gaagattcag cgatattaag aacgttgatc cgaaatcatt taaattagtc
     4021 caaaataagt atctgggagt aataatccag tgtttagtga cagagacaaa gacaagcgtt
     4081 agtaggcaca tatacttctt tagcgcaagg ggtaggatcg atccacttgt atatttggat
     4141 gaatttttga ggaattctga accagtccta aaacgagtaa ataggaccgg caattcttca
     4201 agcaacaagc aggaatacca attattaaaa gataacttag tcagatcgta caacaaagct
     4261 ttgaagaaaa atgcgcctta ttcaatcttt gctataaaaa atggcccaaa atctcacatt
     4321 ggaagacatt tgatgacctc atttctttca atgaagggcc taacggagtt gactaatgtt
     4381 gtgggaaatt ggagcgataa gcgtgcttct gccgtggcca ggacaacgta tactcatcag
     4441 ataacagcaa tacctgatca ctacttcgca ctagtttctc ggtactatgc atatgatcca
     4501 atatcaaagg aaatgatagc attgaaggat gagactaatc caattgagga gtggcagcat
     4561 atagaacagc taaagggtag tgctgaagga agcatacgat accccgcatg gaatgggata
     4621 atatcacagg aggtactaga ctacctttca tcctacataa atagacgcat ataatagatt
     4681 atattactaa ttaattgggg accctagagg tccccttttt tattttaaaa attttttcac
     4741 aaaacggttt acaagcataa ataaaatggc gccacaggtg cggttgctgg cgcctatatc
     4801 gccgacatca ccgatgggag atcctaagat atcgcttagg ccacacgttc aagtgcagcc
     4861 acaggataaa tttgcactga gcctgggtgg gattcggact cgaccgcata gccttcagga
     4921 gtgagttttg tgcaatacca accgacgact tgaccctgcc aagcggcacc agatttcttg
     4981 cgtacgcgat cccctaagcc aaaggtggca ctcaggggaa gcgcaaactg ccctgcaacg
     5041 ggagcgttgg cttcatcgct actttgaccc atgtcgaatc cttcttgtga atctattatg
     5101 gcgacaagca aatcgagctc tgactgccta ccccacaaca actactagaa agcaccagca
     5161 caacggctgc ctaactttgt tttagggcga ctaccctgct gcgtaacatc gttgctgttc
     5221 cataacatca aacatcgacc cacggcgtaa cgcgcttagc tagcttggat gcccgaggct
     5281 agactgtaca aaaaaacagt cataacaagc catgaaaacc gccactgcgc cgttaccacc
     5341 gctgcgttcg gtcaaggttc tggaccagtt gcgtgagcgc atacgctact tgcattacag
     5401 tttacgaacc gaacaggctt atgtcaactg ggttcgtgcg agctcatcga tatgcattta
     5461 atctttctgc gaattgagat gacgccactg gctgggcgtc atcccggttt cccgggtaaa
     5521 caccaccgaa aaatagttac tatcttcaaa gccacattcg gtcgaaatat cactgattaa
     5581 caggcggcta tgctggagaa gatattgcgc atgacacact ctgacctgtc gcagatattg
     5641 attgatggtc attccagtct gctggcgaaa ttgctgacgc aaaacgcgct cactgcacga
     5701 tgcctcatca caaaatttat ccagcgcaaa gggacttttc aggctagccg ccagccgggt
     5761 aatcagctta tccagcaacg tttcgctgga tgttggcggc aacgaatcac tggtgtaacg
     5821 atggcgattc agcaacatca ccaactgccc gaacagcaac tcagccattt cgttagcaaa
     5881 cggcacatgc tgactacttt catgctcaag ctgaccgata acctgccgcg cctgcgccat
     5941 ccccatgcta cctaagcgcc agtgtggttg ccctgcgctg gcgttaaatc ccggaatcgc
     6001 cccctgccag tcaagattca gcttcagacg ctccgggcaa taaataatat tctgcaaaac
     6061 cagatcgtta acggaagcgt aggagtgttt atcgtcagca tgaatgtaaa agagatcgcc
     6121 acgggtaatg cgataagggc gatcgttgag tacatgcagg ccattaccgc gccagacaat
     6181 caccagctca caaaaatcat gtgtatgttc agcaaagaca tcttgcggat aacggtcagc
     6241 cacagcgact gcctgctggt cgctggcaaa aaaatcatct ttgagaagtt ttaactgatg
     6301 cgccaccgtg gctacctcgg ccagagaacg aagttgatta ttcgcaatat ggcgtacaaa
     6361 tacgttgaga agattcgcgt tattgcagaa agccatcccg tccctggcga atatcacgcg
     6421 gtgaccagtt aaactctcgg cgaaaaagcg tcgaaaagtg gttactgtcg ctgaatccac
     6481 agcgataggc gatgtcagta acgctggcct cgctgtggcg tagcagatgt cgggctttca
     6541 tcagtcgcag gcggttcagg tatcgctgag gcgtcagtcc cgtttgctgc ttaagctgcc
     6601 gatgtagcgt acgcagtgaa agagaaaatt gatccgccac ggcatcccaa ttcacctcat
     6661 cggcaaaatg gtcctccagc caggccagaa gcaagttgag acgtgatgcg ctgttttcca
     6721 ggttctcctg caaactgctt ttacgcagca agagcagtaa ttgcataaac aagatctcgc
     6781 gactggcggt cgagggtaaa tcattttccc cttcctgctg ttccatctgt gcaaccagct
     6841 gtcgcacctg ctgcaatacg ctgtggttaa cgcgccagtg agacggatac tgcccatcca
     6901 gctcttgtgg cagcaactga ttcagcccgg cgagaaactg aaatcgatcc ggcgagcgat
     6961 acagcacatt ggtcagacac agattatcgg tatgttcata cagatgccga tcatgatcgc
     7021 gtacgaaaca gaccgtgcca ccggtgatgg tatagggctg cccattaaac acatgaatac
     7081 ccgtgccatg ttcgacaatc acaatttcat gaaaatcatg atgatgttca ggaaaatccg
     7141 cctgcgggag ccggggttct atcgccacgg acgcgttacc agacggaaaa aaatccacac
     7201 tatgtaatac ggtcatactg gcctcctgat gtcgtcaaca cggcgaaata gtaatcacga
     7261 ggtcaggttc ttaccttaaa ttttcgacgg aaaaccacgt aaaaaacgtc gatttttcaa
     7321 gatacagcgt gaattttcag gaaatgcggt gagcatcaca tcaccacaat tcagcaaatt
     7381 gtgaacatca tcacgttcat ctttccctgg ttgccaatgg cccattttcc tgtcagtaac
     7441 gagaaggtcg cgaattcagg cgctttttag actggtcgta atgaaattca gcaggatcac
     7501 attatggata ttaatactga aactgagatc aagcaaaagc attcactaac cccctttcct
     7561 gttttcctaa tcagcccggc atttcgcggg cgatattttc acagctattt caggagttca
     7621 gccatgaacg cttattacat tcaggatcgt cttgaggctc agagctgggc gcgtcactac
     7681 cagcagctcg cccgtgaaga gaaagaggca gaactggcag acgacatgga aaaaggcctg
     7741 ccccagcacc tgtttgaatc gctatgcatc gatcatttgc aacgccacgg ggccagcaaa
     7801 aaatccatta cccgtgcgtt tgatgacgat gttgagtttc aggagcgcat ggcagaacac
     7861 atccggtaca tggttgaaac cattgctcac caccaggttg atattgattc agaggtataa
     7921 aacgaatgag tactgcactc gcaacgctgg ctgggaagct ggctgaacgt gtcggcatgg
     7981 attctgtcga cccacaggaa ctgatcacca ctcttcgcca gacggcattt aaaggtgatg
     8041 ccagcgatgc gcagttcatc gcattactga tcgttgccaa ccagtacggc cttaatccgt
     8101 ggacgaaaga aatttacgcc tttcctgata agcagaatgg catcgttccg gtggtgggcg
     8161 ttgatggctg gtcccgcatc atcaatgaaa accagcagtt tgatggcatg gactttgagc
     8221 aggacaatga atcctgtaca tgccggattt accgcaagga ccgtaatcat ccgatctgcg
     8281 ttaccgaatg gatggatgaa tgccgccgcg aaccattcaa aactcgcgaa ggcagagaaa
     8341 tcacggggcc gtggcagtcg catcccaaac ggatgttacg tcataaagcc atgattcagt
     8401 gtgcccgtct ggccttcgga tttgctggta tctatgacaa ggatgaagcc gagcgcattg
     8461 tcgaaaatac tgcatacact gcagaacgtc agccggaacg cgacatcact ccggttaacg
     8521 atgaaaccat gcaggagatt aacactctgc tgatcgccct ggataaaaca tgggatgacg
     8581 acttattgcc gctctgttcc cagatatttc gccgcgacat tcgtgcatcg tcagaactga
     8641 cacaggccga agcagtaaaa gctcttggat tcctgaaaca gaaagccgca gagcagaagg
     8701 tggcagcatg acaccggaca ttatcctgca gcgtaccggg atcgatgtga gagctgtcga
     8761 acagggggat gatgcgtggc acaaattacg gctcggcgtc atcaccgctt cagaagttca
     8821 caacgtgata gcaaaacccc gctccggaaa gaagtggcct gacatgaaaa tgtcctactt
     8881 ccacaccctg cttgctgagg tttgcaccgg tgtggctccg gaagttaacg ctaaagcact
     8941 ggcctgggga aaacagtacg agaacgacgc cagaaccctg tttgaattca cttccggcgt
     9001 gaatgttact gaatccccga tcatctatcg cgacgaaagt atgcgtaccg cctgctctcc
     9061 cgatggttta tgcagtgacg gcaacggcct tgaactgaaa tgcccgttta cctcccggga
     9121 tttcatgaag ttccggctcg gtggtttcga ggccataaag tcagcttaca tggcccaggt
     9181 gcagtacagc atgtgggtga cgcgaaaaaa tgcctggtac tttgccaact atgacccgcg
     9241 tatgaagcgt gaaggcctgc attatgtcgt gattgagcgg gatgaaaagt acatggcgag
     9301 ttttgacgag atcgtgccgg agttcatcga aaaaatggac gaggcactgg ctgaaattgg
     9361 ttttgtattt ggggagcaat ggcgatagat cctagcaagg aggaaatcat catggctatc
     9421 gacgaaaaca aacagaaagc gttggcggca gcactgggcc agattgagaa acaatttggt
     9481 aaaggctcca tcatgcgcct gggtgaagac cgttccatgg atgtggaaac catctctacc
     9541 ggttcgcttt cactggatat cgcgcttggg gcaggtggtc tgccgatggg ccgtatcgtc
     9601 gaaatctacg gaccggaatc ttccggtaaa accacgctga cgctgcaggt gatcgccgca
     9661 gcgcagcgtg aaggtaaaac ctgtgcgttt atcgatgctg aacacgcgct ggacccaatc
     9721 tacgcacgta aactgggcgt cgatatcgac aacctgctgt gctcccagcc ggacaccggc
     9781 gagcaggcac tggaaatctg tgacgccctg gcgcgttctg gcgcagtaga cgttatcgtc
     9841 gttgactccg tggcggcact gacgccgaaa gcggaaatcg aaggcgaaat cggcgactct
     9901 cacatgggcc ttgcggcacg tatgatgagc caggcgatgc gtaagctggc gggtaacctg
     9961 aagcagtcca acacgctgct gatcttcatc aaccagatcc gtatgaaaat tggtgtgatg
    10021 ttcggtaacc cggaaaccac taccggtggt aacgcgctga aattctacgc ctctgttcgt
    10081 ctcgacatcc gtcgtatcgg cgcggtgaaa gagggcgaaa acgtggtggg tagcgaaacc
    10141 cgcgtgaaag tggtgaagaa caaaatcgct gcgccgttta aacaggctga attccagatc
    10201 ctctacggcg aaggtatcaa cttctacggc gaactggttg acctgggcgt aaaagagaag
    10261 ctgatcgaga aagcaggcgc gtggtacagc tacaaaggtg agaagatcgg tcagggtaaa
    10321 gcgaatgcga ctgcctggct gaaagataac ccggaaaccg cgaaagagat cgagaagaaa
    10381 gtacgtgagt tgctgctgag caacccgaac tcaacgccgg atttctctgt agatgatagc
    10441 gaaggcgtag cagaaactaa cgaagatttt taagctagga tctcgagaag cttgacctgt
    10501 gaagtgaaaa atggcgcaca ttgtgcgaca ttttttttgt ctgccgttta ccgctactgc
    10561 gtcacggatc tccacgcgcc ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg
    10621 cgcagcgtga ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct
    10681 tcctttctcg ccacgttcgc cggctttccc cgtcaagctc taaatcgggg gctcccttta
    10741 gggttccgat ttagtgcttt acggcacctc gaccccaaaa aacttgatta gggtgatggt
    10801 tcacgtagtg ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg
    10861 ttctttaata gtggactctt gttccaaact ggaacaacac tcaaccctat ctcggtctat
    10921 tcttttgatt tataagggat tttgccgatt tcggcctatt ggttaaaaaa tgagctgatt
    10981 taacaaaaat ttaacgcgaa ttttaacaaa atattaacgc ttacaatttc aggtggcact
    11041 tttcggggaa atgtgcgcgg aacccctatt tgtttatttt tctaaataca ttcaaatatg
    11101 tatccgctca tgagacaata accctgataa atgcttcaat aatattgaaa aaggaagagt
//

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

P4022/pCMV-SPORT6-CALCOCO2人源基因质粒

P4023/pCMV-SPORT6-BCL2A1人源基因质粒

P4024/pCMV-SPORT6-TDP2人源基因质粒

P4025/pCMV-SPORT6-HSPA8（1同义突变）人源基因质粒

P4026/pCMV-SPORT6-DYNC1I2人源基因质粒

P4027/pCMV-SPORT6-STYX人源基因质粒

P4028/pCMV-SPORT6-SUB1人源基因质粒

P4029/pCMV-SPORT6-KPNA1人源基因质粒

P4030/pCMV-SPORT6-PSMD14人源基因质粒

P4031/pCMV-SPORT6-XAGE1D人源基因质粒

P4032/pCMV-SPORT6-OTUB2人源基因质粒