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pmCherry-EGFP-LC3B-h,Plasmid pmCherry-EGFP-LC3B-h,pmCherry-EGFP-LC3B-h vector
pmCherry-EGFP-LC3B-h Information
Alias: mRFP-GFP-LC3; MAP1LC3B; LC3B; ATG8F;
Promoter: CMV
Replicon: pUC ori, F1 ori, SV40 ori
Terminator: SV40 poly (A) signal
Plasmid classification: mammalian cells, autophagic expression vectors
Plasmid size: 5833 BP
Prokaryotic resistance: kanamycin
Screening Marker: Hereditary Mycin G418
Cloning strain: DH5alpha Escherichia coli
Culture conditions: 37鈩? aerobic, LB
Expression host: mammalian cells
Induction mode: no induction, transient expression
5'Sequencing Primers: CMV-F: CGCAAATGGGGGGGTAGGGGGGTG
3'Sequencing primers: Sv40-polyA-R: GAAATTTGATGCTATTGC
pmCherry-EGFP-LC3B-h Description
PmCherry-EGFP-LC3b is a human mammalian autophagy double fluorescent reporter plasmid.
pmCherry-EGFP-LC3B-h Sequence
LOCUS Exported 5833 bp ds-DNA circular SYN 12-MAY-2016
FEATURES Location/Qualifiers
source 1..5833
/organism="synthetic DNA construct"
/mol_type="other DNA"
enhancer 61..364
/note="CMV enhancer"
/note="human cytomegalovirus immediate early enhancer"
promoter 365..568
/note="CMV promoter"
/note="human cytomegalovirus (CMV) immediate early
promoter"
CDS 604..1311
/codon_start=1
/product="monomeric derivative of DsRed fluorescent protein
(Shaner et al., 2004)"
/note="mCherry"
/note="mammalian codon-optimized"
/translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
EGRHSTGGMDELYK"
CDS 1333..2049
/codon_start=1
/product="enhanced GFP"
/note="EGFP"
/note="mammalian codon-optimized"
/translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
EFVTAAGITLGMDELYK"
misc_feature 2083..2457
/note="LC3B-h"
polyA_signal 2621..2742
/note="SV40 poly(A) signal"
/note="SV40 polyadenylation signal"
rep_origin complement(2749..3204)
/direction=LEFT
/note="f1 ori"
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
promoter 3231..3335
/gene="bla"
/note="AmpR promoter"
promoter 3337..3694
/note="SV40 promoter"
/note="SV40 enhancer and early promoter"
rep_origin 3545..3680
/note="SV40 ori"
/note="SV40 origin of replication"
CDS 3729..4523
/codon_start=1
/gene="aph(3')-II (or nptII)"
/product="aminoglycoside phosphotransferase from Tn5"
/note="NeoR/KanR"
/note="confers resistance to neomycin, kanamycin, and G418
(Geneticin(R))"
/translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
polyA_signal 4755..4802
/note="HSV TK poly(A) signal"
/note="herpes simplex virus thymidine kinase
polyadenylation signal (Cole and Stacy, 1985)"
rep_origin 5131..5719
/direction=RIGHT
/note="ori"
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
ORIGIN
1 tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg
61 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
121 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
181 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
241 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
301 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
361 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
421 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
481 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
541 acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatcc gctagccgcc
601 accatggtga gcaagggcga ggaggataac atggccatca tcaaggagtt catgcgcttc
661 aaggtgcaca tggagggctc cgtgaacggc cacgagttcg agatcgaggg cgagggcgag
721 ggccgcccct acgagggcac ccagaccgcc aagctgaagg tgaccaaggg tggccccctg
781 cccttcgcct gggacatcct gtcccctcag ttcatgtacg gctccaaggc ctacgtgaag
841 caccccgccg acatccccga ctacttgaag ctgtccttcc ccgagggctt caagtgggag
901 cgcgtgatga acttcgagga cggcggcgtg gtgaccgtga cccaggactc ctccctgcag
961 gacggcgagt tcatctacaa ggtgaagctg cgcggcacca acttcccctc cgacggcccc
1021 gtaatgcaga agaagaccat gggctgggag gcctcctccg agcggatgta ccccgaggac
1081 ggcgccctga agggcgagat caagcagagg ctgaagctga aggacggcgg ccactacgac
1141 gctgaggtca agaccaccta caaggccaag aagcccgtgc agctgcccgg cgcctacaac
1201 gtcaacatca agttggacat cacctcccac aacgaggact acaccatcgt ggaacagtac
1261 gaacgcgccg agggccgcca ctccaccggc ggcatggacg agctgtacaa gtccggactc
1321 agatctcgag gcatggtgag caagggcgag gagctgttca ccggggtggt gcccatcctg
1381 gtcgagctgg acggcgacgt aaacggccac aagttcagcg tgtccggcga gggcgagggc
1441 gatgccacct acggcaagct gaccctgaag ttcatctgca ccaccggcaa gctgcccgtg
1501 ccctggccca ccctcgtgac caccctgacc tacggcgtgc agtgcttcag ccgctacccc
1561 gaccacatga agcagcacga cttcttcaag tccgccatgc ccgaaggcta cgtccaggag
1621 cgcaccatct tcttcaagga cgacggcaac tacaagaccc gcgccgaggt gaagttcgag
1681 ggcgacaccc tggtgaaccg catcgagctg aagggcatcg acttcaagga ggacggcaac
1741 atcctggggc acaagctgga gtacaactac aacagccaca acgtctatat catggccgac
1801 aagcagaaga acggcatcaa ggtgaacttc aagatccgcc acaacatcga ggacggcagc
1861 gtgcagctcg ccgaccacta ccagcagaac acccccatcg gcgacggccc cgtgctgctg
1921 cccgacaacc actacctgag cacccagtcc gccctgagca aagaccccaa cgagaagcgc
1981 gatcacatgg tcctgctgga gttcgtgacc gccgccggga tcactctcgg catggacgag
2041 ctgtacaagt actcagatct cgagctcaag cttcgaattc ccatgccgtc ggagaagacc
2101 ttcaagcagc gccgcacctt cgaacaaaga gtagaagatg tccgacttat tcgagagcag
2161 catccaacca aaatcccggt gataatagaa cgatacaagg gtgagaagca gcttcctgtt
2221 ctggataaaa caaagttcct tgtacctgac catgtcaaca tgagtgagct catcaagata
2281 attagaaggc gcttacagct caatgctaat caggccttct tcctgttggt gaacggacac
2341 agcatggtca gcgtctccac accaatctca gaggtgtatg agagtgagaa agatgaagat
2401 ggattcctgt acatggtcta tgcctcccag gagacgttcg ggatgaaatt gtcagtgtaa
2461 gaattctgca gtcgacggta ccgcgggccc gggatccacc ggatctagat aactgatcat
2521 aatcagccat accacatttg tagaggtttt acttgcttta aaaaacctcc cacacctccc
2581 cctgaacctg aaacataaaa tgaatgcaat tgttgttgtt aacttgttta ttgcagctta
2641 taatggttac aaataaagca atagcatcac aaatttcaca aataaagcat ttttttcact
2701 gcattctagt tgtggtttgt ccaaactcat caatgtatct taacgcgtaa attgtaagcg
2761 ttaatatttt gttaaaattc gcgttaaatt tttgttaaat cagctcattt tttaaccaat
2821 aggccgaaat cggcaaaatc ccttataaat caaaagaata gaccgagata gggttgagtg
2881 ttgttccagt ttggaacaag agtccactat taaagaacgt ggactccaac gtcaaagggc
2941 gaaaaaccgt ctatcagggc gatggcccac tacgtgaacc atcaccctaa tcaagttttt
3001 tggggtcgag gtgccgtaaa gcactaaatc ggaaccctaa agggagcccc cgatttagag
3061 cttgacgggg aaagccggcg aacgtggcga gaaaggaagg gaagaaagcg aaaggagcgg
3121 gcgctagggc gctggcaagt gtagcggtca cgctgcgcgt aaccaccaca cccgccgcgc
3181 ttaatgcgcc gctacagggc gcgtcaggtg gcacttttcg gggaaatgtg cgcggaaccc
3241 ctatttgttt atttttctaa atacattcaa atatgtatcc gctcatgaga caataaccct
3301 gataaatgct tcaataatat tgaaaaagga agagtcctga ggcggaaaga accagctgtg
3361 gaatgtgtgt cagttagggt gtggaaagtc cccaggctcc ccagcaggca gaagtatgca
3421 aagcatgcat ctcaattagt cagcaaccag gtgtggaaag tccccaggct ccccagcagg
3481 cagaagtatg caaagcatgc atctcaatta gtcagcaacc atagtcccgc ccctaactcc
3541 gcccatcccg cccctaactc cgcccagttc cgcccattct ccgccccatg gctgactaat
3601 tttttttatt tatgcagagg ccgaggccgc ctcggcctct gagctattcc agaagtagtg
3661 aggaggcttt tttggaggcc taggcttttg caaagatcga tcaagagaca ggatgaggat
3721 cgtttcgcat gattgaacaa gatggattgc acgcaggttc tccggccgct tgggtggaga
3781 ggctattcgg ctatgactgg gcacaacaga caatcggctg ctctgatgcc gccgtgttcc
3841 ggctgtcagc gcaggggcgc ccggttcttt ttgtcaagac cgacctgtcc ggtgccctga
3901 atgaactgca agacgaggca gcgcggctat cgtggctggc cacgacgggc gttccttgcg
3961 cagctgtgct cgacgttgtc actgaagcgg gaagggactg gctgctattg ggcgaagtgc
4021 cggggcagga tctcctgtca tctcaccttg ctcctgccga gaaagtatcc atcatggctg
4081 atgcaatgcg gcggctgcat acgcttgatc cggctacctg cccattcgac caccaagcga
4141 aacatcgcat cgagcgagca cgtactcgga tggaagccgg tcttgtcgat caggatgatc
4201 tggacgaaga gcatcagggg ctcgcgccag ccgaactgtt cgccaggctc aaggcgagca
4261 tgcccgacgg cgaggatctc gtcgtgaccc atggcgatgc ctgcttgccg aatatcatgg
4321 tggaaaatgg ccgcttttct ggattcatcg actgtggccg gctgggtgtg gcggaccgct
4381 atcaggacat agcgttggct acccgtgata ttgctgaaga gcttggcggc gaatgggctg
4441 accgcttcct cgtgctttac ggtatcgccg ctcccgattc gcagcgcatc gccttctatc
4501 gccttcttga cgagttcttc tgagcgggac tctggggttc gaaatgaccg accaagcgac
4561 gcccaacctg ccatcacgag atttcgattc caccgccgcc ttctatgaaa ggttgggctt
4621 cggaatcgtt ttccgggacg ccggctggat gatcctccag cgcggggatc tcatgctgga
4681 gttcttcgcc caccctaggg ggaggctaac tgaaacacgg aaggagacaa taccggaagg
4741 aacccgcgct atgacggcaa taaaaagaca gaataaaacg cacggtgttg ggtcgtttgt
4801 tcataaacgc ggggttcggt cccagggctg gcactctgtc gataccccac cgagacccca
4861 ttggggccaa tacgcccgcg tttcttcctt ttccccaccc caccccccaa gttcgggtga
4921 aggcccaggg ctcgcagcca acgtcggggc ggcaggccct gccatagcct caggttactc
4981 atatatactt tagattgatt taaaacttca tttttaattt aaaaggatct aggtgaagat
5041 cctttttgat aatctcatga ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc
5101 agaccccgta gaaaagatca aaggatcttc ttgagatcct ttttttctgc gcgtaatctg
5161 ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt tgtttgccgg atcaagagct
5221 accaactctt tttccgaagg taactggctt cagcagagcg cagataccaa atactgtcct
5281 tctagtgtag ccgtagttag gccaccactt caagaactct gtagcaccgc ctacatacct
5341 cgctctgcta atcctgttac cagtggctgc tgccagtggc gataagtcgt gtcttaccgg
5401 gttggactca agacgatagt taccggataa ggcgcagcgg tcgggctgaa cggggggttc
5461 gtgcacacag cccagcttgg agcgaacgac ctacaccgaa ctgagatacc tacagcgtga
5521 gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg
5581 cagggtcgga acaggagagc gcacgaggga gcttccaggg ggaaacgcct ggtatcttta
5641 tagtcctgtc gggtttcgcc acctctgact tgagcgtcga tttttgtgat gctcgtcagg
5701 ggggcggagc ctatggaaaa acgccagcaa cgcggccttt ttacggttcc tggccttttg
5761 ctggcctttt gctcacatgt tctttcctgc gttatcccct gattctgtgg ataaccgtat
5821 taccgccatg cat
//
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