质粒名称: | pBABE-puro mCherry-EGFP-LC3B |
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出品公司: | 爱迪基因 |
目录编号: | 22418 |
存储实验室: | Jayanta Debnath |
载体骨架基本信息 |
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载体名称: | pBABE-puro |
载体出品公司: | -- |
载体抗性: | 氨苄青霉素 |
空载体大小: | 5169 bp |
完整质粒大小: | -- |
载体修饰: | -- |
载体类型: | 哺乳动物表达,逆转录病毒 |
筛选标记: | 嘌呤霉素 |
高拷贝/低拷贝: | 高拷贝 |
生长条件: | 37 ℃ |
插入基因信息 |
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插入基因名称: | microtubule-associated protein 1 light chain 3 beta,MAP1LC3B,LC3B,LC3 |
物种来源: | H. sapiens (human) |
插入大小: | 1900 bp |
基因突变: | -- |
GenBank编号 | BC045759 |
启动子 | -- |
标签/融合蛋白: |
mCherry(插入件上的N端子)
eGFP(插入件上的N端子)
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克隆方法: | 限制性内切酶 |
5' 克隆位点: | BamHI(克隆过程中销毁) |
3' 克隆位点: | SalI (未破坏) |
5' 测序引物 | -- |
3' 测序引物 | -- |
文献引用方法 |
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材料和方法部分: | pBABE-puro mCherry-EGFP-LC3B was a gift from Jayanta Debnath (Addgene plasmid # 22418 ; http://n2t.net/addgene:22418 ; RRID:Addgene_22418) |
参考文献部分: | PLIC proteins or ubiquilins regulate autophagy-dependent cell survival during nutrient starvation. N'Diaye EN, Kajihara KK, Hsieh I, Morisaki H, Debnath J, Brown EJ. EMBO Rep. 2009 Feb;10(2):173-9. doi: 10.1038/embor.2008.238. Epub 2009 Jan 16. 10.1038/embor.2008.238 PubMed 19148225 |
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载体基本信息
载体图谱质粒图谱和多克隆位点信息
载体简介
载体序列
LOCUS pBABE-puro mCherry-EGFP-LC3B 6952 bp ds-DNA circular SYN 06-AUG-2018 DEFINITION retroviral expression of mCherry, EGFP-tagged LC3. ACCESSION . VERSION . KEYWORDS pBABE-puro mCherry-EGFP-LC3B SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 6952) AUTHORS N'Diaye EN, Kajihara KK, Hsieh I, Morisaki H, Debnath J, Brown EJ TITLE PLIC proteins or ubiquilins regulate autophagy-dependent cell survival during nutrient starvation. JOURNAL EMBO Rep. 2009 Feb;10(2):173-9. doi: 10.1038/embor.2008.238. Epub 2009 Jan 16. PUBMED 19148225 REFERENCE 2 (bases 1 to 6952) AUTHORS . TITLE Direct Submission JOURNAL Exported Aug 6, 2018 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..6952 /organism="synthetic DNA construct" /mol_type="other DNA" LTR 1..470 /note="long terminal repeat from Moloney murine leukemia virus" misc_feature 535..734 /label=MMLV Psi /note="packaging signal of Moloney murine leukemia virus (MMLV)" CDS 935..1351 /label=gag (truncated) /note="truncated Moloney murine leukemia virus (MMLV) gag gene lacking the start codon" primer_bind 1281..1303 /label=pLXSN 5' /note="Murine stem cell virus, forward primer. Also called MSCV" primer_bind 1319..1335 /label=pBABE 5' /note="Psi packaging signal, forward primer for pBABE vectors" regulatory 1362..1371 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" CDS 1368..2075 /codon_start=1 /product="monomeric derivative of DsRed fluorescent protein (Shaner et al., 2004)" /label=mCherry /note="mammalian codon-optimized" /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA EGRHSTGGMDELYK" primer_bind complement(1513..1531) /label=mCherry-R /note="mCherry, reverse primer" primer_bind 1781..1800 /label=mCherry-F /note="mCherry, forward primer" protein_bind 2103..2127 /gene="mutant version of attB" /label=attB1 /bound_moiety="BP Clonase(TM)" /note="recombination site for the Gateway(R) BP reaction" regulatory 2136..2145 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" CDS 2142..2858 /codon_start=1 /product="enhanced GFP" /label=EGFP /note="mammalian codon-optimized" /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL EFVTAAGITLGMDELYK" primer_bind complement(2187..2208) /label=EGFP-N /note="EGFP, reverse primer" primer_bind complement(2448..2467) /label=EXFP-R /note="For distinguishing EGFP variants, reverse primer" primer_bind 2795..2816 /label=EGFP-C /note="EGFP, forward primer" CDS 2886..3260 /codon_start=1 /gene="MAP1LC3B" /product="human LC3 protein" /label=LC3B /note="Atg8 family member found on autophagosomes" /translation="MPSEKTFKQRRTFEQRVEDVRLIREQHPTKIPVIIERYKGEKQLP VLDKTKFLVPDHVNMSELIKIIRRRLQLNANQAFFLLVNGHSMVSVSTPISEVYESEKD EDGFLYMVYASQETFGMKLSV" primer_bind complement(3275..3295) /label=pBABE 3' /note="SV40 enhancer, reverse primer for pBABE vectors" promoter 3280..3609 /label=SV40 promoter /note="SV40 enhancer and early promoter" rep_origin 3460..3595 /label=SV40 ori /note="SV40 origin of replication" primer_bind 3522..3541 /label=SV40pro-F /note="SV40 promoter/origin, forward primer" CDS 3619..4218 /codon_start=1 /gene="pac from Streptomyces alboniger" /product="puromycin N-acetyltransferase" /label=PuroR /note="confers resistance to puromycin" /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA" primer_bind complement(3619..3638) /label=Puro-R /note="Puromycin resistance gene, reverse primer. Also called puro-variant-R" primer_bind 4115..4135 /label=Puro-F /note="Puromycin resistance gene, forward primer" LTR 4439..4908 /note="long terminal repeat from Moloney murine leukemia virus" primer_bind complement(5038..5055) /label=L4440 /note="L4440 vector, forward primer" rep_origin complement(5209..5797) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(5289..5308) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS complement(5957..6817) /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" primer_bind 6580..6599 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(6818..6922) /gene="bla" /label=AmpR promoter ORIGIN 1 ctgaatatgg gccaaacagg atatctgtgg taagcagttc ctgccccggc tcagggccaa 61 gaacagatgg aacagctgaa tatgggccaa acaggatatc tgtggtaagc agttcctgcc 121 ccggctcagg gccaagaaca gatggtcccc agatgcggtc cagccctcag cagtttctag 181 agaaccatca gatgtttcca gggtgcccca aggacctgaa atgaccctgt gccttatttg 241 aactaaccaa tcagttcgct tctcgcttct gttcgcgcgc ttctgctccc cgagctcaat 301 aaaagagccc acaacccctc actcggggcg ccagtcctcc gattgactga gtcgcccggg 361 tacccgtgta tccaataaac cctcttgcag ttgcatccga cttgtggtct cgctgttcct 421 tgggagggtc tcctctgagt gattgactac ccgtcagcgg gggtctttca tttgggggct 481 cgtccgggat cgggagaccc ctgcccaggg accaccgacc caccaccggg aggcaagctg 541 gccagcaact tatctgtgtc tgtccgattg tctagtgtct atgactgatt ttatgcgcct 601 gcgtcggtac tagttagcta actagctctg tatctggcgg acccgtggtg gaactgacga 661 gttctgaaca cccggccgca accctgggag acgtcccagg gactttgggg gccgtttttg 721 tggcccgacc tgaggaaggg agtcgatgtg gaatccgacc ccgtcaggat atgtggttct 781 ggtaggagac gagaacctaa aacagttccc gcctccgtct gaatttttgc tttcggtttg 841 gaaccgaagc cgcgcgtctt gtctgctgca gcgctgcagc atcgttctgt gttgtctctg 901 tctgactgtg tttctgtatt tgtctgaaaa ttagggccag actgttacca ctcccttaag 961 tttgacctta ggtcactgga aagatgtcga gcggatcgct cacaaccagt cggtagatgt 1021 caagaagaga cgttgggtta ccttctgctc tgcagaatgg ccaaccttta acgtcggatg 1081 gccgcgagac ggcaccttta accgagacct catcacccag gttaagatca aggtcttttc 1141 acctggcccg catggacacc cagaccaggt cccctacatc gtgacctggg aagccttggc 1201 ttttgacccc cctccctggg tcaagccctt tgtacaccct aagcctccgc ctcctcttcc 1261 tccatccgcc ccgtctctcc cccttgaacc tcctcgttcg accccgcctc gatcctccct 1321 ttatccagcc ctcactcctt ctctaggcgc cggccggatc tgccaccatg gtgagcaagg 1381 gcgaggagga taacatggcc atcatcaagg agttcatgcg cttcaaggtg cacatggagg 1441 gctccgtgaa cggccacgag ttcgagatcg agggcgaggg cgagggccgc ccctacgagg 1501 gcacccagac cgccaagctg aaggtgacca agggtggccc cctgcccttc gcctgggaca 1561 tcctgtcccc tcagttcatg tacggctcca aggcctacgt gaagcacccc gccgacatcc 1621 ccgactactt gaagctgtcc ttccccgagg gcttcaagtg ggagcgcgtg atgaacttcg 1681 aggacggcgg cgtggtgacc gtgacccagg actcctccct gcaggacggc gagttcatct 1741 acaaggtgaa gctgcgcggc accaacttcc cctccgacgg ccccgtaatg cagaagaaga 1801 ccatgggctg ggaggcctcc tccgagcgga tgtaccccga ggacggcgcc ctgaagggcg 1861 agatcaagca gaggctgaag ctgaaggacg gcggccacta cgacgctgag gtcaagacca 1921 cctacaaggc caagaagccc gtgcagctgc ccggcgccta caacgtcaac atcaagttgg 1981 acatcacctc ccacaacgag gactacacca tcgtggaaca gtacgaacgc gccgagggcc 2041 gccactccac cggcggcatg gacgagctgt acaagtccgg actcagatcc gctggcccag 2101 gaacaagttt gtacaaaaaa gcaggctttc cggtcgccac catggtgagc aagggcgagg 2161 agctgttcac cggggtggtg cccatcctgg tcgagctgga cggcgacgta aacggccaca 2221 agttcagcgt gtccggcgag ggcgagggcg atgccaccta cggcaagctg accctgaagt 2281 tcatctgcac caccggcaag ctgcccgtgc cctggcccac cctcgtgacc accctgacct 2341 acggcgtgca gtgcttcagc cgctaccccg accacatgaa gcagcacgac ttcttcaagt 2401 ccgccatgcc cgaaggctac gtccaggagc gcaccatctt cttcaaggac gacggcaact 2461 acaagacccg cgccgaggtg aagttcgagg gcgacaccct ggtgaaccgc atcgagctga 2521 agggcatcga cttcaaggag gacggcaaca tcctggggca caagctggag tacaactaca 2581 acagccacaa cgtctatatc atggccgaca agcagaagaa cggcatcaag gtgaacttca 2641 agatccgcca caacatcgag gacggcagcg tgcagctcgc cgaccactac cagcagaaca 2701 cccccatcgg cgacggcccc gtgctgctgc ccgacaacca ctacctgagc acccagtccg 2761 ccctgagcaa agaccccaac gagaagcgcg atcacatggt cctgctggag ttcgtgaccg 2821 ccgccgggat cactctcggc atggacgagc tgtacaagtc cggactcaga tccggtaccg 2881 aattcatgcc gtcggagaag accttcaagc agcgccgcac cttcgaacaa agagtagaag 2941 atgtccgact tattcgagag cagcatccaa ccaaaatccc ggtgataata gaacgataca 3001 agggtgagaa gcagcttcct gttctggata aaacaaagtt ccttgtacct gaccatgtca 3061 acatgagtga gctcatcaag ataattagaa ggcgcttaca gctcaatgct aatcaggcct 3121 tcttcctgtt ggtgaacgga cacagcatgg tcagcgtctc cacaccaatc tcagaggtgt 3181 atgagagtga gaaagatgaa gatggattcc tgtacatggt ctatgcctcc caggagacgt 3241 tcgggatgaa attgtcagtg taagtcgacc ctgtggaatg tgtgtcagtt agggtgtgga 3301 aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa ttagtcagca 3361 accaggtgtg gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag catgcatctc 3421 aattagtcag caaccatagt cccgccccta actccgccca tcccgcccct aactccgccc 3481 agttccgccc attctccgcc ccatggctga ctaatttttt ttatttatgc agaggccgag 3541 gccgcctcgg cctctgagct attccagaag tagtgaggag gcttttttgg aggcctaggc 3601 ttttgcaaaa agcttaccat gaccgagtac aagcccacgg tgcgcctcgc cacccgcgac 3661 gacgtcccca gggccgtacg caccctcgcc gccgcgttcg ccgactaccc cgccacgcgc 3721 cacaccgtcg atccggaccg ccacatcgag cgggtcaccg agctgcaaga actcttcctc 3781 acgcgcgtcg ggctcgacat cggcaaggtg tgggtcgcgg acgacggcgc cgcggtggcg 3841 gtctggacca cgccggagag cgtcgaagcg ggggcggtgt tcgccgagat cggcccgcgc 3901 atggccgagt tgagcggttc ccggctggcc gcgcagcaac agatggaagg cctcctggcg 3961 ccgcaccggc ccaaggagcc cgcgtggttc ctggccaccg tcggcgtctc gcccgaccac 4021 cagggcaagg gtctgggcag cgccgtcgtg ctccccggag tggaggcggc cgagcgcgcc 4081 ggggtgcccg ccttcctgga gacctccgcg ccccgcaacc tccccttcta cgagcggctc 4141 ggcttcaccg tcaccgccga cgtcgaggtg cccgaaggac cgcgcacctg gtgcatgacc 4201 cgcaagcccg gtgcctgacg cccgccccac gacccgcagc gcccgaccga aaggagcgca 4261 cgaccccatg catcgataaa ataaaagatt ttatttagtc tccagaaaaa ggggggaatg 4321 aaagacccca cctgtaggtt tggcaagcta gcttaagtaa cgccattttg caaggcatgg 4381 aaaaatacat aactgagaat agagaagttc agatcaaggt caggaacaga tggaacagct 4441 gaatatgggc caaacaggat atctgtggta agcagttcct gccccggctc agggccaaga 4501 acagatggaa cagctgaata tgggccaaac aggatatctg tggtaagcag ttcctgcccc 4561 ggctcagggc caagaacaga tggtccccag atgcggtcca gccctcagca gtttctagag 4621 aaccatcaga tgtttccagg gtgccccaag gacctgaaat gaccctgtgc cttatttgaa 4681 ctaaccaatc agttcgcttc tcgcttctgt tcgcgcgctt ctgctccccg agctcaataa 4741 aagagcccac aacccctcac tcggggcgcc agtcctccga ttgactgagt cgcccgggta 4801 cccgtgtatc caataaaccc tcttgcagtt gcatccgact tgtggtctcg ctgttccttg 4861 ggagggtctc ctctgagtga ttgactaccc gtcagcgggg gtctttcaca tgcagcatgt 4921 atcaaaatta atttggtttt ttttcttaag tatttacatt aaatggccat agttgcatta 4981 atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt gggcgctctt ccgcttcctc 5041 gctcactgac tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa 5101 ggcggtaata cggttatcca cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa 5161 aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt tccataggct 5221 ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc gaaacccgac 5281 aggactataa agataccagg cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc 5341 gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg tggcgctttc 5401 tcatagctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca agctgggctg 5461 tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact atcgtcttga 5521 gtccaacccg gtaagacacg acttatcgcc actggcagca gccactggta acaggattag 5581 cagagcgagg tatgtaggcg gtgctacaga gttcttgaag tggtggccta actacggcta 5641 cactagaaga acagtatttg gtatctgcgc tctgctgaag ccagttacct tcggaaaaag 5701 agttggtagc tcttgatccg gcaaacaaac caccgctggt agcggtggtt tttttgtttg 5761 caagcagcag attacgcgca gaaaaaaagg atctcaagaa gatcctttga tcttttctac 5821 ggggtctgac gctcagtgga acgaaaactc acgttaaggg attttggtca tgagattatc 5881 aaaaaggatc ttcacctaga tccttttgcg gccgcaaatc aatctaaagt atatatgagt 5941 aaacttggtc tgacagttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc 6001 tatttcgttc atccatagtt gcctgactcc ccgtcgtgta gataactacg atacgggagg 6061 gcttaccatc tggccccagt gctgcaatga taccgcgaga cccacgctca ccggctccag 6121 atttatcagc aataaaccag ccagccggaa gggccgagcg cagaagtggt cctgcaactt 6181 tatccgcctc catccagtct attaattgtt gccgggaagc tagagtaagt agttcgccag 6241 ttaatagttt gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt 6301 ttggtatggc ttcattcagc tccggttccc aacgatcaag gcgagttaca tgatccccca 6361 tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat cgttgtcaga agtaagttgg 6421 ccgcagtgtt atcactcatg gttatggcag cactgcataa ttctcttact gtcatgccat 6481 ccgtaagatg cttttctgtg actggtgagt actcaaccaa gtcattctga gaatagtgta 6541 tgcggcgacc gagttgctct tgcccggcgt caatacggga taataccgcg ccacatagca 6601 gaactttaaa agtgctcatc attggaaaac gttcttcggg gcgaaaactc tcaaggatct 6661 taccgctgtt gagatccagt tcgatgtaac ccactcgtgc acccaactga tcttcagcat 6721 cttttacttt caccagcgtt tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa 6781 agggaataag ggcgacacgg aaatgttgaa tactcatact cttccttttt caatattatt 6841 gaagcattta tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa 6901 ataaacaaat aggggttccg cgcacatttc cccgaaaagt gccacctgca gc //