| 质粒名称: | pZac2.1 gfaABC1D-BioID2-BioID2-HA |
|---|---|
| 出品公司: | addgene |
| 目录编号： | 176740 |
| 存储实验室: | Baljit Khakh |
|
载体骨架基本信息 |
|
| 载体名称: | PZac2.1 |
| 载体出品公司: | Khakh lab |
| 空载体大小: | 5109bp |
| 完整质粒大小: | 6630 bp |
| 载体类型: | 细菌细胞表达、AAV |
| 载体抗性: | 氨苄霉素 |
| 生长温度: | 37 ℃ |
| 宿主菌: | NEB Stable |
| 拷贝数: | 未知 |
|
插入基因信息 |
|
| 插入基因名称: | BioID2-BioID2-HA / BioID2-HA |
| 物种来源: | Aquifex aeolicus |
| 插入大小: | 1521 bp |
| 启动子: | gfaABC1D |
| 标签/融合蛋白: | HA (C terminal on insert) |
| 克隆方法: | Ligation Independent Cloning |
| 5’克隆位点: | AgeI(未破坏) |
| 3’克隆位点: | NotI (未破坏) |
| 5’测序引物 | taatacgactcactatagg |
| 3’测序引物 | gtggtttgtccaaactcatc |
|
文献引用方法 |
|
| 材料和方法部分: | pZac2.1 gfaABC1D-BioID2-BioID2-HA was a gift from Baljit Khakh (Addgene plasmid # 176740 ; http://n2t.net/addgene:176740 ; RRID:Addgene_176740) |
| 参考文献部分: | Astrocyte-neuron subproteomes and obsessive-compulsive disorder mechanisms. Soto JS, Jami-Alahmadi Y, Chacon J, Moye SL, Diaz-Castro B, Wohlschlegel JA, Khakh BS. Nature. 2023 Apr;616(7958):764-773. doi: 10.1038/s41586-023-05927-7. Epub 2023 Apr 12. 10.1038/s41586-023-05927-7 PubMed 37046092 |
买家导航
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 pZac2.1 gfaABC1D-BioID2-BioID2-HA 6630 bp ds-DNA circular SYN 24-NOV-2021
DEFINITION Expresses the promiscuous small biotin ligase, BioID2, in
astrocytes.
ACCESSION .
VERSION .
KEYWORDS pZac2.1 gfaABC1D-BioID2-BioID2-HA
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 6630)
TITLE Bacterial expression plasmids for production of protein-fused BioID2
JOURNAL Unpublished
REFERENCE 2 (bases 1 to 6630)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported Nov 24, 2021 from SnapGene Server 1.1.58
http://www.biofeng.com
FEATURES Location/Qualifiers
source 1..6630
/organism="synthetic DNA construct"
/mol_type="other DNA"
protein_bind 107..128
/label=CAP binding site
/bound_moiety="E. coli catabolite activator protein"
/note="CAP binding activates transcription in the presence
of cAMP."
promoter 143..173
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind 181..197
/label=lac operator
/bound_moiety="lac repressor encoded by lacI"
/note="The lac repressor binds to the lac operator to
inhibit transcription in E. coli. This inhibition can be
relieved by adding lactose or
isopropyl-beta-D-thiogalactopyranoside (IPTG)."
primer_bind 186..208
/label=M13/pUC Reverse
/note="In lacZ gene"
primer_bind 205..221
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
primer_bind 205..221
/label=M13 Reverse
/note="In lacZ gene. Also called M13-rev"
repeat_region 259..388
/label=AAV2 ITR
intron 1239..1371
/label=chimeric intron
/note="chimera between introns from human beta-globin and
immunoglobulin heavy chain genes"
primer_bind 1354..1373
/label=pMT2-F
/note="Synthetic intron, forward primer"
promoter 1416..1434
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
CDS 1461..2159
/codon_start=1
/product="promiscuous R40G mutant of a biotin protein
ligase from Aquifex aeolicus (Kim et al., 2016)"
/label=BioID2
/translation="MFKNLIWLKEVDSTQERLKEWNVSYGTALVADRQTKGRGGLGRKW
LSQEGGLYFSFLLNPKEFENLLQLPLVLGLSVSEALEEITEIPFSLKWPNDVYFQEKKV
SGVLCELSKDKLIVGIGINVNQREIPEEIKDRATTLYEITGKDWDRKEVLLKVLKRISE
NLKKFKEKSFKEFKGKIESKMLYLGEEVKLLGEGKITGKLVGLSEKGGALILTEEGIKE
ILSGEFSLRRS"
CDS 2244..2942
/codon_start=1
/product="promiscuous R40G mutant of a biotin protein
ligase from Aquifex aeolicus (Kim et al., 2016)"
/label=BioID2
/translation="MFKNLIWLKEVDSTQERLKEWNVSYGTALVADRQTKGRGGLGRKW
LSQEGGLYFSFLLNPKEFENLLQLPLVLGLSVSEALEEITEIPFSLKWPNDVYFQEKKV
SGVLCELSKDKLIVGIGINVNQREIPEEIKDRATTLYEITGKDWDRKEVLLKVLKRISE
NLKKFKEKSFKEFKGKIESKMLYLGEEVKLLGEGKITGKLVGLSEKGGALILTEEGIKE
ILSGEFSLRRS"
CDS 2952..2978
/codon_start=1
/product="HA (human influenza hemagglutinin) epitope tag"
/label=HA
/translation="YPYDVPDYA"
misc_feature 3003..3591
/label=WPRE
/note="woodchuck hepatitis virus posttranscriptional
regulatory element"
primer_bind complement(3056..3076)
/label=WPRE-R
/note="WPRE, reverse primer"
CDS complement(3474..3485)
/codon_start=1
/product="Factor Xa recognition and cleavage site"
/label=Factor Xa site
/translation="IEGR"
polyA_signal 3655..3776
/label=SV40 poly(A) signal
/note="SV40 polyadenylation signal"
primer_bind complement(3666..3685)
/label=EBV-rev
/note="SV40 polyA terminator, reverse primer"
primer_bind 3720..3739
/label=SV40pA-R
/note="SV40 polyA, reverse primer"
repeat_region 3938..4067
/label=AAV2 ITR
primer_bind complement(4086..4103)
/label=M13 Forward
/note="In lacZ gene. Also called M13-F20 or M13 (-21)
Forward"
primer_bind complement(4086..4102)
/label=M13 fwd
/note="common sequencing primer, one of multiple similar
variants"
primer_bind complement(4095..4117)
/label=M13/pUC Forward
/note="In lacZ gene"
rep_origin 4244..4699
/direction=RIGHT
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
primer_bind complement(4331..4350)
/label=F1ori-R
/note="F1 origin, reverse primer"
primer_bind 4541..4562
/label=F1ori-F
/note="F1 origin, forward primer"
promoter 4725..4829
/gene="bla"
/label=AmpR promoter
CDS 4830..5690
/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 complement(5048..5067)
/label=Amp-R
/note="Ampicillin resistance gene, reverse primer"
rep_origin 5861..6449
/direction=RIGHT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind 6350..6369
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
primer_bind 6603..6620
/label=L4440
/note="L4440 vector, forward primer"
ORIGIN
1 agcgcccaat acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagctggc
61 acgacaggtt tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat gtgagttagc
121 tcactcatta ggcaccccag gctttacact ttatgcttcc ggctcgtatg ttgtgtggaa
181 ttgtgagcgg ataacaattt cacacaggaa acagctatga ccatgattac gccagattta
241 attaaggcct taattaggct gcgcgctcgc tcgctcactg aggccgcccg ggcaaagccc
301 gggcgtcggg cgacctttgg tcgcccggcc tcagtgagcg agcgagcgcg cagagaggga
361 gtggccaact ccatcactag gggttccttg tagttaatga ttaacccgcc atgctactta
421 tctacgtagc catgctctag gaagatctaa catatcctgg tgtggagtag gggacgctgc
481 tctgacagag gctcgggggc ctgagctggc tctgtgagct ggggaggagg cagacagcca
541 ggccttgtct gcaagcagac ctggcagcat tgggctggcc gccccccagg gcctcctctt
601 catgcccagt gaatgactca ccttggcaca gacacaatgt tcggggtggg cacagtgcct
661 gcttcccgcc gcaccccagc ccccctcaaa tgccttccga gaagcccatt gagcaggggg
721 cttgcattgc accccagcct gacagcctgg catcttggga taaaagcagc acagccccct
781 aggggctgcc cttgctgtgt ggcgccaccg gcggtggaga acaaggctct attcagcctg
841 tgcccaggaa aggggatcag gggatgccca ggcatggaca gtgggtggca gggggggaga
901 ggagggctgt ctgcttccca gaagtccaag gacacaaatg ggtgagggga gagctctccc
961 catagctggg ctgcggccca accccacccc ctcaggctat gccagggggt gttgccaggg
1021 gcacccgggc atcgccagtc tagcccactc cttcataaag ccctcgcatc ccaggagcga
1081 gcagagccag agcaggttgg agaggagacg catcacctcc gctgctcgca agctttattg
1141 cggtagttta tcacagttaa attgctaacg cagtcagtgc ttctgacaca acagtctcga
1201 acttaagctg cagaagttgg tcgtgaggca ctgggcaggt aagtatcaag gttacaagac
1261 aggtttaagg agaccaatag aaactgggct tgtcgagaca gagaagactc ttgcgtttct
1321 gataggcacc tattggtctt actgacatcc actttgcctt tctctccaca ggtgtccact
1381 cccagttcaa ttacagctct taaggctaga gtacttaata cgactcacta taggctagcc
1441 tcgagctcgg atccaccacc atgttcaaaa atcttatttg gctgaaggaa gttgatagca
1501 cccaggagcg gcttaaggaa tggaatgtct cctatggcac tgcactcgtc gccgatagac
1561 agaccaaagg tagaggcggt ctcggacgga agtggttgag tcaggagggc ggtctttact
1621 tctctttctt gttgaatcca aaagagtttg aaaatttgtt gcaacttcca ctggtcttgg
1681 gactctccgt gagcgaggca ctcgaagaga ttacagagat accatttagt ctgaagtggc
1741 ccaacgacgt ttattttcag gagaaaaagg tttcaggggt cctttgtgag ctcagtaagg
1801 acaagctgat agtgggaatc ggtataaacg tgaatcaaag agagatccct gaagagatca
1861 aggatcgggc cacaacactt tatgaaatca ctggaaagga ttgggacaga aaggaggtct
1921 tgctgaaggt tctcaagcga atttccgaaa atctcaagaa atttaaggaa aaatccttta
1981 aagagttcaa aggaaaaatt gagtcaaaga tgttgtacct tggagaggag gttaaactct
2041 tgggagaagg caagattacc gggaaacttg tgggcctgag tgaaaagggc ggtgctttga
2101 tacttacaga ggaaggcata aaggagatac tgtccggcga atttagcttg cgacggtcaa
2161 gcttaggtgg aggaggttct ggaggcggtg gaagtggtgg cggaggtagc ggtggaggag
2221 gttctatggt gtctaaggaa ttcatgttca aaaatcttat ttggctgaag gaagttgata
2281 gcacccagga gcggcttaag gaatggaatg tctcctatgg cactgcactc gtcgccgata
2341 gacagaccaa aggtagaggc ggtctcggac ggaagtggtt gagtcaggag ggcggtcttt
2401 acttctcttt cttgttgaat ccaaaagagt ttgaaaattt gttgcaactt ccactggtct
2461 tgggactctc cgtgagcgag gcactcgaag agattacaga gataccattt agtctgaagt
2521 ggcccaacga cgtttatttt caggagaaaa aggtttcagg ggtcctttgt gagctcagta
2581 aggacaagct gatagtggga atcggtataa acgtgaatca aagagagatc cctgaagaga
2641 tcaaggatcg ggccacaaca ctttatgaaa tcactggaaa ggattgggac agaaaggagg
2701 tcttgctgaa ggttctcaag cgaatttccg aaaatctcaa gaaatttaag gaaaaatcct
2761 ttaaagagtt caaaggaaaa attgagtcaa agatgttgta ccttggagag gaggttaaac
2821 tcttgggaga aggcaagatt accgggaaac ttgtgggcct gagtgaaaag ggcggtgctt
2881 tgatacttac agaggaaggc ataaaggaga tactgtccgg cgaatttagc ttgcgacggt
2941 cagctagcgc ttacccttac gatgtaccgg attacgcata gggatccaat tcgcttatcg
3001 ataatcaacc tctggattac aaaatttgtg aaagattgac tggtattctt aactatgttg
3061 ctccttttac gctatgtgga tacgctgctt taatgccttt gtatcatgct attgcttccc
3121 gtatggcttt cattttctcc tccttgtata aatcctggtt gctgtctctt tatgaggagt
3181 tgtggcccgt tgtcaggcaa cgtggcgtgg tgtgcactgt gtttgctgac gcaaccccca
3241 ctggttgggg cattgccacc acctgtcagc tcctttccgg gactttcgct ttccccctcc
3301 ctattgccac ggcggaactc atcgccgcct gccttgcccg ctgctggaca ggggctcggc
3361 tgttgggcac tgacaattcc gtggtgttgt cggggaaatc atcgtccttt ccttggctgc
3421 tcgcctatgt tgccacctgg attctgcgcg ggacgtcctt ctgctacgtc ccttcggccc
3481 tcaatccagc ggaccttcct tcccgcggcc tgctgccggc tctgcggcct cttccgcgtc
3541 ttcgccttcg ccctcagacg agtcggatct ccctttgggc cgcctccccg catcgatacc
3601 ggaattctgc agatatctag agtcgacccg ggcggccgct tcgagcagac atgataagat
3661 acattgatga gtttggacaa accacaacta gaatgcagtg aaaaaaatgc tttatttgtg
3721 aaatttgtga tgctattgct ttatttgtaa ccattataag ctgcaataaa caagttaaca
3781 acaacaattg cattcatttt atgtttcagg ttcaggggga gatgtgggag gttttttaaa
3841 gcaagtaaaa cctctacaaa tgtggtaaaa tcgataagga tcttcctaga gcatggctac
3901 gtagataagt agcatggcgg gttaatcatt aactacaagg aacccctagt gatggagttg
3961 gccactccct ctctgcgcgc tcgctcgctc actgaggccg ggcgaccaaa ggtcgcccga
4021 cgcccgggct ttgcccgggc ggcctcagtg agcgagcgag cgcgcagcct taattaacct
4081 aattcactgg ccgtcgtttt acaacgtcgt gactgggaaa accctggcgt tacccaactt
4141 aatcgccttg cagcacatcc ccctttcgcc agctggcgta atagcgaaga ggcccgcacc
4201 gatcgccctt cccaacagtt gcgcagcctg aatggcgaat gggacgcgcc ctgtagcggc
4261 gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga ccgctacact tgccagcgcc
4321 ctagcgcccg ctcctttcgc tttcttccct tcctttctcg ccacgttcgc cggctttccc
4381 cgtcaagctc taaatcgggg gctcccttta gggttccgat ttagtgcttt acggcacctc
4441 gaccccaaaa aacttgatta gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg
4501 gtttttcgcc ctttgacgtt ggagtccacg ttctttaata gtggactctt gttccaaact
4561 ggaacaacac tcaaccctat ctcggtctat tcttttgatt tataagggat tttgccgatt
4621 tcggcctatt ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa ttttaacaaa
4681 atattaacgc ttacaattta ggtggcactt ttcggggaaa tgtgcgcgga acccctattt
4741 gtttattttt ctaaatacat tcaaatatgt atccgctcat gagacaataa ccctgataaa
4801 tgcttcaata atattgaaaa aggaagagta tgagtattca acatttccgt gtcgccctta
4861 ttcccttttt tgcggcattt tgccttcctg tttttgctca cccagaaacg ctggtgaaag
4921 taaaagatgc tgaagatcag ttgggtgcac gagtgggtta catcgaactg gatctcaaca
4981 gcggtaagat ccttgagagt tttcgccccg aagaacgttt tccaatgatg agcactttta
5041 aagttctgct atgtggcgcg gtattatccc gtattgacgc cgggcaagag caactcggtc
5101 gccgcataca ctattctcag aatgacttgg ttgagtactc accagtcaca gaaaagcatc
5161 ttacggatgg catgacagta agagaattat gcagtgctgc cataaccatg agtgataaca
5221 ctgcggccaa cttacttctg acaacgatcg gaggaccgaa ggagctaacc gcttttttgc
5281 acaacatggg ggatcatgta actcgccttg atcgttggga accggagctg aatgaagcca
5341 taccaaacga cgagcgtgac accacgatgc ctgtagcaat ggcaacaacg ttgcgcaaac
5401 tattaactgg cgaactactt actctagctt cccggcaaca attaatagac tggatggagg
5461 cggataaagt tgcaggacca cttctgcgct cggcccttcc ggctggctgg tttattgctg
5521 ataaatctgg agccggtgag cgtgggtctc gcggtatcat tgcagcactg gggccagatg
5581 gtaagccctc ccgtatcgta gttatctaca cgacggggag tcaggcaact atggatgaac
5641 gaaatagaca gatcgctgag ataggtgcct cactgattaa gcattggtaa ctgtcagacc
5701 aagtttactc atatatactt tagattgatt taaaacttca tttttaattt aaaaggatct
5761 aggtgaagat cctttttgat aatctcatga ccaaaatccc ttaacgtgag ttttcgttcc
5821 actgagcgtc agaccccgta gaaaagatca aaggatcttc ttgagatcct ttttttctgc
5881 gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt tgtttgccgg
5941 atcaagagct accaactctt tttccgaagg taactggctt cagcagagcg cagataccaa
6001 atactgttct tctagtgtag ccgtagttag gccaccactt caagaactct gtagcaccgc
6061 ctacatacct cgctctgcta atcctgttac cagtggctgc tgccagtggc gataagtcgt
6121 gtcttaccgg gttggactca agacgatagt taccggataa ggcgcagcgg tcgggctgaa
6181 cggggggttc gtgcacacag cccagcttgg agcgaacgac ctacaccgaa ctgagatacc
6241 tacagcgtga gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc
6301 cggtaagcgg cagggtcgga acaggagagc gcacgaggga gcttccaggg ggaaacgcct
6361 ggtatcttta tagtcctgtc gggtttcgcc acctctgact tgagcgtcga tttttgtgat
6421 gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa cgcggccttt ttacggttcc
6481 tggccttttg ctggcctttt gctcacatgt tctttcctgc gttatcccct gattctgtgg
6541 ataaccgtat taccgccttt gagtgagctg ataccgctcg ccgcagccga acgaccgagc
6601 gcagcgagtc agtgagcgag gaagcggaag
//
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