| 质粒名称: | pJRH-1362 scFv entry plasmid |
|---|---|
| 出品公司: | Addgene |
| 目录编号: | 201912 |
| 存储实验室: | Jennifer Doudna |
|
载体骨架基本信息 |
|
| 载体名称: | pCAGGS |
| 载体出品公司: | Clontech |
| 空载体大小: | 4957 bp |
| 完整质粒大小: | 4937 bp |
| 载体类型: | 哺乳细胞表达 |
| 载体抗性: | 氨苄霉素 |
| 生长温度: | 37 ℃ |
| 宿主菌: | DH5a |
| 拷贝数: | 高拷贝 |
|
插入基因信息 |
|
| 插入基因名称: | Stuffer sequence (drop out for scFv cloning) + CD8 hinge and transmembrane domain |
| 启动子: | CAG |
| 克隆方法: | Gibson Cloning |
| 5’测序引物 | CTCTAGAGCCTCTGCTAACC |
| 3’测序引物 | GCCAGAAGTCAGATGCTCAAGG |
|
文献引用方法 |
|
| 材料和方法部分: | pJRH-1362 scFv entry plasmid was a gift from Jennifer Doudna (Addgene plasmid # 201912 ; http://n2t.net/addgene:201912 ; RRID:Addgene_201912) |
| 参考文献部分: | In vivo human T cell engineering with enveloped delivery vehicles. Hamilton JR, Chen E, Perez BS, Sandoval Espinoza CR, Kang MH, Trinidad M, Ngo W, Doudna JA. Nat Biotechnol. 2024 Jan 11. doi: 10.1038/s41587-023-02085-z. 10.1038/s41587-023-02085-z PubMed 38212493 |
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载体基本信息
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载体简介
载体序列
LOCUS pJRH-1362 scFv entry plasmid 4937 bp ds-DNA circular SYN 04-AUG-2023
DEFINITION Entry vector for cloning single chain variable fragments displayed
on the human CD8a hinge and transmembrane domain (CAG promoter).
ACCESSION .
VERSION .
KEYWORDS pJRH-1362 scFv entry plasmid
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 4937)
TITLE bioRxiv - Programmable enveloped delivery vehicles for human genome
engineering in vivo
JOURNAL Unpublished
REFERENCE 2 (bases 1 to 4937)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported Aug 4, 2023 from SnapGene Server 1.1.58
http://www.biofeng.com
FEATURES Location/Qualifiers
source 1..4937
/organism="synthetic DNA construct"
/mol_type="other DNA"
enhancer 5..384
/label=CMV enhancer
/note="human cytomegalovirus immediate early enhancer"
intron 642..1658
/label=chimeric intron
/note="chimera between introns from chicken beta-actin and
rabbit beta-globin"
primer_bind 1582..1604
/label=pCAGGS-5
/note="Chimeric intron in CAG promoter, forward primer"
primer_bind 1666..1685
/label=pCAG-F
/note="Rabbit beta-globin intron, for pCAG plasmids,
forward primer"
regulatory 1713..1722
/regulatory_class="other"
/note="vertebrate consensus sequence for strong initiation
of translation (Kozak, 1987)"
primer_bind complement(1959..1978)
/label=Bglob-pA-R
/note="Rabbit beta-globin polyA region, reverse primer"
polyA_signal 2024..2079
/label=-beta-globin poly(A) signal
/note="rabbit beta-globin polyadenylation signal (Gil and
Proudfoot, 1987)"
primer_bind complement(2078..2097)
/label=rbglobpA-R
/note="Rabbit beta-globin polyA, reverse primer. Also
called rb-glob-pA-term-R"
primer_bind complement(2440..2456)
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
primer_bind complement(2440..2456)
/label=M13 Reverse
/note="In lacZ gene. Also called M13-rev"
primer_bind complement(2453..2475)
/label=M13/pUC Reverse
/note="In lacZ gene"
protein_bind 2464..2480
/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)."
promoter complement(2488..2518)
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind 2533..2554
/label=CAP binding site
/bound_moiety="E. coli catabolite activator protein"
/note="CAP binding activates transcription in the presence
of cAMP."
promoter 2613..2808
/label=SV40 promoter
/note="SV40 early promoter"
rep_origin 2659..2794
/label=SV40 ori
/note="SV40 origin of replication"
primer_bind 2721..2740
/label=SV40pro-F
/note="SV40 promoter/origin, forward primer"
polyA_signal 2814..2948
/label=SV40 poly(A) signal
/note="SV40 polyadenylation signal"
primer_bind complement(2851..2870)
/label=SV40pA-R
/note="SV40 polyA, reverse primer"
primer_bind 2905..2924
/label=EBV-rev
/note="SV40 polyA terminator, reverse primer"
primer_bind complement(3016..3033)
/label=L4440
/note="L4440 vector, forward primer"
rep_origin complement(3187..3775)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind complement(3267..3286)
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
CDS complement(3946..4806)
/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 4569..4588
/label=Amp-R
/note="Ampicillin resistance gene, reverse primer"
promoter complement(4807..4911)
/gene="bla"
/label=AmpR promoter
ORIGIN
1 ggtcgacatt gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat
61 agcccatata tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg
121 cccaacgacc cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata
181 gggactttcc attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta
241 catcaagtgt atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc
301 gcctggcatt atgcccagta catgacctta tgggactttc ctacttggca gtacatctac
361 gtattagtca tcgctattac catggtcgag gtgagcccca cgttctgctt cactctcccc
421 atctcccccc cctccccacc cccaattttg tatttattta ttttttaatt attttgtgca
481 gcgatggggg cggggggggg ggggrgmsrg gcgaggggcg gggcggggcg aggcggagag
541 gtgcggcggc agccaatcag agcggcgcgc tccgaaagtt tccttttatg gcgaggcggc
601 ggcggcggcg gccctataaa aagcgaagcg cgcggcgggc gggagtcgct gcgcgctgcc
661 ttcgccccgt gccccgctcc gccgccgcct cgcgccgccc gccccggctc tgactgaccg
721 cgttactccc acaggtgagc gggcgggacg gcccttctcc tccgggctgt aattagcgct
781 tggtttaatg acggcttgtt tcttttctgt ggctgcgtga aagccttgag gggctccggg
841 agggcccttt gtgcgggggg agcggctcgg ggggtgcgtg cgtgtgtgtg tgcgtgggga
901 gcgccgcgtg cggctccgcg ctgcccggcg gctgtgagcg ctgcgggcgc ggcgcggggc
961 tttgtgcgct ccgcagtgtg cgcgagggga gcgcggccgg gggcggtgcc ccgcggtgcg
1021 gggggggctg cgaggggaac aaaggctgcg tgcggggtgt gtgcgtgggg gggtgagcag
1081 ggggtgtggg cgcgtcggtc gggctgcaac cccccctgca cccccctccc cgagttgctg
1141 agcacggccc ggcttcgggt gcggggctcc gtacggggcg tggcgcgggg ctcgccgtgc
1201 cgggcggggg gtggcggcag gtgggggtgc cgggcggggc ggggccgcct cgggccgggg
1261 agggctcggg ggaggggcgc ggcggccccc ggagcgccgg cggctgtcga ggcgcggcga
1321 gccgcagcca ttgcctttta tggtaatcgt gcgagagggc gcagggactt cctttgtccc
1381 aaatctgtgc ggagccgaaa tctgggaggc gccgccgcac cccctctagc gggcgcgggg
1441 cgaagcggtg cggcgccggc aggaaggaaa tgggcgggga gggccttcgt gcgtcgccgc
1501 gccgccgtcc ccttctccct ctccagcctc ggggctgtcc gcggggggac ggctgccttc
1561 gggggggacg gggcagggcg gggttcggct tctggcgtgt gaccggcggc tctagagcct
1621 ctgctaacca tgttcatgcc ttcttctttt tcctacagct cctgggcaac gtgctggtta
1681 ttgtgctgtc tcatcatttt ggcaaagaat tcgccaccat ggccttaacc gtctcctcaa
1741 ccacgacgcc agcgccgcga ccaccaacac cggcgcccac catcgcgtcg cagcccctgt
1801 ccctgcgccc agaggcgtgc cggccagcgg cggggggcgc agtgcacacg agggggctgg
1861 acttcgcctg tgatatctac atctgggcgc ccttggccgg gacttgtggg gttcttctcc
1921 tgtcactggt tatcaccctt tactgctaag ctagcagatc tttttccctc tgccaaaaat
1981 tatggggaca tcatgaagcc ccttgagcat ctgacttctg gctaataaag gaaatttatt
2041 ttcattgcaa tagtgtgttg gaattttttg tgtctctcac tcggaaggac atatgggagg
2101 gcaaatcatt taaaacatca gaatgagtat ttggtttaga gtttggcaac atatgcccat
2161 atgctggctg ccatgaacaa aggttggcta taaagaggtc atcagtatat gaaacagccc
2221 cctgctgtcc attccttatt ccatagaaaa gccttgactt gaggttagat tttttttata
2281 ttttgttttg tgttattttt ttctttaaca tccctaaaat tttccttaca tgttttacta
2341 gccagatttt tcctcctctc ctgactactc ccagtcatag ctgtccctct tctcttatgg
2401 agatccctcg acctgcagcc caagcttggc gtaatcatgg tcatagctgt ttcctgtgtg
2461 aaattgttat ccgctcacaa ttccacacaa catacgagcc ggaagcataa agtgtaaagc
2521 ctggggtgcc taatgagtga gctaactcac attaattgcg ttgcgctcac tgcccgcttt
2581 ccagtcggga aacctgtcgt gccagcggat ccgcatctca attagtcagc aaccatagtc
2641 ccgcccctaa ctccgcccat cccgccccta actccgccca gttccgccca ttctccgccc
2701 catggctgac taattttttt tatttatgca gaggccgagg ccgcctcggc ctctgagcta
2761 ttccagaagt agtgaggagg cttttttgga ggcctaggct tttgcaaaaa gctaacttgt
2821 ttattgcagc ttataatggt tacaaataaa gcaatagcat cacaaatttc acaaataaag
2881 catttttttc actgcattct agttgtggtt tgtccaaact catcaatgta tcttatcatg
2941 tctggatccg ctgcattaat gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg
3001 gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc tgcggcgagc
3061 ggtatcagct cactcaaagg cggtaatacg gttatccaca gaatcagggg ataacgcagg
3121 aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct
3181 ggcgtttttc cataggctcc gcccccctga cgagcatcac aaaaatcgac gctcaagtca
3241 gaggtggcga aacccgacag gactataaag ataccaggcg tttccccctg gaagctccct
3301 cgtgcgctct cctgttccga ccctgccgct taccggatac ctgtccgcct ttctcccttc
3361 gggaagcgtg gcgctttctc atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt
3421 tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct gcgccttatc
3481 cggtaactat cgtcttgagt ccaacccggt aagacacgac ttatcgccac tggcagcagc
3541 cactggtaac aggattagca gagcgaggta tgtaggcggt gctacagagt tcttgaagtg
3601 gtggcctaac tacggctaca ctagaagaac agtatttggt atctgcgctc tgctgaagcc
3661 agttaccttc ggaaaaagag ttggtagctc ttgatccggc aaacaaacca ccgctggtag
3721 cggtggtttt tttgtttgca agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga
3781 tcctttgatc ttttctacgg ggtctgacgc tcagtggaac gaaaactcac gttaagggat
3841 tttggtcatg agattatcaa aaaggatctt cacctagatc cttttaaatt aaaaatgaag
3901 ttttaaatca atctaaagta tatatgagta aacttggtct gacagttacc aatgcttaat
3961 cagtgaggca cctatctcag cgatctgtct atttcgttca tccatagttg cctgactccc
4021 cgtcgtgtag ataactacga tacgggaggg cttaccatct ggccccagtg ctgcaatgat
4081 accgcgagac ccacgctcac cggctccaga tttatcagca ataaaccagc cagccggaag
4141 ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc atccagtcta ttaattgttg
4201 ccgggaagct agagtaagta gttcgccagt taatagtttg cgcaacgttg ttgccattgc
4261 tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct tcattcagct ccggttccca
4321 acgatcaagg cgagttacat gatcccccat gttgtgcaaa aaagcggtta gctccttcgg
4381 tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg ttatggcagc
4441 actgcataat tctcttactg tcatgccatc cgtaagatgc ttttctgtga ctggtgagta
4501 ctcaaccaag tcattctgag aatagtgtat gcggcgaccg agttgctctt gcccggcgtc
4561 aatacgggat aataccgcgc cacatagcag aactttaaaa gtgctcatca ttggaaaacg
4621 ttcttcgggg cgaaaactct caaggatctt accgctgttg agatccagtt cgatgtaacc
4681 cactcgtgca cccaactgat cttcagcatc ttttactttc accagcgttt ctgggtgagc
4741 aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat
4801 actcatactc ttcctttttc aatattattg aagcatttat cagggttatt gtctcatgag
4861 cggatacata tttgaatgta tttagaaaaa taaacaaata ggggttccgc gcacatttcc
4921 ccgaaaagtg ccacctg
//
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