| 质粒名称: | pET28a-uvsX-H6 |
|---|---|
| 出品公司: | Addgene |
| 目录编号: | 163913 |
| 存储实验室: | Paul Freemont |
|
载体骨架基本信息 |
|
| 载体名称: | pET28a |
| 载体出品公司: | Novagen |
| 空载体大小: | 5369 bp |
| 完整质粒大小: | 6391 bp |
| 载体修饰: | 移除N端子His标签 |
| 载体类型: | 细菌表达 |
| 筛选标记: | -- |
| 载体抗性: | 卡那霉素，50μg/mL |
| 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: | uvsX |
| 物种来源: | 肠道细菌噬菌体T4 |
| 插入大小: | 1173 bp |
| 基因突变: | -- |
| GenBank编号 | -- |
| 启动子: | T7启动子 |
| 标签/融合蛋白: | His（插入C端子） |
| 克隆方法: | 限制性内切酶 |
| 5’克隆位点: | BsaI(破坏) |
| 3’克隆位点: | BsaI (破坏) |
| 5’测序引物 | TAATACGACTCACTATAGGG |
| 3’测序引物 | GCTAGTTATTGCTCAGCGG |
|
文献引用方法 |
|
| 材料和方法部分: | pET28a-uvsX-H6 was a gift from Paul Freemont (Addgene plasmid # 163913 ; http://n2t.net/addgene:163913 ; RRID:Addgene_163913) |
| 参考文献部分: | A Multiplexed Cas13-Based Assay with Point-of-Care Attributes for Simultaneous COVID-19 Diagnosis and Variant Surveillance. Patchsung M, Homchan A, Aphicho K, Suraritdechachai S, Wanitchanon T, Pattama A, Sappakhaw K, Meesawat P, Wongsatit T, Athipanyasilp A, Jantarug K, Athipanyasilp N, Buahom J, Visanpattanasin S, Niljianskul N, Chaiyen P, Tinikul R, Wichukchinda N, Mahasirimongkol S, Sirijatuphat R, Angkasekwinai N, Crone MA, Freemont PS, Joung J, Ladha A, Abudayyeh O, Gootenberg J, Zhang F, Chewapreecha C, Chanarat S, Horthongkham N, Pakotiprapha D, Uttamapinant C. CRISPR J. 2022 Nov 11. doi: 10.1089/crispr.2022.0048. 10.1089/crispr.2022.0048 PubMed 36367987 |
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 pET28a-uvsX-H6 6391 bp ds-DNA circular SYN 13-MAY-2021
DEFINITION Expresses uvsX for bacterial expression and affinity purification.
ACCESSION .
VERSION .
KEYWORDS pET28a-uvsX-H6
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 6391)
TITLE One Pot CRISPR RPA
JOURNAL Unpublished
REFERENCE 2 (bases 1 to 6391)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported May 13, 2021 from SnapGene Server 1.1.58
http://www.biofeng.com
FEATURES Location/Qualifiers
source 1..6391
/organism="synthetic DNA construct"
/mol_type="other DNA"
CDS complement(646..1728)
/codon_start=1
/gene="lacI"
/product="lac repressor"
/label=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)."
/translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
ALADSLMQLARQVSRLESGQ"
primer_bind 1690..1709
/label=LacI-R
/note="LacI, reverse primer"
promoter complement(1729..1806)
/gene="lacI"
/label=lacI promoter
primer_bind 2012..2031
/label=pBRrevBam
/note="pBR322 vectors, tet region, downstream of BamHI,
reverse primer"
primer_bind 2115..2134
/label=T7
/note="T7 promoter, forward primer"
promoter 2115..2133
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
protein_bind 2134..2158
/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 2167..2196
/label=lac promoter
/note="promoter for the E. coli lac operon"
RBS 2216..2238
/note="efficient ribosome binding site from bacteriophage
T7 gene 10 (Olins and Rangwala, 1989)"
CDS 3419..3436
/codon_start=1
/product="6xHis affinity tag"
/label=6xHis
/translation="HHHHHH"
primer_bind complement(3440..3458)
/label=T7 Term
/note="T7 terminator, reverse primer"
terminator 3454..3501
/label=T7 terminator
/note="transcription terminator for bacteriophage T7 RNA
polymerase"
rep_origin 3538..3993
/direction=RIGHT
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
primer_bind complement(3625..3644)
/label=F1ori-R
/note="F1 origin, reverse primer"
primer_bind 3835..3856
/label=F1ori-F
/note="F1 origin, forward primer"
CDS complement(4085..4900)
/codon_start=1
/gene="aph(3')-Ia"
/product="aminoglycoside phosphotransferase"
/label=KanR
/note="confers resistance to kanamycin in bacteria or G418
(Geneticin(R)) in eukaryotes"
/translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
primer_bind 4808..4827
/label=Kan-R
/note="Kanamycin resistance gene, reverse primer"
primer_bind 4892..4911
/label=pENTR-R
/note="pENTR vectors, reverse primer"
rep_origin 5022..5610
/direction=RIGHT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind 5511..5530
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
primer_bind 5764..5781
/label=L4440
/note="L4440 vector, forward primer"
misc_feature 5796..5935
/label=bom
/note="basis of mobility region from pBR322"
primer_bind complement(5861..5880)
/label=pRS-marker
/note="pRS vectors, use to sequence yeast selectable
marker"
primer_bind 6021..6043
/label=pGEX 3'
/note="pGEX vectors, reverse primer"
CDS complement(6037..6228)
/codon_start=1
/gene="rop"
/product="Rop protein, which maintains plasmids at low copy
number"
/label=rop
/translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
DELYRSCLARFGDDGENL"
ORIGIN
1 atcactcagg gtcaatgcca gcgcttcgtt aatacagatg taggtgttcc acagggtagc
61 cagcagcatc ctgcgatgca gatccggaac ataatggtgc agggcgctga cttccgcgtt
121 tccagacttt acgaaacacg gaaaccgaag accattcatg ttgttgctca ggtcgcagac
181 gttttgcagc agcagtcgct tcacgttcgc tcgcgtatcg gtgattcatt ctgctaacca
241 gtaaggcaac cccgccagcc tagccgggtc ctcaacgaca ggagcacgat catgcgcacc
301 cgtggggccg ccatgccggc gataatggcc tgcttctcgc cgaaacgttt ggtggcggga
361 ccagtgacga aggcttgagc gagggcgtgc aagattccga ataccgcaag cgacaggccg
421 atcatcgtcg cgctccagcg aaagcggtcc tcgccgaaaa tgacccagag cgctgccggc
481 acctgtccta cgagttgcat gataaagaag acagtcataa gtgcggcgac gatagtcatg
541 ccccgcgccc accggaagga gctgactggg ttgaaggctc tcaagggcat cggtcgagat
601 cccggtgcct aatgagtgag ctaacttaca ttaattgcgt tgcgctcact gcccgctttc
661 cagtcgggaa acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc ggggagaggc
721 ggtttgcgta ttgggcgcca gggtggtttt tcttttcacc agtgagacgg gcaacagctg
781 attgcccttc accgcctggc cctgagagag ttgcagcaag cggtccacgc tggtttgccc
841 cagcaggcga aaatcctgtt tgatggtggt taacggcggg atataacatg agctgtcttc
901 ggtatcgtcg tatcccacta ccgagatatc cgcaccaacg cgcagcccgg actcggtaat
961 ggcgcgcatt gcgcccagcg ccatctgatc gttggcaacc agcatcgcag tgggaacgat
1021 gccctcattc agcatttgca tggtttgttg aaaaccggac atggcactcc agtcgccttc
1081 ccgttccgct atcggctgaa tttgattgcg agtgagatat ttatgccagc cagccagacg
1141 cagacgcgcc gagacagaac ttaatgggcc cgctaacagc gcgatttgct ggtgacccaa
1201 tgcgaccaga tgctccacgc ccagtcgcgt accgtcttca tgggagaaaa taatactgtt
1261 gatgggtgtc tggtcagaga catcaagaaa taacgccgga acattagtgc aggcagcttc
1321 cacagcaatg gcatcctggt catccagcgg atagttaatg atcagcccac tgacgcgttg
1381 cgcgagaaga ttgtgcaccg ccgctttaca ggcttcgacg ccgcttcgtt ctaccatcga
1441 caccaccacg ctggcaccca gttgatcggc gcgagattta atcgccgcga caatttgcga
1501 cggcgcgtgc agggccagac tggaggtggc aacgccaatc agcaacgact gtttgcccgc
1561 cagttgttgt gccacgcggt tgggaatgta attcagctcc gccatcgccg cttccacttt
1621 ttcccgcgtt ttcgcagaaa cgtggctggc ctggttcacc acgcgggaaa cggtctgata
1681 agagacaccg gcatactctg cgacatcgta taacgttact ggtttcacat tcaccaccct
1741 gaattgactc tcttccgggc gctatcatgc cataccgcga aaggttttgc gccattcgat
1801 ggtgtccggg atctcgacgc tctcccttat gcgactcctg cattaggaag cagcccagta
1861 gtaggttgag gccgttgagc accgccgccg caaggaatgg tgcatgcaag gagatggcgc
1921 ccaacagtcc cccggccacg gggcctgcca ccatacccac gccgaaacaa gcgctcatga
1981 gcccgaagtg gcgagcccga tcttccccat cggtgatgtc ggcgatatag gcgccagcaa
2041 ccgcacctgt ggcgccggtg atgccggcca cgatgcgtcc ggcgtagagg atcgagatct
2101 cgatcccgcg aaattaatac gactcactat aggggaattg tgagcggata acaattcccc
2161 tctagattta cactttatgc ttccggctcg tatgtttcta gagaattcaa ataattttgt
2221 ttaactttaa gaaggagatt taaatatgag cgatctgaaa agccgtctga ttaaagcaag
2281 caccagcaaa ctgaccgcag aactgacagc aagcaaattt ttcaacgaaa aagatgtggt
2341 gcgcacgaaa attccgatga tgaatattgc actgagcggt gaaattaccg gtggtatgca
2401 gagcggtctg ctgattctgg ctggtccgag caaaagcttt aaaagcaatt ttggtctgac
2461 catggtgagc agctatatgc gtcagtatcc ggatgcagtt tgtctgtttt atgatagcga
2521 atttggtatc acaccggcat atctgcgtag catgggtgtt gatccggaac gtgttattca
2581 tacaccggtt cagagcctgg aacagctgcg tattgatatg gttaatcagc tggatgcaat
2641 tgaacgtggt gaaaaagtgg tggtgtttat tgatagcctg ggtaatctgg caagcaaaaa
2701 agaaaccgaa gatgccctga atgaaaaagt tgttagcgat atgacccgtg ccaaaaccat
2761 gaaaagcctg tttcgtattg ttaccccgta tttcagcacc aaaaacattc cgtgtattgc
2821 catcaaccat acctatgaaa cccaagagat gtttagcaaa accgttatgg gtggtggcac
2881 cggtccgatg tatagcgcag ataccgtttt tatcattggc aaacgccaga ttaaagatgg
2941 tagcgatctg cagggttatc agtttgttct gaatgttgaa aaaagccgca ccgtgaaaga
3001 gaaaagcaaa ttctttatcg acgtgaaatt cgatggtggc attgatccgt atagtggtct
3061 gctggatatg gcactggaac tgggttttgt tgttaaaccg aaaaatggtt ggtatgcccg
3121 tgaatttctg gatgaagaaa ccggtgaaat gatccgtgaa gaaaaaagct ggcgtgcaaa
3181 agataccaat tgtaccacct tttggggtcc gctgttcaaa catcagccgt ttcgtgatgc
3241 cattaaacgt gcatatcagc tgggtgccat cgatagcaat gaaattgttg aagccgaagt
3301 ggatgaactg atcaatagca aagtggaaaa attcaagagt ccggaatcca aaagcaaaag
3361 cgcagcagat ctggaaaccg atctggaaca actgagcgat atggaagaat ttaacgaaca
3421 ccatcaccat caccactaac cgctgagcaa taactagcat aaccccttgg ggcctctaaa
3481 cgggtcttga ggggtttttt gctgaaagga ggaactatat ccggattggc gaatgggacg
3541 cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta
3601 cacttgccag cgccctagcg cccgctcctt tcgctttctt cccttccttt ctcgccacgt
3661 tcgccggctt tccccgtcaa gctctaaatc gggggctccc tttagggttc cgatttagtg
3721 ctttacggca cctcgacccc aaaaaacttg attagggtga tggttcacgt agtgggccat
3781 cgccctgata gacggttttt cgccctttga cgttggagtc cacgttcttt aatagtggac
3841 tcttgttcca aactggaaca acactcaacc ctatctcggt ctattctttt gatttataag
3901 ggattttgcc gatttcggcc tattggttaa aaaatgagct gatttaacaa aaatttaacg
3961 cgaattttaa caaaatatta acgcttacaa tttaggtggc acttttcggg gaaatgtgcg
4021 cggaacccct atttgtttat ttttctaaat acattcaaat atgtatccgc tcatgaatta
4081 attcttagaa aaactcatcg agcatcaaat gaaactgcaa tttattcata tcaggattat
4141 caataccata tttttgaaaa agccgtttct gtaatgaagg agaaaactca ccgaggcagt
4201 tccataggat ggcaagatcc tggtatcggt ctgcgattcc gactcgtcca acatcaatac
4261 aacctattaa tttcccctcg tcaaaaataa ggttatcaag tgagaaatca ccatgagtga
4321 cgactgaatc cggtgagaat ggcaaaagtt tatgcatttc tttccagact tgttcaacag
4381 gccagccatt acgctcgtca tcaaaatcac tcgcatcaac caaaccgtta ttcattcgtg
4441 attgcgcctg agcgagacga aatacgcgat cgctgttaaa aggacaatta caaacaggaa
4501 tcgaatgcaa ccggcgcagg aacactgcca gcgcatcaac aatattttca cctgaatcag
4561 gatattcttc taatacctgg aatgctgttt tcccggggat cgcagtggtg agtaaccatg
4621 catcatcagg agtacggata aaatgcttga tggtcggaag aggcataaat tccgtcagcc
4681 agtttagtct gaccatctca tctgtaacat cattggcaac gctacctttg ccatgtttca
4741 gaaacaactc tggcgcatcg ggcttcccat acaatcgata gattgtcgca cctgattgcc
4801 cgacattatc gcgagcccat ttatacccat ataaatcagc atccatgttg gaatttaatc
4861 gcggcctaga gcaagacgtt tcccgttgaa tatggctcat aacacccctt gtattactgt
4921 ttatgtaagc agacagtttt attgttcatg accaaaatcc cttaacgtga gttttcgttc
4981 cactgagcgt cagaccccgt agaaaagatc aaaggatctt cttgagatcc tttttttctg
5041 cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg
5101 gatcaagagc taccaactct ttttccgaag gtaactggct tcagcagagc gcagatacca
5161 aatactgtcc ttctagtgta gccgtagtta ggccaccact tcaagaactc tgtagcaccg
5221 cctacatacc tcgctctgct aatcctgtta ccagtggctg ctgccagtgg cgataagtcg
5281 tgtcttaccg ggttggactc aagacgatag ttaccggata aggcgcagcg gtcgggctga
5341 acggggggtt cgtgcacaca gcccagcttg gagcgaacga cctacaccga actgagatac
5401 ctacagcgtg agctatgaga aagcgccacg cttcccgaag ggagaaaggc ggacaggtat
5461 ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg agcttccagg gggaaacgcc
5521 tggtatcttt atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg atttttgtga
5581 tgctcgtcag gggggcggag cctatggaaa aacgccagca acgcggcctt tttacggttc
5641 ctggcctttt gctggccttt tgctcacatg ttctttcctg cgttatcccc tgattctgtg
5701 gataaccgta ttaccgcctt tgagtgagct gataccgctc gccgcagccg aacgaccgag
5761 cgcagcgagt cagtgagcga ggaagcggaa gagcgcctga tgcggtattt tctccttacg
5821 catctgtgcg gtatttcaca ccgcaatggt gcactctcag tacaatctgc tctgatgccg
5881 catagttaag ccagtataca ctccgctatc gctacgtgac tgggtcatgg ctgcgccccg
5941 acacccgcca acacccgctg acgcgccctg acgggcttgt ctgctcccgg catccgctta
6001 cagacaagct gtgaccgtct ccgggagctg catgtgtcag aggttttcac cgtcatcacc
6061 gaaacgcgcg aggcagctgc ggtaaagctc atcagcgtgg tcgtgaagcg attcacagat
6121 gtctgcctgt tcatccgcgt ccagctcgtt gagtttctcc agaagcgtta atgtctggct
6181 tctgataaag cgggccatgt taagggcggt tttttcctgt ttggtcactg atgcctccgt
6241 gtaaggggga tttctgttca tgggggtaat gataccgatg aaacgagaga ggatgctcac
6301 gatacgggtt actgatgatg aacatgcccg gttactggaa cgttgtgagg gtaaacaact
6361 ggcggtatgg atgcggcggg accagagaaa a
//
