Price Information
| Cat No. | Plasmid Name | Availability | Add to cart |
|---|---|---|---|
| V012689 | Twinstrep-SUMO-huLwCas13a | In stock (lyophilized plasmid) |
Buy one, get one free! |
Two vials of lyophilized plasmid will be delivered, each vial is about 5µg.
Basic Vector Information
Twinstrep-SUMO-huLwCas13a for recombinant protein bacterial expression. Insert is human codon optimized but expresses well in bacteria.
Twinstrep-SUMO-huLwCas13a vector Vector Map
Plasmid Resuspension Protocol:
1. Centrifuge at 5,000×g for 5 min.
2. Carefully open the tube and add 20 μl of sterile water to dissolve the DNA.
3. Close the tube and incubate for 10 minutes at room temperature.
4. Briefly vortex the tube and then do a quick spin to concentrate the liquid at the bottom. Speed is less than 5000×g.
5.Store the plasmid at -20 ℃.
Twinstrep-SUMO-huLwCas13a vector Sequence
LOCUS 40924_48982 9542 bp DNA circular SYN 06-APR-2021
DEFINITION Twinstrep-SUMO-huLwCas13a for recombinant protein bacterial
expression. Insert is human codon optimized but expresses well in
bacteria..
ACCESSION .
VERSION .
KEYWORDS .
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 9542)
AUTHORS Gootenberg JS, Abudayyeh OO, Lee JW, Essletzbichler P, Dy AJ, Joung
J, Verdine V, Donghia N, Daringer NM, Freije CA, Myhrvold C,
Bhattacharyya RP, Livny J, Regev A, Koonin EV, Hung DT, Sabeti PC,
Collins JJ, Zhang F
TITLE Nucleic acid detection with CRISPR-Cas13a/C2c2.
JOURNAL Science. 2017 Apr 13. pii: eaam9321. doi: 10.1126/science.aam9321.
PUBMED 28408723
REFERENCE 2 (bases 1 to 9542)
TITLE Direct Submission
REFERENCE 3 (bases 1 to 9542)
AUTHORS .
TITLE Direct Submission
COMMENT SGRef: number: 1; type: "Journal Article"; journalName: "Science.
2017 Apr 13. pii: eaam9321. doi: 10.1126/science.aam9321."
COMMENT SGRef: number: 2; type: "Journal Article"
FEATURES Location/Qualifiers
source 1..9542
/mol_type="other DNA"
/organism="synthetic DNA construct"
misc_feature 278..417
/label=bom
/note="basis of mobility region from pBR322"
rep_origin complement(603..1191)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
CDS complement(1365..2222)
/codon_start=1
/label=AmpR
/note="beta-lactamase"
/translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
LIKHW"
promoter complement(2223..2327)
/label=AmpR promoter
promoter 2440..2468
/label=tet promoter
/note="E. coli promoter for tetracycline efflux protein
gene"
terminator complement(2643..2690)
/label=T7 terminator
/note="transcription terminator for bacteriophage T7 RNA
polymerase"
CDS complement(2754..6209)
/codon_start=1
/label=LwCas13a
/note="Leptotrichia wadei CRISPR-Cas effector that acts as
an RNA-guided RNAse (Abudayyeh et al., 2017)"
/translation="MKVTKVDGISHKKYIEEGKLVKSTSEENRTSERLSELLSIRLDIY
IKNPDNASEEENRIRRENLKKFFSNKVLHLKDSVLYLKNRKEKNAVQDKNYSEEDISEY
DLKNKNSFSVLKKILLNEDVNSEELEIFRKDVEAKLNKINSLKYSFEENKANYQKINEN
NVEKVGGKSKRNIIYDYYRESAKRNDYINNVQEAFDKLYKKEDIEKLFFLIENSKKHEK
YKIREYYHKIIGRKNDKENFAKIIYEEIQNVNNIKELIEKIPDMSELKKSQVFYKYYLD
KEELNDKNIKYAFCHFVEIEMSQLLKNYVYKRLSNISNDKIKRIFEYQNLKKLIENKLL
NKLDTYVRNCGKYNYYLQVGEIATSDFIARNRQNEAFLRNIIGVSSVAYFSLRNILETE
NENDITGRMRGKTVKNNKGEEKYVSGEVDKIYNENKQNEVKENLKMFYSYDFNMDNKNE
IEDFFANIDEAISSIRHGIVHFNLELEGKDIFAFKNIAPSEISKKMFQNEINEKKLKLK
IFKQLNSANVFNYYEKDVIIKYLKNTKFNFVNKNIPFVPSFTKLYNKIEDLRNTLKFFW
SVPKDKEEKDAQIYLLKNIYYGEFLNKFVKNSKVFFKITNEVIKINKQRNQKTGHYKYQ
KFENIEKTVPVEYLAIIQSREMINNQDKEEKNTYIDFIQQIFLKGFIDYLNKNNLKYIE
SNNNNDNNDIFSKIKIKKDNKEKYDKILKNYEKHNRNKEIPHEINEFVREIKLGKILKY
TENLNMFYLILKLLNHKELTNLKGSLEKYQSANKEETFSDELELINLLNLDNNRVTEDF
ELEANEIGKFLDFNENKIKDRKELKKFDTNKIYFDGENIIKHRAFYNIKKYGMLNLLEK
IADKAKYKISLKELKEYSNKKNEIEKNYTMQQNLHRKYARPKKDEKFNDEDYKEYEKAI
GNIQKYTHLKNKVEFNELNLLQGLLLKILHRLVGYTSIWERDLRFRLKGEFPENHYIEE
IFNFDNSKNVKYKSGQIVEKYINFYKELYKDNVEKRSIYSDKKVKKLKQEKKDLYIRNY
IAHFNYIPHAEISLLEVLENLRKLLSYDRKLKNAIMKSIVDILKEYGFVATFKIGADKK
IEIQTLESEKIVHLKNLKKKKLMTDRNSEELCELVKVMFEYKALE"
CDS complement(6213..6506)
/codon_start=1
/label=SUMO
/note="cleavable ubiquitin-like protein tag"
/translation="MSDSEVNQEAKPEVKPEVKPETHINLKVSDGSSEIFFKIKKTTPL
RRLMEAFAKRQGKEMDSLRFLYDGIRIQADQTPEDLDMEDNDIIEAHREQIGG"
CDS complement(6507..6590)
/codon_start=1
/label=Twin-Strep-tag
/note="two Strep-Tag(R)II moieties connected by a linker
for enhanced binding to Strep-Tactin(R), an engineered form
of streptavidin (Schmidt et al., 2013)"
/translation="WSHPQFEKGGGSGGGSGGSAWSHPQFEK"
CDS complement(6603..6620)
/codon_start=1
/label=thrombin site
/note="thrombin recognition and cleavage site"
/translation="LVPRGS"
CDS complement(6630..6647)
/codon_start=1
/label=6xHis
/note="6xHis affinity tag"
/translation="HHHHHH"
RBS complement(6666..6688)
/label=RBS
/note="efficient ribosome binding site from bacteriophage
T7 gene 10 (Olins and Rangwala, 1989)"
protein_bind complement(6703..6727)
/label=lac operator
/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(6728..6746)
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
promoter 7055..7132
/label=lacI promoter
CDS 7133..8212
/codon_start=1
/label=lacI
/note="lac repressor"
/translation="VKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
ALADSLMQLARQVSRLESGQ"
protein_bind 8228..8249
/label=CAP binding site
/note="CAP binding activates transcription in the presence
of cAMP."
CDS join(9527..9542,1..173)
/codon_start=1
/label=rop
/note="Rop protein, which maintains plasmids at low copy
number"
/translation="VTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
DELYRSCLARFGDDGENL"