Price Information
| Cat No. | Plasmid Name | Availability | Add to cart |
|---|---|---|---|
| V001870 | pYS1 | In stock, 1 week for quality controls |
Buy one, get one free! |
Two tubes of lyophilized plasmid will be delivered, each tube is about 5µg.
Basic Vector Information
- Vector Name:
- pYS1
- Antibiotic Resistance:
- Kanamycin
- Length:
- 11783 bp
- Type:
- Gene deletion vector
- Replication origin:
- ori
- Source/Author:
- Shenkerman Y, Elharar Y, Vishkautzan M, Gur E.
- Promoter:
- sacB
pYS1 vector Vector Map
Plasmid 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 ℃.
6. The concentration of plasmid re-measurement sometimes differs from the nominal value, which may be due to the position of the lyophilized plasmid in the tube, the efficiency of the re-dissolution, the measurement bias, and adsorption on the wall of the tube, therefore, it is recommended to transform and extract the plasmid before using it
General Plasmid Transform Protocol
1. Take one 100μl of the competent cells and thaw it on ice for 10min, add 2μl of plasmid, then ice bath for 30min, then heat-shock it at 42℃ for 60s, do not stir, and then ice bath for 2min.
2. Add 900μl of LB liquid medium without antibiotics, and incubate at 37℃ for 45min (30℃ for 1-1.5 hours) with 180rpm shaking.
3. Centrifuge at 6000rpm for 5min, leave only 100μl of supernatant to resuspend the bacterial precipitate and spread it onto the target plasmid-resistant LB plate.
4. Invert the plate and incubate at 37℃ for 14h, or at 30℃ for 20h.
5. Pick a single colony into LB liquid medium, add the corresponding antibiotics, incubate at 220rpm for 14h, and extract the plasmid according to the experimental needs and the instructions of the plasmid extraction kit.
pYS1 vector Sequence
LOCUS 40924_48258 11783 bp DNA circular SYN 18-DEC-2018
DEFINITION Gene deletion vector pYS1, complete sequence.
ACCESSION .
VERSION .
KEYWORDS .
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 11783)
AUTHORS Shenkerman Y, Elharar Y, Vishkautzan M, Gur E.
TITLE Efficient and simple generation of unmarked gene deletions in
Mycobacterium smegmatis
JOURNAL Gene 533 (1), 374-378 (2014)
PUBMED 24100088
REFERENCE 2 (bases 1 to 11783)
AUTHORS Shenkerman Y, Elharar Y, Vishkautzan M, Gur E.
TITLE Direct Submission
JOURNAL Submitted (27-AUG-2013) Life Sciences, Ben-Gurion University of the
Negev, Rager Blvd, Beer-Sheva 84105, Israel
REFERENCE 3 (bases 1 to 11783)
TITLE Direct Submission
REFERENCE 4 (bases 1 to 11783)
AUTHORS .
TITLE Direct Submission
COMMENT SGRef: number: 1; type: "Journal Article"; journalName: "Gene";
date: "2014"; volume: "533"; issue: "1"; pages: "374-378"
COMMENT SGRef: number: 2; type: "Journal Article"; journalName: "Submitted
(27-AUG-2013) Life Sciences, Ben-Gurion University of the Negev,
Rager Blvd, Beer-Sheva 84105, Israel"
COMMENT SGRef: number: 3; type: "Journal Article"
COMMENT ##Assembly-Data-START##
Sequencing Technology :: Sanger dideoxy sequencing
##Assembly-Data-END##
FEATURES Location/Qualifiers
source 1..11783
/mol_type="other DNA"
/organism="synthetic DNA construct"
promoter 204..648
/label=sacB promoter
/note="sacB promoter and control region"
CDS 649..2043
/codon_start=1
/label=SacB
/note="secreted levansucrase that renders bacterial growth
sensitive to sucrose"
/translation="MNIKKFAKQATVLTFTTALLAGGATQAFAKETNQKPYKETYGISH
ITRHDMLQIPEQQKNEKYQVPEFDSSTIKNISSAKGLDVWDSWPLQNADGTVANYHGYH
IVFALAGDPKNADDTSIYMFYQKVGETSIDSWKNAGRVFKDSDKFDANDSILKDQTQEW
SGSATFTSDGKIRLFYTDFSGKHYGKQTLTTAQVNVSASDSSLNINGVEDYKSIFDGDG
KTYQNVQQFIDEGNYSSGDNHTLRDPHYVEDKGHKYLVFEANTGTEDGYQGEESLFNKA
YYGKSTSFFRQESQKLLQSDKKRTAELANGALGMIELNDDYTLKKVMKPLIASNTVTDE
IERANVFKMNGKWYLFTDSRGSKMTIDGITSNDIYMLGYVSNSLTGPYKPLNKTGLVLK
MDLDPNDVTFTYSHFAVPQAKGNNVVITSYMTNRGFYADKQSTFAPSFLLNIKGKKTSV
VKDSILE"
rep_origin complement(2210..2798)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
rep_origin 2964..5717
/label=mycobacterial PAL5000ts replication origin
/note="mycobacterial PAL5000ts replication origin"
CDS complement(5956..6768)
/codon_start=1
/label=KanR
/note="aminoglycoside phosphotransferase"
/translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
KPHAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
SDFDDERNGWPVEQVWKEMHNLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
terminator complement(6929..6975)
/label=rrnB T1 terminator
/note="transcription terminator T1 from the E. coli rrnB
gene"
CDS complement(7040..8101)
/codon_start=1
/gene="che9c 61"
/product="Che9c 61"
/label=che9c 61
/protein_id="AGY30910.1"
/translation="MAENAVTKQDSPKAPETISQVLQVLVPQLARAVPKGMDPDRIARI
VQTEIRKSRNAKAAGIAKQSLDDCTQESFAGALLTSAALGLEPGVNGECYLVPYRDTRR
GVVECQLIIGYQGIVKLFWQHPRASRIDAQWVGANDEFHYTMGLNPTLKHVKAKGDRGN
PVYFYAIVEVTGAEPLWDVFTADEIRELRRGKVGSSGDIKDPQRWMERKTALKQVLKLA
PKTTRLDAAIRADDRPGTDLSQSQALALPSTVKPTADYIDGEIAEPHEVDTPPKSSRAQ
RAQRATAPAPDVQMANPDQLKRLGEIQKAEKYNDADWFKFLADSAGVKATRAADLTFDE
AKAVIDMFDGPNA"
gene complement(7040..8101)
/gene="che9c 61"
/label=che9c 61
CDS complement(8094..9038)
/codon_start=1
/gene="che9c 60"
/product="Che9c 60"
/label=che9c 60
/protein_id="AGY30909.1"
/translation="MSVPTQDGMHRFVDEDVYHADRGSLSVSGAKLLLPPSCPAKFRWE
MDNTRKPKKVWDFGHVAHKLVLGKGAEFEILDPEVHGLKADGTPSEKPTATGMWRKAEA
EARKQGKVPIHVDLFTKAYDMAEKVRQHPTAGPIFANPDGEAEVALYYTDPETGVRLRG
RIDWLTDDIDDYKTSMTANPAELKTKFYKLGYFMQAAWYIDLLVALGLAENPRFRFITQ
EKEPPYVVTPIQYDDEAIEEGRRRNRQAIRLYADCMESGKWPDYSDDVVTISLPSWGLP
RPQTVGDVVTDSYIYDTDPLEEADPIEGDYIYG"
gene complement(8094..9038)
/gene="che9c 60"
/label=che9c 60
CDS complement(9127..9768)
/codon_start=1
/gene="amiS"
/product="AmiS acetamidase regulator"
/label=amiS
/protein_id="AGY30908.1"
/translation="MGGVGLFYVGAVLIIDGLMLLGRISPRGATPLNFFVGGLQVVTPT
VLILQSGGDAAVIFAASGLYLFGFTYLWVAINNVTDWDGEGLGWFSLFVAIAALGYSWH
AFTAEADPAFGVIWLLWAVLWFMLFLLLGLGHDALGPAVGFVAVAEGVITAAVPAFLIV
SGNWETGPLPAAVIAVIGFAAVVLAYPIGRRLAAPSVTNPPPAALAATTR"
gene complement(9127..9768)
/gene="amiS"
/label=amiS
CDS complement(9773..10057)
/codon_start=1
/gene="amiD"
/product="AmiD acetamidase regulator"
/label=amiD
/protein_id="AGY30907.1"
/translation="MPTYTFRCSHCGPFDLTCAISERDAAATCPECRTPARRVFGSVGL
TTFTAGHHRAFDAASASAESPTVVKSIPAGADRPRAPRRNPGLPSLPRY"
gene complement(9773..10057)
/gene="amiD"
/label=amiD
CDS complement(10137..10565)
/codon_start=1
/gene="amiA"
/product="AmiA acetamidase regulator"
/label=amiA
/protein_id="AGY30906.1"
/translation="MAAGQQRRPNLLLPLVRLTHLAESAIERVLADSSLKIEDWRVLDE
LAGRRTVPMSDLAQATLITGPTLTRTVDRLVSQGIIYRTADLHDRRRVLVALTPRGRTL
RNRLVDAVAEAECAAFESCGLDVDQLRELVDTTSNLTS"
gene complement(10137..10565)
/gene="amiA"
/label=amiA
CDS 10668..11741
/codon_start=1
/gene="amiC"
/product="AmiC acetamidase regulator"
/label=amiC
/protein_id="AGY30905.1"
/translation="MQDGEVEFRVGLVIPLQGPAGIFAPSCEAVAELAAKEVNDRGGLQ
GRKVTIEVLDGGRPGDDVARTVADRLRGHGLDAVTGWHISAVRNRISPVVRDRIPYVYT
SLYEGGERTPGVFCTGETPQIQIAPALAWLRDHFGIRSWCLVGDDYIWPRRSAAAARAY
CRDLDLELRREIYVPYGTDDFRAPVRKAIASGAQAVLMLLVGQDAVLFNREFARAGGHD
RMARFSPLMEENMLLASGAGSTENLYVAAAYFSSLATAGAMDLMGSYVARYGADAPPLN
AMAESCYEGLVALEAIFQRAHSPEIPDLMASAHDVGFDGPRGPMCMRDSQFDQQVYIAS
ADGYDFDILDTLTTLDA"
gene 10668..11741
/gene="amiC"
/label=amiC