pJL-SaKKH-BE3
PVT10482 2ug
pJL-SaKKH-BE3 Information
Prokaryotic resistance: ampicillin Amp
Clone strain: Escherichia coli Stbl3
Culture conditions: 37 C
pJL-SaKKH-BE3 Description
PJL-SaKKH-BE3 is a rat mammalian expression plasmid.
pJL-SaKKH-BE3 Sequence
LOCUS Exported 7689 bp ds-DNA circular SYN 01-DEC-2017
DEFINITION synthetic circular DNA
FEATURES Location/Qualifiers
source 1..7689
/organism="synthetic DNA construct"
/mol_type="other DNA"
CDS 13..261
/codon_start=1
/product="uracil-DNA glycosylase inhibitor from a Bacillus
subtilis bacteriophage (Mol et al., 1995)"
/label=UGI
/translation="TNLSDIIEKETGKQLVIQESILMLPEEVEEVIGNKPESDILVHTA
YDESTDENVMLLTSDAPEYKPWALVIQDSNGENKIKML"
CDS 274..294
/codon_start=1
/product="nuclear localization signal of SV40 (simian virus
40) large T antigen"
/label=SV40 NLS
/translation="PKKKRKV"
CDS 303..320
/codon_start=1
/product="6xHis affinity tag"
/label=6xHis
/translation="HHHHHH"
polyA_signal 349..573
/label=bGH poly(A) signal
/note="bovine growth hormone polyadenylation signal"
rep_origin complement(1046..1634)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
CDS complement(1805..2665)
/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"
promoter complement(2666..2770)
/gene="bla"
/label=AmpR promoter
enhancer 3040..3419
/label=CMV enhancer
/note="human cytomegalovirus immediate early enhancer"
promoter 3420..3623
/label=CMV promoter
/note="human cytomegalovirus (CMV) immediate early
promoter"
promoter 3665..3683
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
CDS 3697..4383
/codon_start=1
/product="cytidine deaminase (C to U editing enzyme) from
rat"
/label=APOBEC-1
/note="can use ssDNA as a substrate (Komor et al., 2016)"
/translation="MSSETGPVAVDPTLRRRIEPHEFEVFFDPRELRKETCLLYEINWG
GRHSIWRHTSQNTNKHVEVNFIEKFTTERYFCPNTRCSITWFLSWSPCGECSRAITEFL
SRYPHVTLFIYIARLYHHADPRNRQGLRDLISSGVTIQIMTEQESGYCWRNFVNYSPSN
EAHWPRYPHLWVRLYVLELYCIILGLPPCLNILRRKQPQLTFFTIALQSCHYQRLPPHI
LWATGLK"
misc_feature 4432..7596
/label=SaKKHCas9n
CDS 7597..7617
/codon_start=1
/product="nuclear localization signal of SV40 (simian virus
40) large T antigen"
/label=SV40 NLS
/translation="PKKKRKV"
CDS 7624..7689
/codon_start=1
/product="three tandem FLAG(R) epitope tags, followed by an
enterokinase cleavage site"
/label=3xFLAG
/translation="DYKDHDGDYKDHDIDYKDDDDK"
ORIGIN
1 tctggtggtt ctactaatct gtcagatatt attgaaaagg agaccggtaa gcaactggtt
61 atccaggaat ccatcctcat gctcccagag gaggtggaag aagtcattgg gaacaagccg
121 gaaagcgata tactcgtgca caccgcctac gacgagagca ccgacgagaa tgtcatgctt
181 ctgactagcg acgcccctga atacaagcct tgggctctgg tcatacagga tagcaacggt
241 gagaacaaga ttaagatgct ctctggtggt tctcccaaga agaagaggaa agtctaaccg
301 gtcatcatca ccatcaccat tgagtttaaa cccgctgatc agcctcgact gtgccttcta
361 gttgccagcc atctgttgtt tgcccctccc ccgtgccttc cttgaccctg gaaggtgcca
421 ctcccactgt cctttcctaa taaaatgagg aaattgcatc gcattgtctg agtaggtgtc
481 attctattct ggggggtggg gtggggcagg acagcaaggg ggaggattgg gaagacaata
541 gcaggcatgc tggggatgcg gtgggctcta tggcttctga ggcggaaaga accagctggg
601 gctcgatacc gtcgacctct agctagagct tggcgtaatc atggtcatag ctgtttcctg
661 tgtgaaattg ttatccgctc acaattccac acaacatacg agccggaagc ataaagtgta
721 aagcctaggg tgcctaatga gtgagctaac tcacattaat tgcgttgcgc tcactgcccg
781 ctttccagtc gggaaacctg tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga
841 gaggcggttt gcgtattggg cgctcttccg cttcctcgct cactgactcg ctgcgctcgg
901 tcgttcggct gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg ttatccacag
961 aatcagggga taacgcagga aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc
1021 gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac gagcatcaca
1081 aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga taccaggcgt
1141 ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt accggatacc
1201 tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc
1261 tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc
1321 ccgaccgctg cgccttat
