基本信息
| 启动子: | CspA promoter |
| 原核抗性: | 氨苄青霉素Amp |
| 克隆菌株: | 大肠杆菌DH5α |
| 培养条件: | 37℃，有氧，LB |
| 表达宿主: | 大肠杆菌 |
| 培养条件: | 37℃，有氧，LB |
| 诱导方式: | IPTG或乳糖及其类似物 |
质粒简介
pCold-SUMO-10×His-3C-Strep II-FLAG质粒是一个可以在低温环境下表达的大肠杆菌载体。质粒图谱
质粒序列
LOCUS Exported 4930 bp ds-DNA circular SYN 12-OCT-2017
DEFINITION synthetic circular DNA
KEYWORDS pCold-SUMO-10×His-3C-Strep II-FLAG
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 4930)
TITLE Direct Submission
FEATURES Location/Qualifiers
source 1..4930
/organism="synthetic DNA construct"
/mol_type="other DNA"
misc_feature 15..81
/label=cspA promoter
protein_bind 84..100
/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)."
CDS 271..343
/codon_start=1
/product="6xHis affinity tag"
/label=6xHis
/translation="HMNWSHPQFEKSSGSSGGHHHHHH"
CDS 365..653
/codon_start=1
/gene="S. cerevisiae SMT3 (truncated)"
/product="cleavable ubiquitin-like protein tag"
/label=SUMO
/translation="DSEVNQEAKPEVKPEVKPETHINLKVSDGSSEIFFKIKKTTPLRR
LMEAFAKRQGKEMDSLRFLYDGIRIQADQAPEDLDMEDNDIIEAHREQIG"
CDS 676..705
/codon_start=1
/product="10xHis affinity tag"
/label=10xHis
/translation="HHHHHHHHHH"
CDS 706..729
/codon_start=1
/product="recognition and cleavage site for human
rhinovirus 3C and PreScission proteases"
/label=HRV 3C site
/translation="LEVLFQGP"
CDS 775..798
/codon_start=1
/product="peptide that binds Strep-Tactin(R), an engineered
form of streptavidin"
/label=Strep-Tag II
/translation="WSHPQFEK"
CDS 850..873
/codon_start=1
/product="FLAG(R) epitope tag, followed by an enterokinase
cleavage site"
/label=FLAG
/translation="DYKDDDDK"
rep_origin complement(1234..1689)
/direction=LEFT
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
promoter 1789..1893
/gene="bla"
/label=AmpR promoter
CDS 1894..2754
/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"
rep_origin 2925..3513
/direction=RIGHT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
CDS complement(3685..4767)
/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"
promoter complement(4768..4845)
/gene="lacI"
/label=lacI promoter
ORIGIN
1 aaggaatggt gtggccgatt aatcataaat atgaaaaata attgttgcat cacccgccaa
61 tgcgtggctt aatgcacatc aaattgtgag cggataacaa tttgatgtgc tagcgcatat
121 ccagtgtagt aaggcaagtc ccttcaagag ttatcgttga tacccctcgt agtgcacatt
181 cctttaacgc ttcaaaatct gtaaagcacg ccatatcgcc gaaaggcaca cttaattatt
241 aagaggtaat acaccatgaa tcacaaagtg catatgaatt ggagccaccc gcagttcgaa
301 aaaagcagcg gcagcagcgg cggtcatcac catcatcatc acggcggcag cggcggcagc
361 gggtcggact cagaagtcaa tcaagaagct aagccagagg tcaagccaga agtcaagcct
421 gagactcaca tcaatttaaa ggtgtccgat ggatcttcag agatcttctt caagatcaaa
481 aagaccactc ctttaagaag gctgatggaa gcgttcgcta aaagacaggg taaggaaatg
541 gactccttaa gattcttgta cgacggtatt agaattcaag ctgatcaggc ccctgaagat
601 ttggacatgg aggataacga tattattgag gctcaccgcg aacagattgg aggcctggag
661 ctcggtaccc tcgagcatca ccatcaccat caccatcacc atcacctgga agttctgttc
721 caggggcccg aattctatgg caggaagaag cggagacagc gacgaagagg atcctggagc
781 cacccgcagt tcgagaaagc ggccgcatat ggcaggaaga agcggagaca gcgacgaaga
841 taagtcgacg attacaagga tgacgacgat aagtaatcta gataggtaat ctctgcttaa
901 aagcacagaa tctaagatcc ctgccattgg gcggggattt ttttatttgt tttcaggaaa
961 taaataatcg atcgcgtaat aaaatctatt attatttttg tgaagaataa atttgggtgc
1021 aatgagaatg cgcaggccct ttcgtctcgc gccgtttcgg tgatgacggt gaaaacctct
1081 gacacatgca gctcccggag acggtcacag cttgtctgta agcggatgcc gggagcagac
1141 aagcccgtca gggcgcgtca gcgggtgttg gcgggtgtcg gggctggctt aactatgcgg
1201 catcagagca gattgtactg agagtgcacc ataaaattgt aaacgttaat attttgttaa
1261 aattcgcgtt aaatttttgt taaatcagct cattttttaa ccaataggcc gaaatcggca
1321 aaatccctta taaatcaaaa gaatagcccg agatagggtt gagtgttgtt ccagtttgga
1381 acaagagtcc actattaaag aacgtggact ccaacgtcaa agggcgaaaa accgtctatc
1441 agggcgatgg cccactacgt gaaccatcac ccaaatcaag ttttttgggg tcgaggtgcc
1501 gtaaagcact aaatcggaac cctaaaggga gcccccgatt tagagcttga cggggaaagc
1561 cggcgaacgt ggcgagaaag gaagggaaga aagcgaaagg agcgggcgct agggcgctgg
1621 caagtgtagc ggtcacgctg cgcgtaacca ccacacccgc cgcgcttaat gcgccgctac
1681 agggcgcgta ctatggttgc tttgacgtat gcggtgtgaa ataccgcaca gatgcgtaag
1741 gagaaaatac cgcatcaggc gtcaggtggc acttttcggg gaaatgtgcg cggaacccct
1801 atttgtttat ttttctaaat acattcaaat atgtatccgc tcatgagaca ataaccctga
1861 taaatgcttc aataatattg aaaaaggaag agtatgagta ttcaacattt ccgtgtcgcc
1921 cttattccct tttttgcggc attttgcctt cctgtttttg ctcacccaga aacgctggtg
1981 aaagtaaaag atgctgaaga tcagttgggt gcacgagtgg gttacatcga actggatctc
2041 aacagcggta agatccttga gagttttcgc cccgaagaac gttttccaat gatgagcact
2101 tttaaagttc tgctatgtgg cgcggtatta tcccgtattg acgccgggca agagcaactc
2161 ggtcgccgca tacactattc tcagaatgac ttggttgagt actcaccagt cacagaaaag
2221 catcttacgg atggcatgac agtaagagaa ttatgcagtg ctgccataac catgagtgat
2281 aacactgcgg ccaacttact tctgacaacg atcggaggac cgaaggagct aaccgctttt
2341 ttgcacaaca tgggggatca tgtaactcgc cttgatcgtt gggaaccgga gctgaatgaa
2401 gccataccaa acgacgagcg tgacaccacg atgcctgtag caatggcaac aacgttgcgc
2461 aaactattaa ctggcgaact acttactcta gcttcccggc aacaattaat agactggatg
2521 gaggcggata aagttgcagg accacttctg cgctcggccc ttccggctgg ctggtttatt
2581 gctgataaat ctggagccgg tgagcgtggg tctcgcggta tcattgcagc actggggcca
2641 gatggtaagc cctcccgtat cgtagttatc tacacgacgg ggagtcaggc aactatggat
2701 gaacgaaata gacagatcgc tgagataggt gcctcactga ttaagcattg gtaactgtca
2761 gaccaagttt actcatatat actttagatt gatttaaaac ttcattttta atttaaaagg
2821 atctaggtga agatcctttt tgataatctc atgaccaaaa tcccttaacg tgagttttcg
2881 ttccactgag cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga tccttttttt
2941 ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg
3001 ccggatcaag agctaccaac tctttttccg aaggtaactg gcttcagcag agcgcagata
3061 ccaaatactg ttcttctagt gtagccgtag ttaggccacc acttcaagaa ctctgtagca
3121 ccgcctacat acctcgctct gctaatcctg ttaccagtgg ctgctgccag tggcgataag
3181 tcgtgtctta ccgggttgga ctcaagacga tagttaccgg ataaggcgca gcggtcgggc
3241 tgaacggggg gttcgtgcac acagcccagc ttggagcgaa cgacctacac cgaactgaga
3301 tacctacagc gtgagctatg agaaagcgcc acgcttcccg aagggagaaa ggcggacagg
3361 tatccggtaa gcggcagggt cggaacagga gagcgcacga gggagcttcc agggggaaac
3421 gcctggtatc tttatagtcc tgtcgggttt cgccacctct gacttgagcg tcgatttttg
3481 tgatgctcgt caggggggcg gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg
3541 ttcctggcct tttgctggcc ttttgctcac atagtcatgc cccgcgccca ccggaaggag
3601 ctgactgggt tgaaggctct caagggcatc ggtcgagatc ccggtgccta atgagtgagc
3661 taacttacat taattgcgtt gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc
3721 cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat tgggcgccag
3781 ggtggttttt cttttcacca gtgagacggg caacagctga ttgcccttca ccgcctggcc
3841 ctgagagagt tgcagcaagc ggtccacgct ggtttgcccc agcaggcgaa aatcctgttt
3901 gatggtggtt aacggcggga tataacatga gctgtcttcg gtatcgtcgt atcccactac
3961 cgagatatcc gcaccaacgc gcagcccgga ctcggtaatg gcgcgcattg cgcccagcgc
4021 catctgatcg ttggcaacca gcatcgcagt gggaacgatg ccctcattca gcatttgcat
4081 ggtttgttga aaaccggaca tggcactcca gtcgccttcc cgttccgcta tcggctgaat
4141 ttgattgcga gtgagatatt tatgccagcc agccagacgc agacgcgccg agacagaact
4201 taatgggccc gctaacagcg cgatttgctg gtgacccaat gcgaccagat gctccacgcc
4261 cagtcgcgta ccgtcttcat gggagaaaat aatactgttg atgggtgtct ggtcagagac
4321 atcaagaaat aacgccggaa cattagtgca ggcagcttcc acagcaatgg catcctggtc
4381 atccagcgga tagttaatga tcagcccact gacgcgttgc gcgagaagat tgtgcaccgc
4441 cgctttacag gcttcgacgc cgcttcgttc taccatcgac accaccacgc tggcacccag
4501 ttgatcggcg cgagatttaa tcgccgcgac aatttgcgac ggcgcgtgca gggccagact
4561 ggaggtggca acgccaatca gcaacgactg tttgcccgcc agttgttgtg ccacgcggtt
4621 gggaatgtaa ttcagctccg ccatcgccgc ttccactttt tcccgcgttt tcgcagaaac
4681 gtggctggcc tggttcacca cgcgggaaac ggtctgataa gagacaccgg catactctgc
4741 gacatcgtat aacgttactg gtttcacatt caccaccctg aattgactct cttccgggcg
4801 ctatcatgcc ataccgcgaa aggttttgcg ccattcgatg gtgtccggga tctcgacgct
4861 ctcccttatg cgactcctgc attaggaagc agcccagtag taggttgagg ccgttgagca
4921 ccgccgccgc
温馨提示：不可用于临床ZL。
