| 质粒名称: | pET22B_PcrA_2Bchimera |
|---|---|
| 出品公司: | 爱迪基因 |
| 目录编号: | 102998 |
| 存储实验室: | Martin Webb |
|
载体骨架基本信息 |
|
| 载体名称: | pET22B |
| 载体出品公司: | -- |
| 载体抗性: | 氨苄青霉素 |
| 空载体大小: | -- |
| 完整质粒大小: | -- |
| 载体修饰: | -- |
| 载体类型: | 细菌表达 |
| 筛选标记: | -- |
| 高拷贝/低拷贝: | 高拷贝 |
| 生长条件: | 37 ℃ |
|
插入基因信息 |
|
| 插入基因名称: | PcrA_2Bchimera |
| 物种来源: | Staphylococcus aureus, Bacilllus stearothermophilus |
| 插入大小: | -- |
| 基因突变: | Bacilllus stearothermophilus PcrA in which the 2B domain has been substituted with the equivalent sequence of Staphylococcus aureus PcrA. |
| GenBank编号 | -- |
| 启动子 | T7 |
| 标签/融合蛋白: | -- |
| 克隆方法: | 限制性内切酶 |
| 5' 克隆位点: | NdeI（销毁后未知） |
| 3' 克隆位点: | BamHI （销毁后未知） |
| 5' 测序引物 | T7 |
| 3' 测序引物 | -- |
|
文献引用方法 |
|
| 材料和方法部分: | pET22B_PcrA_2Bchimera was a gift from Martin Webb (Addgene plasmid # 102998 ; http://n2t.net/addgene:102998 ; RRID:Addgene_102998) |
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 pET22B_PcrA_2Bchimera 7584 bp ds-DNA circular SYN 12-MAY-2021
DEFINITION Express a DNA helicase, a chimeric construct of Bacilllus
stearothermophilus PcrA and 2B domain of Staphylococcus aureus PcrA.
ACCESSION .
VERSION .
KEYWORDS pET22B_PcrA_2Bchimera
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 7584)
TITLE Webb Lab plasmids
JOURNAL Unpublished
REFERENCE 2 (bases 1 to 7584)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58
http://www.snapgene.com
FEATURES Location/Qualifiers
source 1..7584
/organism="synthetic DNA construct"
/mol_type="other DNA"
primer_bind 1..20
/label=T7
/note="T7 promoter, forward primer"
promoter 1..19
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
protein_bind 20..44
/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)."
RBS 59..81
/note="efficient ribosome binding site from bacteriophage
T7 gene 10 (Olins and Rangwala, 1989)"
CDS 2316..2333
/codon_start=1
/product="6xHis affinity tag"
/label=6xHis
/translation="HHHHHH"
primer_bind complement(2386..2404)
/label=T7 Term
/note="T7 terminator, reverse primer"
terminator 2400..2447
/label=T7 terminator
/note="transcription terminator for bacteriophage T7 RNA
polymerase"
rep_origin 2484..2939
/direction=RIGHT
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
primer_bind complement(2571..2590)
/label=F1ori-R
/note="F1 origin, reverse primer"
primer_bind 2781..2802
/label=F1ori-F
/note="F1 origin, forward primer"
promoter 2965..3069
/gene="bla"
/label=AmpR promoter
CDS 3070..3930
/codon_start=1
/gene="bla"
/product="beta-lactamase"
/label=AmpR
/note="confers resistance to ampicillin, carbenicillin, and
related antibiotics"
/translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
LIKHW"
primer_bind complement(3288..3307)
/label=Amp-R
/note="Ampicillin resistance gene, reverse primer"
rep_origin 4101..4689
/direction=RIGHT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind 4590..4609
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
primer_bind 4843..4860
/label=L4440
/note="L4440 vector, forward primer"
misc_feature 4875..5014
/label=bom
/note="basis of mobility region from pBR322"
primer_bind complement(4940..4959)
/label=pRS-marker
/note="pRS vectors, use to sequence yeast selectable
marker"
primer_bind 5100..5122
/label=pGEX 3'
/note="pGEX vectors, reverse primer"
CDS complement(5116..5307)
/codon_start=1
/gene="rop"
/product="Rop protein, which maintains plasmids at low copy
number"
/label=rop
/translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
DELYRSCLARFGDDGENL"
CDS complement(6116..7198)
/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 7160..7179
/label=LacI-R
/note="LacI, reverse primer"
promoter complement(7199..7276)
/gene="lacI"
/label=lacI promoter
primer_bind 7482..7501
/label=pBRrevBam
/note="pBR322 vectors, tet region, downstream of BamHI,
reverse primer"
ORIGIN
1 taatacgact cactataggg gaattgtgag cggataacaa ttcccctcta gaaataattt
61 tgtttaactt taagaaggag atatacatat gaactttctg agcgaacagc tgctggcaca
121 tctgaataaa gaacagcaag aagcagttcg taccaccgaa ggtccgctgc tgattatggc
181 aggcgcaggt agcggtaaaa cccgtgttct gacccatcgt attgcatatc tgatggcaga
241 aaaacatgtt gcaccgtgga atattctggc aattaccttt accaataaag ccgcacgtga
301 aatgcgtgaa cgtgttcaga gcctgctggg tggtgcagca gaagatgttt ggattagcac
361 ctttcatagc atgtgtgttc gtattctgcg tcgtgatatt gatcgcattg gtattaatcg
421 caactttagc attctggacc cgaccgatca gctgagcgtt atgaaaacca ttctgaaaga
481 aaaaaacatc gatccgaaaa aattcgagcc tcgtaccatt ctgggcacca ttagcgcagc
541 aaaaaatgaa ctgctgcctc cggaacagtt tgcaaaacgt gcaagcacct attatgaaaa
601 agttgtgtcc gatgtgtacc aagaatatca gcaacgtctg ctgcgtaatc atagcctgga
661 ttttgatgat ctgattatga ccaccatcca gctgtttgat cgtgttccgg atgttctgca
721 ttattatcag tacaaattcc agtacatcca catcgatgaa tatcaggata ccaatcgtgc
781 acagtatacc ctggttaaaa aactggcaga acgctttcag aacatttgtg cagttggtga
841 tgcagatcag agcatttatc gttggcgtgg tgcagatatt caaaacattc tgagctttga
901 acgcgattat ccgaatgcca aagttattct gctggaacag aattatcgta gcaccaaacg
961 cattctgcag gcagcaaatg aagtgattga acataatgtg aaccgtaaac cgaaacgtct
1021 gtggaccgaa aatccggaag gtaaaccgat tctgtattat gaagcaatga acgaagcaga
1081 tgaagcacag tttgttgcag gtcgtattcg tgaagcagtt gaacgtggtg aacgtcgtta
1141 tcgtgatttt gcagttctgt atcgtaccaa tgcacagagc cgtgttatgg aagaaatgct
1201 gctgaaagca aatatcccgt atcagattgt tggtggccag aaattctatg atcgcaaaga
1261 aattaaagat ctgctgagct atctgcgcat tattgccaat agcaatgatg atattagcct
1321 gcagcgtatt atcaatgttc cgaaacgtgg tgttggtccg agcagcgttg aaaaagtgca
1381 gaattatgcc ctgcagaaca acatcagcat gtttgatgca ctgggtgaag cagatttcat
1441 tggtctgagc aaaaaagtta cccaagaatg cctgaacttc tatgaactga ttcagtccct
1501 gatcaaagaa caagaattcc tggaaatcca tgagatcgtt gatgaagtgc tgcagaaaag
1561 cggttatcgc gaaatgctgg aacgtgaaaa taccctggaa agccgtagcc gtctggaaaa
1621 tattgatgaa tttatgagcg tgccgaaaga ttatgaagaa aatacaccgc tggaagaaca
1681 gagcctgatc aattttctga ccgatctgag cctggttgca gatatcgatg aagcaaatgg
1741 caccgaacag gcagccgaag gtgatgccgt tatgctgatg accctgcatg ccgcaaaagg
1801 tctggaattt ccggttgtgt ttctgattgg catggaagaa ggtatttttc cgcataatcg
1861 ttccctggaa gatgatgatg aaatggaaga ggaacgtcgt ctggcctatg ttggtattac
1921 ccgtgcagaa gaagaactgg ttctgaccag cgcacagatg cgtaccctgt ttggtaacat
1981 tcagatgaat ccgcctagcc gttttctgaa tgaaattcct gcacatctgc tggaaaccgc
2041 aagccgtcgt caggctggtg ccagccgtcc ggcagttagt cgtccgcagg caagcggtgc
2101 cgttggtagc tggaaagtgg gtgatcgtgc aaatcatcgt aaatggggta ttggcaccgt
2161 tgttagcgtt cgtggtggcg gtgatgatca agaactggat attgcatttc cgagcccgat
2221 tggcattaaa cgcctgctgg caaaatttgc accgatcgaa aaagtgtaag gatccgaatt
2281 cgagctccgt cgacaagctt gcggccgcac tcgagcacca ccaccaccac cactgagatc
2341 cggctgctaa caaagcccga aaggaagctg agttggctgc tgccaccgct gagcaataac
2401 tagcataacc ccttggggcc tctaaacggg tcttgagggg ttttttgctg aaaggaggaa
2461 ctatatccgg attggcgaat gggacgcgcc ctgtagcggc gcattaagcg cggcgggtgt
2521 ggtggttacg cgcagcgtga ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc
2581 tttcttccct tcctttctcg ccacgttcgc cggctttccc cgtcaagctc taaatcgggg
2641 gctcccttta gggttccgat ttagtgcttt acggcacctc gaccccaaaa aacttgatta
2701 gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt
2761 ggagtccacg ttctttaata gtggactctt gttccaaact ggaacaacac tcaaccctat
2821 ctcggtctat tcttttgatt tataagggat tttgccgatt tcggcctatt ggttaaaaaa
2881 tgagctgatt taacaaaaat ttaacgcgaa ttttaacaaa atattaacgc ttacaattta
2941 ggtggcactt ttcggggaaa tgtgcgcgga acccctattt gtttattttt ctaaatacat
3001 tcaaatatgt atccgctcat gagacaataa ccctgataaa tgcttcaata atattgaaaa
3061 aggaagagta tgagtattca acatttccgt gtcgccctta ttcccttttt tgcggcattt
3121 tgccttcctg tttttgctca cccagaaacg ctggtgaaag taaaagatgc tgaagatcag
3181 ttgggtgcac gagtgggtta catcgaactg gatctcaaca gcggtaagat ccttgagagt
3241 tttcgccccg aagaacgttt tccaatgatg agcactttta aagttctgct atgtggcgcg
3301 gtattatccc gtattgacgc cgggcaagag caactcggtc gccgcataca ctattctcag
3361 aatgacttgg ttgagtactc accagtcaca gaaaagcatc ttacggatgg catgacagta
3421 agagaattat gcagtgctgc cataaccatg agtgataaca ctgcggccaa cttacttctg
3481 acaacgatcg gaggaccgaa ggagctaacc gcttttttgc acaacatggg ggatcatgta
3541 actcgccttg atcgttggga accggagctg aatgaagcca taccaaacga cgagcgtgac
3601 accacgatgc ctgcagcaat ggcaacaacg ttgcgcaaac tattaactgg cgaactactt
3661 actctagctt cccggcaaca attaatagac tggatggagg cggataaagt tgcaggacca
3721 cttctgcgct cggcccttcc ggctggctgg tttattgctg ataaatctgg agccggtgag
3781 cgtgggtctc gcggtatcat tgcagcactg gggccagatg gtaagccctc ccgtatcgta
3841 gttatctaca cgacggggag tcaggcaact atggatgaac gaaatagaca gatcgctgag
3901 ataggtgcct cactgattaa gcattggtaa ctgtcagacc aagtttactc atatatactt
3961 tagattgatt taaaacttca tttttaattt aaaaggatct aggtgaagat cctttttgat
4021 aatctcatga ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc agaccccgta
4081 gaaaagatca aaggatcttc ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa
4141 acaaaaaaac caccgctacc agcggtggtt tgtttgccgg atcaagagct accaactctt
4201 tttccgaagg taactggctt cagcagagcg cagataccaa atactgtcct tctagtgtag
4261 ccgtagttag gccaccactt caagaactct gtagcaccgc ctacatacct cgctctgcta
4321 atcctgttac cagtggctgc tgccagtggc gataagtcgt gtcttaccgg gttggactca
4381 agacgatagt taccggataa ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag
4441 cccagcttgg agcgaacgac ctacaccgaa ctgagatacc tacagcgtga gctatgagaa
4501 agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga
4561 acaggagagc gcacgaggga gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc
4621 gggtttcgcc acctctgact tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc
4681 ctatggaaaa acgccagcaa cgcggccttt ttacggttcc tggccttttg ctggcctttt
4741 gctcacatgt tctttcctgc gttatcccct gattctgtgg ataaccgtat taccgccttt
4801 gagtgagctg ataccgctcg ccgcagccga acgaccgagc gcagcgagtc agtgagcgag
4861 gaagcggaag agcgcctgat gcggtatttt ctccttacgc atctgtgcgg tatttcacac
4921 cgcaatggtg cactctcagt acaatctgct ctgatgccgc atagttaagc cagtatacac
4981 tccgctatcg ctacgtgact gggtcatggc tgcgccccga cacccgccaa cacccgctga
5041 cgcgccctga cgggcttgtc tgctcccggc atccgcttac agacaagctg tgaccgtctc
5101 cgggagctgc atgtgtcaga ggttttcacc gtcatcaccg aaacgcgcga ggcagctgcg
5161 gtaaagctca tcagcgtggt cgtgaagcga ttcacagatg tctgcctgtt catccgcgtc
5221 cagctcgttg agtttctcca gaagcgttaa tgtctggctt ctgataaagc gggccatgtt
5281 aagggcggtt ttttcctgtt tggtcactga tgcctccgtg taagggggat ttctgttcat
5341 gggggtaatg ataccgatga aacgagagag gatgctcacg atacgggtta ctgatgatga
5401 acatgcccgg ttactggaac gttgtgaggg taaacaactg gcggtatgga tgcggcggga
5461 ccagagaaaa atcactcagg gtcaatgcca gcgcttcgtt aatacagatg taggtgttcc
5521 acagggtagc cagcagcatc ctgcgatgca gatccggaac ataatggtgc agggcgctga
5581 cttccgcgtt tccagacttt acgaaacacg gaaaccgaag accattcatg ttgttgctca
5641 ggtcgcagac gttttgcagc agcagtcgct tcacgttcgc tcgcgtatcg gtgattcatt
5701 ctgctaacca gtaaggcaac cccgccagcc tagccgggtc ctcaacgaca ggagcacgat
5761 catgcgcacc cgtggggccg ccatgccggc gataatggcc tgcttctcgc cgaaacgttt
5821 ggtggcggga ccagtgacga aggcttgagc gagggcgtgc aagattccga ataccgcaag
5881 cgacaggccg atcatcgtcg cgctccagcg aaagcggtcc tcgccgaaaa tgacccagag
5941 cgctgccggc acctgtccta cgagttgcat gataaagaag acagtcataa gtgcggcgac
6001 gatagtcatg ccccgcgccc accggaagga gctgactggg ttgaaggctc tcaagggcat
6061 cggtcgagat cccggtgcct aatgagtgag ctaacttaca ttaattgcgt tgcgctcact
6121 gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc
6181 ggggagaggc ggtttgcgta ttgggcgcca gggtggtttt tcttttcacc agtgagacgg
6241 gcaacagctg attgcccttc accgcctggc cctgagagag ttgcagcaag cggtccacgc
6301 tggtttgccc cagcaggcga aaatcctgtt tgatggtggt taacggcggg atataacatg
6361 agctgtcttc ggtatcgtcg tatcccacta ccgagatatc cgcaccaacg cgcagcccgg
6421 actcggtaat ggcgcgcatt gcgcccagcg ccatctgatc gttggcaacc agcatcgcag
6481 tgggaacgat gccctcattc agcatttgca tggtttgttg aaaaccggac atggcactcc
6541 agtcgccttc ccgttccgct atcggctgaa tttgattgcg agtgagatat ttatgccagc
6601 cagccagacg cagacgcgcc gagacagaac ttaatgggcc cgctaacagc gcgatttgct
6661 ggtgacccaa tgcgaccaga tgctccacgc ccagtcgcgt accgtcttca tgggagaaaa
6721 taatactgtt gatgggtgtc tggtcagaga catcaagaaa taacgccgga acattagtgc
6781 aggcagcttc cacagcaatg gcatcctggt catccagcgg atagttaatg atcagcccac
6841 tgacgcgttg cgcgagaaga ttgtgcaccg ccgctttaca ggcttcgacg ccgcttcgtt
6901 ctaccatcga caccaccacg ctggcaccca gttgatcggc gcgagattta atcgccgcga
6961 caatttgcga cggcgcgtgc agggccagac tggaggtggc aacgccaatc agcaacgact
7021 gtttgcccgc cagttgttgt gccacgcggt tgggaatgta attcagctcc gccatcgccg
7081 cttccacttt ttcccgcgtt ttcgcagaaa cgtggctggc ctggttcacc acgcgggaaa
7141 cggtctgata agagacaccg gcatactctg cgacatcgta taacgttact ggtttcacat
7201 tcaccaccct gaattgactc tcttccgggc gctatcatgc cataccgcga aaggttttgc
7261 gccattcgat ggtgtccggg atctcgacgc tctcccttat gcgactcctg cattaggaag
7321 cagcccagta gtaggttgag gccgttgagc accgccgccg caaggaatgg tgcatgcaag
7381 gagatggcgc ccaacagtcc cccggccacg gggcctgcca ccatacccac gccgaaacaa
7441 gcgctcatga gcccgaagtg gcgagcccga tcttccccat cggtgatgtc ggcgatatag
7501 gcgccagcaa ccgcacctgt ggcgccggtg atgccggcca cgatgcgtcc ggcgtagagg
7561 atcgagatct cgatcccgcg aaat
//
