| 出品公司: | Invitrogen |
|---|---|
| 载体名称: | pHIL-D2, pHIL D2 |
| 质粒类型: | 毕赤酵母蛋白表达载体 |
| 表达水平: | 高拷贝 |
| 启动子: | AOX1 |
| 克隆方法: | 多克隆位点，限制性内切酶 |
| 载体大小: | 8210 bp |
| 5' 测序引物及序列: | 5´ AOX1:5´-GACTGGTTCCAATTGACAAGC-3´; |
| 3' 测序引物及序列: | 3´ AOX1:5´-GCAAATGGCATTCTGACATCC-3´ |
| 载体标签: | / |
| 载体抗性: | 氨苄 |
| 筛选标记: | HIS4 |
| 备注: |
细胞内蛋白表达载体
|
| 产品目录号: | K1710-01 |
| 稳定性: | 稳定 Stable |
| 组成型: | 组成型 Constitutive |
| 病毒/非病毒: | 非病毒 |
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
pHIL-D2载体的详细信息如下：
• pHIL-D2载体大小8,210 bp，是融合表达载体。
• 载体构建过程中，可供使用的单一限制性内切酶位点为EcoR I.
• 胞内表达蛋白基因。
• 在载体构建过程中，需要在基因上添加起始密码子ATG。
• 毕赤酵母中利用HIS4进行筛选。
• 为了将基因插入GS115或KM71的AOX区, 使用SacI限制性内切酶线性化质粒(GS115中产生His+Mut+基因型，KM71中产生His+ MutS基因型)
•为了将基因插入GS115或KM71的His4区, 使用Sal I或者Stu I限制性内切酶线性化质粒(GS115中产生His+ Mut+基因型，KM71中产生His+ MutS基因型)
•将基因插入GS115的AOX1区域，需要使用Bgl II线性化质粒 (产生His+MutS基因型)。
载体序列
LOCUS pHIL-D2 8210 bp ds-DNA circular SYN 05-二月-2013
DEFINITION Pichia pastoris HIS4 vector for methanol-inducible intracellular
expression of a protein.
ACCESSION .
VERSION .
KEYWORDS pHIL-D2
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
COMMENT This file is created by Vector NTI
http://www.biofeng.com/
COMMENT VNTAUTHORNAME|biofeng.com|
COMMENT The gene of interest can be inserted at the unique EcoRI site.
FEATURES Location/Qualifiers
source 1..8210
/organism="synthetic DNA construct"
/lab_host="Pichia pastoris"
/mol_type="other DNA"
promoter 14..953
/gene="Pichia pastoris AOX1"
/note="AOX1 promoter"
/note="inducible promoter, regulated by methanol"
/note="color: #ffffff; direction: RIGHT"
terminator 1031..1277
/gene="Pichia pastoris AOX1"
/note="AOX1 terminator"
/note="transcription terminator for AOX1"
/note="color: #ffffff"
CDS complement(1690..4224)
/codon_start=1
/gene="Pichia pastoris HIS4"
/locus_tag="
"
/product="multifunctional enzyme, required for histidine
biosynthesis"
/note="PpHIS4"
/note="auxotrophic marker for Pichia pastoris"
/note="color: #ff7f50"
misc_feature 4579..5335
/note="AOX1 3' fragment"
/note="region downstream of Pichia pastoris AOX1 gene"
/note="color: #ffffff"
promoter 5582..5686
/gene="bla"
/note="AmpR promoter"
/note="color: #ffffff; direction: RIGHT"
CDS 5687..6547
/codon_start=1
/gene="bla"
/product="beta-lactamase"
/note="AmpR"
/note="confers resistance to ampicillin, carbenicillin, and
related antibiotics"
rep_origin complement(6589..7044)
/direction=LEFT
/note="f1 ori"
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
/note="color: #ffff00"
rep_origin 7155..7743
/direction=RIGHT
/note="ori"
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
/note="color: #ffff00"
ORIGIN
1 agatcgcggc cgcgatctaa catccaaaga cgaaaggttg aatgaaacct ttttgccatc
61 cgacatccac aggtccattc tcacacataa gtgccaaacg caacaggagg ggatacacta
121 gcagcagacc gttgcaaacg caggacctcc actcctcttc tcctcaacac ccacttttgc
181 catcgaaaaa ccagcccagt tattgggctt gattggagct cgctcattcc aattccttct
241 attaggctac taacaccatg actttattag cctgtctatc ctggcccccc tggcgaggtc
301 atgtttgttt atttccgaat gcaacaagct ccgcattaca cccgaacatc actccagatg
361 agggctttct gagtgtgggg tcaaatagtt tcatgttccc aaatggccca aaactgacag
421 tttaaacgct gtcttggaac ctaatatgac aaaagcgtga tctcatccaa gatgaactaa
481 gtttggttcg ttgaaatgct aacggccagt tggtcaaaaa gaaacttcca aaagtcgcca
541 taccgtttgt cttgtttggt attgattgac gaatgctcaa aaataatctc attaatgctt
601 agcgcagtct ctctatcgct tctgaacccg gtggcacctg tgccgaaacg caaatgggga
661 aacaacccgc tttttggatg attatgcatt gtcctccaca ttgtatgctt ccaagattct
721 ggtgggaata ctgctgatag cctaacgttc atgatcaaaa tttaactgtt ctaaccccta
781 cttgacaggc aatatataaa cagaaggaag ctgccctgtc ttaaaccttt ttttttatca
841 tcattattag cttactttca taattgcgac tggttccaat tgacaagctt ttgattttaa
901 cgacttttaa cgacaacttg agaagatcaa aaaacaacta attattcgaa acgaggaatt
961 cgccttagac atgactgttc ctcagttcaa gttgggcact tacgagaaga ccggtcttgc
1021 tagattctaa tcaagaggat gtcagaatgc catttgcctg agagatgcag gcttcatttt
1081 tgatactttt ttatttgtaa cctatatagt ataggatttt ttttgtcatt ttgtttcttc
1141 tcgtacgagc ttgctcctga tcagcctatc tcgcagctga tgaatatctt gtggtagggg
1201 tttgggaaaa tcattcgagt ttgatgtttt tcttggtatt tcccactcct cttcagagta
1261 cagaagatta agtgagaagt tcgtttgtgc aagcttatcg ataagcttta atgcggtagt
1321 ttatcacagt taaattgcta acgcagtcag gcaccgtgta tgaaatctaa caatgcgctc
1381 atcgtcatcc tcggcaccgt caccctggat gctgtaggca taggcttggt tatgccggta
1441 ctgccgggcc tcttgcggga tatcgtccat tccgacagca tcgccagtca ctatggcgtg
1501 ctgctagcgc tatatgcgtt gatgcaattt ctatgcgcac ccgttctcgg agcactgtcc
1561 gaccgctttg gccgccgccc agtcctgctc gcttcgctac ttggagccac tatcgactac
1621 gcgatcatgg cgaccacacc cgtcctgtgg atctatcgaa tctaaatgta agttaaaatc
1681 tctaaataat taaataagtc ccagtttctc catacgaacc ttaacagcat tgcggtgagc
1741 atctagacct tcaacagcag ccagatccat cactgcttgg ccaatatgtt tcagtccctc
1801 aggagttacg tcttgtgaag tgatgaactt ctggaaggtt gcagtgttaa ctccgctgta
1861 ttgacgggca tatccgtacg ttggcaaagt gtggttggta ccggaggagt aatctccaca
1921 actctctgga gagtaggcac caacaaacac agatccagcg tgttgtactt gatcaacata
1981 agaagaagca ttctcgattt gcaggatcaa gtgttcagga gcgtactgat tggacatttc
2041 caaagcctgc tcgtaggttg caaccgatag ggttgtagag tgtgcaatac acttgcgtac
2101 aatttcaacc cttggcaact gcacagcttg gttgtgaaca gcatcttcaa ttctggcaag
2161 ctccttgtct gtcatatcga cagccaacag aatcacctgg gaatcaatac catgttcagc
2221 ttgagacaga aggtctgagg caacgaaatc tggatcagcg tatttatcag caataactag
2281 aacttcagaa ggcccagcag gcatgtcaat actacacagg gctgatgtgt cattttgaac
2341 catcatcttg gcagcagtaa cgaactggtt tcctggacca aatattttgt cacacttagg
2401 aacagtttct gttccgtaag ccatagcagc tactgcctgg gcgcctcctg ctagcacgat
2461 acacttagca ccaaccttgt gggcaacgta gatgacttct ggggtaaggg taccatcctt
2521 cttaggtgga gatgcaaaaa caatttcttt gcaaccagca actttggcag gaacacccag
2581 catcagggaa gtggaaggca gaattgcggt tccaccagga atatagaggc caactttctc
2641 aataggtctt gcaaaacgag agcagactac accagggcaa gtctcaactt gcaacgtctc
2701 cgttagttga gcttcatgga atttcctgac gttatctata gagagatcaa tggctctctt
2761 aacgttatct ggcaattgca taagttcctc tgggaaagga gcttctaaca caggtgtctt
2821 caaagcgact ccatcaaact tggcagttag ttctaaaagg gctttgtcac cattttgacg
2881 aacattgtcg acaattggtt tgactaattc cataatctgt tccgttttct ggataggacg
2941 acgaagggca tcttcaattt cttgtgagga ggccttagaa acgtcaattt tgcacaattc
3001 aatacgacct tcagaaggga cttctttagg tttggattct tctttaggtt gttccttggt
3061 gtatcctggc ttggcatctc ctttccttct agtgaccttt agggacttca tatccaggtt
3121 tctctccacc tcgtccaacg tcacaccgta cttggcacat ctaactaatg caaaataaaa
3181 taagtcagca cattcccagg ctatatcttc cttggattta gcttctgcaa gttcatcagc
3241 ttcctcccta attttagcgt tcaacaaaac ttcgtcgtca aataaccgtt tggtataaga
3301 accttctgga gcattgctct tacgatccca caaggtggct tccatggctc taagaccctt
3361 tgattggcca aaacaggaag tgcgttccaa gtgacagaaa ccaacacctg tttgttcaac
3421 cacaaatttc aagcagtctc catcacaatc caattcgata cccagcaact tttgagttgc
3481 tccagatgta gcacctttat accacaaacc gtgacgacga gattggtaga ctccagtttg
3541 tgtccttata gcctccggaa tagacttttt ggacgagtac accaggccca acgagtaatt
3601 agaagagtca gccaccaaag tagtgaatag accatcgggg cggtcagtag tcaaagacgc
3661 caacaaaatt tcactgacag ggaacttttt gacatcttca gaaagttcgt attcagtagt
3721 caattgccga gcatcaataa tggggattat accagaagca acagtggaag tcacatctac
3781 caactttgcg gtctcagaaa aagcataaac agttctacta ccgccattag tgaaactttt
3841 caaatcgccc agtggagaag aaaaaggcac agcgatacta gcattagcgg gcaaggatgc
3901 aactttatca accagggtcc tatagataac cctagcgcct gggatcatcc tttggacaac
3961 tctttctgcc aaatctaggt ccaaaatcac ttcattgata ccattattgt acaacttgag
4021 caagttgtcg atcagctcct caaattggtc ctctgtaacg gatgactcaa cttgcacatt
4081 aacttgaagc tcagtcgatt gagtgaactt gatcaggttg tgcagctggt cagcagcata
4141 gggaaacacg gcttttccta ccaaactcaa ggaattatca aactctgcaa cacttgcgta
4201 tgcaggtagc aagggaaatg tcatacttga agtcggacag tgagtgtagt cttgagaaat
4261 tctgaagccg tatttttatt atcagtgagt cagtcatcag gagatcctct acgccggacg
4321 catcgtggcc ggcatcaccg gcgccacagg tgcggttgct ggcgcctata tcgccgacat
4381 caccgatggg gaagatcggg ctcgccactt cgggctcatg agcgcttgtt tcggcgtggg
4441 tatggtggca ggccccgtgg ccgggggact gttgggcgcc atctccttgc atgcaccatt
4501 ccttgcggcg gcggtgctca acggcctcaa cctactactg ggctgcttcc taatgcagga
4561 gtcgcataag ggagagcgtc gagtatctat gattggaagt atgggaatgg tgatacccgc
4621 attcttcagt gtcttgaggt ctcctatcag attatgccca actaaagcaa ccggaggagg
4681 agatttcatg gtaaatttct ctgacttttg gtcatcagta gactcgaact gtgagactat
4741 ctcggttatg acagcagaaa tgtccttctt ggagacagta aatgaagtcc caccaataaa
4801 gaaatccttg ttatcaggaa caaacttctt gtttcgaact ttttcggtgc cttgaactat
4861 aaaatgtaga gtggatatgt cgggtaggaa tggagcgggc aaatgcttac cttctggacc
4921 ttcaagaggt atgtagggtt tgtagatact gatgccaact tcagtgacaa cgttgctatt
4981 tcgttcaaac cattccgaat ccagagaaat caaagttgtt tgtctactat tgatccaagc
5041 cagtgcggtc ttgaaactga caatagtgtg ctcgtgtttt gaggtcatct ttgtatgaat
5101 aaatctagtc tttgatctaa ataatcttga cgagccaagg cgataaatac ccaaatctaa
5161 aactctttta aaacgttaaa aggacaagta tgtctgcctg tattaaaccc caaatcagct
5221 cgtagtctga tcctcatcaa cttgaggggc actatcttgt tttagagaaa tttgcggaga
5281 tgcgatatcg agaaaaaggt acgctgattt taaacgtgaa atttatctca agatcgcggc
5341 cgcgatctcg aataataact gttatttttc agtgttcccg atctgcgtct atttcacaat
5401 accaacatga gtcagcttat cgatgataag ctgtcaaaca tgagaattaa ttcgatgata
5461 agctgtcaaa catgagaaat cttgaagacg aaagggcctc gtgatacgcc tatttttata
5521 ggttaatgtc atgataataa tggtttctta gacgtcaggt ggcacttttc ggggaaatgt
5581 gcgcggaacc cctatttgtt tatttttcta aatacattca aatatgtatc cgctcatgag
5641 acaataaccc tgataaatgc ttcaataata ttgaaaaagg aagagtatga gtattcaaca
5701 tttccgtgtc gcccttattc ccttttttgc ggcattttgc cttcctgttt ttgctcaccc
5761 agaaacgctg gtgaaagtaa aagatgctga agatcagttg ggtgcacgag tgggttacat
5821 cgaactggat ctcaacagcg gtaagatcct tgagagtttt cgccccgaag aacgttttcc
5881 aatgatgagc acttttaaag ttctgctatg tggcgcggta ttatcccgtg ttgacgccgg
5941 gcaagagcaa ctcggtcgcc gcatacacta ttctcagaat gacttggttg agtactcacc
6001 agtcacagaa aagcatctta cggatggcat gacagtaaga gaattatgca gtgctgccat
6061 aaccatgagt gataacactg cggccaactt acttctgaca acgatcggag gaccgaagga
6121 gctaaccgct tttttgcaca acatggggga tcatgtaact cgccttgatc gttgggaacc
6181 ggagctgaat gaagccatac caaacgacga gcgtgacacc acgatgcctg cagcaatggc
6241 aacaacgttg cgcaaactat taactggcga actacttact ctagcttccc ggcaacaatt
6301 aatagactgg atggaggcgg ataaagttgc aggaccactt ctgcgctcgg cccttccggc
6361 tggctggttt attgctgata aatctggagc cggtgagcgt gggtctcgcg gtatcattgc
6421 agcactgggg ccagatggta agccctcccg tatcgtagtt atctacacga cggggagtca
6481 ggcaactatg gatgaacgaa atagacagat cgctgagata ggtgcctcac tgattaagca
6541 ttggtaactg tcagaccaag tttactcata tatactttag attgatttaa attgtaaacg
6601 ttaatatttt gttaaaattc gcgttaaatt tttgttaaat cagctcattt tttaaccaat
6661 aggccgaaat cggcaaaatc ccttataaat caaaagaata gaccgagata gggttgagtg
6721 ttgttccagt ttggaacaag agtccactat taaagaacgt ggactccaac gtcaaagggc
6781 gaaaaaccgt ctatcagggc gatggcccac tacgtgaacc atcaccctaa tcaagttttt
6841 tggggtcgag gtgccgtaaa gcactaaatc ggaaccctaa agggagcccc cgatttagag
6901 cttgacgggg aaagccggcg aacgtggcga gaaaggaagg gaagaaagcg aaaggagcgg
6961 gcgctagggc gctggcaagt gtagcggtca cgctgcgcgt aaccaccaca cccgccgcgc
7021 ttaatgcgcc gctacagggc gcgtaaaagg atctaggtga agatcctttt tgataatctc
7081 atgaccaaaa tcccttaacg tgagttttcg ttccactgag cgtcagaccc cgtagaaaag
7141 atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa
7201 aaaccaccgc taccagcggt ggtttgtttg ccggatcaag agctaccaac tctttttccg
7261 aaggtaactg gcttcagcag agcgcagata ccaaatactg tccttctagt gtagccgtag
7321 ttaggccacc acttcaagaa ctctgtagca ccgcctacat acctcgctct gctaatcctg
7381 ttaccagtgg ctgctgccag tggcgataag tcgtgtctta ccgggttgga ctcaagacga
7441 tagttaccgg ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac acagcccagc
7501 ttggagcgaa cgacctacac cgaactgaga tacctacagc gtgagcattg agaaagcgcc
7561 acgcttcccg aagggagaaa ggcggacagg tatccggtaa gcggcagggt cggaacagga
7621 gagcgcacga gggagcttcc agggggaaac gcctggtatc tttatagtcc tgtcgggttt
7681 cgccacctct gacttgagcg tcgatttttg tgatgctcgt caggggggcg gagcctatgg
7741 aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct tttgctggcc ttttgctcac
7801 atgttctttc ctgcgttatc ccctgattct gtggataacc gtattaccgc ctttgagtga
7861 gctgataccg ctcgccgcag ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg
7921 gaagagcgcc tgatgcggta ttttctcctt acgcatctgt gcggtatttc acaccgcata
7981 tggtgcactc tcagtacaat ctgctctgat gccgcatagt taagccagta tacactccgc
8041 tatcgctacg tgactgggtc atggctgcgc cccgacaccc gccaacaccc gctgacgcgc
8101 cctgacgggc ttgtctgctc ccggcatccg cttacagaca agctgtgacc gtctccggga
8161 gctgcatgtg tcagaggttt tcaccgtcat caccgaaacg cgcgaggcag
//
