pHIL-D2
pHIL-D2
pHIL-D2
编号 | 载体名称 |
北京华越洋生物VECT2800 | pHIL-D2 |
pHIL-D2载体基本信息
| 出品公司: | -- |
|---|---|
| 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: | -- |
| 稳定性: | 稳定 Stable |
| 组成型: | 组成型 Constitutive |
| 病毒/非病毒: | 非病毒 |
pHIL-D2载体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)。
pHIL-D2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 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
