轉導克隆 的英文怎麼說
中文拼音 [zhuǎndǎokèlōng]
轉導克隆
英文
transduction clone-
The total rna was isolated from pokeweed ( phytolacca americana ) leaves using the method of guanidine isothiocyante and used as template to amplify the total length and deleted mutant pokeweed antiviral protein ( pap ) gene by rt - pcr and then the pap gene was cloned into pgem - t vector. the sequencing results showed that pap gene had 99. 9 % identity comparing with the pap gene nucleotide sequence reported by lin et al ( 1991 ). the iptg - inducible expression vector containing the pap gene was constructed and transferred into e. coli bl21 ( de3 ) - plyss
將缺失型pap基因克隆到植物表達載體pbi121中，通過液氮冷凍法將重組質粒轉入農桿菌lba4404細胞中，然後採用葉盤法，在該農桿菌的介導下將pap基因導入普通煙草中，經過卡那黴素抗性篩選，最後獲得了轉pap基因的工程煙草植株，摩擦接種煙草花葉病毒（ tmv ） ，與非轉基因煙草相比，能夠推遲癥狀表現達2月之久，說明pap基因能夠在其它植物體內產生有活性的高抗病毒的蛋白質。Finally 4 right clones was obtained from 864 transformants by pcr detection. sequencing analysis showed that the hsa gene have been introduced into the right position of the human a - lactalbumin yac. furthermore, works to optimize the conditions of introducing the recombinant yac into goat f ibroblast via cell fusion was explorded
經pcr檢測，從864個轉化子中獲得了4個陽性克隆，測序表明，人血清白蛋白基因已正確的導入到人-乳白蛋白基因yac的特定位點上，並獲得了可進行細胞融合的重組yac載體。In this paper, first strand cdna of 3abc gene was synthesized using template rna extracted from cells infected with fmdv. the complete 3abc gene about isoobp was amplified by pcr and ligated into pgem - t easy vector. after transforming e. coli dh5 a, ampicillin resistant colonies were isolated and plasmid dna was prepared and analyzed by restriction analysis and pcr. presence of the full length 3abc gene was verified by nucleotide sequence analysis and the plasmid containing the expected sequence was named as pgem - 3abc. comparing the aquired sequence of 3abc with that of reference strains, the homology is more than 99 percent. the pgem - 3abc was digested with sal i and bgl ii and ligated into xho i and bgl ii - digested expression vector ptriex - 4 neo. lt was identified by restriction analysis and pcr and sequencing that this fragment had a 17bp deletion hi the nucleotide sequence 708bp of 3abc gene, which happened to form a terminator codon behind 3ab gene, but it contained the complete open reading frame ( orf ) of 3ab gene. positive clones were selected and induced with lmmol / l isopropyl - d - galactoside ( iptg ), bacteria were detected by sds - page and western blotting after properly treated. the results showed that the 3ab gene expressed successfully in e. coli and 33. 5ku fusion protein can be recognized by the positive bovine serum of fmdv. the amount of target protein is over 26 % of the total bacteria protein by gel thin layer scanning analysis
擴增產物連接到pgem - teasy載體中，轉化大腸桿菌dh5菌株，篩選氨芐青霉素抗性菌落，提取質粒經酶切鑒定、 pcr分析以及確證性測序證明，所克隆的1500bp左右的片段含有完整的3abc基因，與國外參考序列相比，同源性在99以上。將重組質粒pgem - 3abc和表達載體ptriex - 4neo分別用sal和bgl與xho和bgl消化后，亞克隆3abc基因至原核表達載體ptriex - 4neo中，通過酶切鑒定、 pcr擴增以及序列分析，發現克隆到ptriex - 4neo載體上的片段於3abc基因708bp處出現了17bp的缺失，碰巧在3ab基因后形成一終止密碼子，但3ab基因的閱讀框架完整，選出含有3ab基因完整閱讀框架的陽性克隆，用iptg誘導表達，收集菌液進行sds - page電泳、 westernblotting分析，結果表明， 3ab基因在大腸桿菌中成功表達，其表達產物為分子量33 . 5ku的融合蛋白，並能被口蹄疫病毒陽性血清識別。經薄層掃描分析，表達量占總蛋白量的26以上。Marek " s disease virus ( mdv ) phosphoprotein 24 ( pp24 ) gene was amplified from md11 strain by polymerase chain reaction ( pcr ). then we cloned it into the downstream of gst gene according to the right open reading frame ( orf ) in pgex - 6p - l vector
本研究將型mdvmd11株的pp24基因的完整orf克隆入原核表達載體pgex - 6p - 1中，重組質粒pgex - pp24轉化bl21宿主菌后，經iptg誘導表達。As a popular vegetable, tomato is rich in vatamin a, b, p and other nutrients ; among them, lyxopene can prevent prostate gland tumor. so introducing stilbene synthase gene into tomato to get new health - protection tomato will have important social and economic effect. our research is made up of four parts ; the cloning and sequencing of stilbene synthase ; tomato genetic transforming mediated by agrobacterium - lba4404 ; quantification of resveratrol in transgenic tomato by hplc analysis
本研究包括四個方面的內容：芪合酶基因（ stilbenesynthasegene ）的克隆與全序列分析；芪合酶基因（ stilbenesynthasegene ）植物表達載體的構建；農桿菌lba4404介導的番茄遺傳轉化： hplc技術分析轉基因植株中表達白藜蘆醇的含量。The hwtx - i gene was chemically synthesized according to its known cdna sequence, the gene was inserted into vector ppic9k which contained aoxj promotor and the sequence of a secreting signal peptide - a - factor, the cloning ppic9k / hwtx - i was constructed and confirmed by two - step pcr and dna sequence analysis, then it was transformed into host strain gs115, a his + muts cell line was screened and multicopy transformants were screened by various g418 concentrations, the multicopy transformant was named gh1. gh1 was cultivated in flasks. after 6 days of induction by 0. 5 % methanol, the supernatant was checked by 16. 5 % tricine - sds page, which showed there was a band in the position of 3. 5 - 6. 1kd, then it was isolated and desalted by ultrofiltration followed by ion exchange of cm column, after reverse phase hplc of ci8 and vacuum drying, the purified rhwtx - 1 was obtained which was proved to be correct recombinant hwtx - i by tricine sds - page, maldi - tof mass spectrometry, amino acid composition analysis, the n - terminal amino acid sequence and its biological activity, the final field of the purified rhwtx - i was about 80mg / l, accounting for 23. 6 % of it total secretory proteins
將帶有hwtx -基因的ppic9k經blgii線性化后，轉化酵母宿主菌gs115原生質體后經篩選陽性克隆並經表型鑒定為his ~ + mut ~ s酵母菌，進一步用遺傳毒素g418篩選多拷貝的轉化菌株，命名為gh1 ；將gh1甲醇酵母菌用0 . 5的甲醇誘導表達，發酵上清經90飽和度的（ nh _ 4 ） _ 2so _ 4沉澱， yw - 3 （ mwc03000 ）的超濾膜超濾，再經cm陽離子交換， c _ （ 18 ）反相hplc純化得到分子量為4kd左右的組分，其中4289 . 05的組分經質譜鑒定，氨基酸組成分析和序列測定為正確的表達產物，生物學活性表明其活性為天然毒素活性70 % ，表達量為80mg / l 。Ssmapkk transformations were screened on media with kanamycin ( 30mg / l ). nineteen individual kanamycin resistant plants were obtained. t2 plants were checked for integration of foreign gene by counting ratio of the number of tolerant plants to the number of non - tolerant plants on selection medium with kanamycin ( 30mg / l )
將ssvp和ssmapkk的全長cdna分別克隆入植物表達載體pcambia1300和prok中，導入根瘤農桿菌gv3101后，由花浸泡法進行擬南芥遺傳轉化，轉化ssvp鹽地堿蓬ssop和ssmapkk基因的克隆與功能鑒定的擬南芥在含潮黴素（ 25mg ）的ms培養基上篩選，獲得t ；代轉基因植株。Our previous work verified that heterotrimeric g protein was involved in regulation of pollen germination, tube growth and in the transmembrane mechanism of extracellular cam signal. in the present study, we provide further evidence that plc signaling pathway was presented in lily pollen and involved in extracellular cam signal transduction mechanism in pollen by molecular, biochemical and physiological approaches. these evidence included : the pip _ ( 2 ) - dependent plc activity was also detected in purified lily pollen protoplasts
為了給上述信號轉導過程提供進一步的證據，本文以百合花粉為材料，從plc活性的測定、 plc基因的克隆及其與花粉萌發的關系、以及plc活性被g蛋白和cam調控等方面，為肌醇磷脂系統參與花粉萌發和細胞外鈣調素的信號轉導過程進一步提供了分子生物學和生物化學的證據。Both the mature genes of gloshedobin and gussurobin were cloned into the vector pet - 32a ( + ), strain bl21 ( de3 ), to study their expression in prokaryotic cell. the gene was expressed under t7 promoter with a fusion protein partner of thx. tag and a 6x his. tag at its n - terminal. having been induced by iptg for 4 hours, the recombinant enzyme was examined in the cytoplasm by sds - page analysis
將大連蛇島蝮蛇和長白山白眉蝮蛇毒類凝血酶基因分別克隆到大腸桿菌表達載體pet - 32 （ a ） +中，在t7啟動子下表達出融合蛋白，融合部分為硫氧還蛋白，位於類凝血酶基因上游，並在其n端帶6xhistag標簽以利於表達產物的分離純化，經熱激轉化至宿主菌bl21 （ de3 ）中， iptg誘導斗小時后收獲菌體。The transfermants with highest resistance to g418 ( 4 g / l in ypd ) were screened to study their expression level in 150 ml shaking flask. having been induced with methanol for 36 hours, the target enzyme could be examined in the supernatant by measuring amidolytic activity
首次將大連蛇島賅蛇毒類凝血酶成熟基回克隆到表達載體ppicgk中，經電激轉化至畢氏酵母菌株gs15中，再經甲醇誘導，在150ml搖瓶畔1獲得細胞外分泌表達產物。A binary plant expression vector with osg6b " driving report gene gus was constructed and transferred into tobacco via agrobacterium tumefaciens mediation
將克隆的啟動子與報告基因gus相連，構建植物表達載體，通過農桿菌介導轉化煙草。Multicopy integrants were screened with g418 from pichia pastoris which contains recombinant plasmid, and induced with methanol to secrete interesting peptide. the supernate of pichia pastoris culture was analysed by sds - page and western blotting. a reactive band, which the apparent molecular weight is 36kd, can be detected with sheep anti - hcmv polyclonal antibodies
重組質粒轉化巴氏畢赤酵母， g418篩選出多拷貝插入的單克隆，甲醇誘導多拷貝插入的單克隆酵母細胞分泌目的蛋白，培養液上清經sds - page電泳分析，在蛋白質印跡中檢測到培養液上清有一表觀分子量為36kd ，能與羊抗hcmv多克隆抗體發生發應的條帶。The e2 genes above of the prevalent strain ( guangxi yulin strain ) were cloned respectively into secreted expression vector ppic9k of eukaryotic expression system p. pastoris and transformed into p. pastoris by electroporation after linearization, 25 high - copied transformants were obtained by g418 screening. it was proved that the e2 genes were integrated stably into chromosome of p. pastoris by dot blot and dna sequencing
豬瘟病毒e2基因的真核表達：分別將csfv兩個代表株的e2基因克隆入畢赤酵母（ p . pastoris ）分泌型表達載體ppic9k中，酶切線型化后電穿孔導入p . pastotis進行整合，經g418篩選得到25個高拷貝轉化子，經dna斑點試驗和dna測序證明外源基因e2穩定地整合到p . pastoris染色體中。The chloroplast shsp gene was screened from the cdna library of tomato flower by pcr strategy and confirmed by sequencing. but difference was found at 3 bases of the sequence from the reported in genbank. then, an integrated vector prok ii of the chloroplast shsp gene and nptii gene ( a kanamycin resistant gene ) with camv35s promoter was constructed and introduced into tomato mediated by agrobacterium tumefaciens lba4404. transgenic tomato were screened by their ability of growing on media containing kanamycin
本實驗採用pcr方法從番茄花cdna文庫中克隆到葉綠體shsp基因，經測序證實與genbank中已發表的序列在編碼區相差2個堿基，其中一個堿基導致1個氨基酸的改變。將葉綠體shsp基因定向克隆于帶有組成性表達啟動子camv35s的植物表達載體prok中，凍融法轉化農桿菌lba4404 ，利用葉圓盤法對番茄進行ti質粒介導的遺傳轉化。The total rna was purified from the germ in the liquid by the guanidine isothiocyantehod method, then the total rna digested by dnase that had not rnase was used for rt - pcr. i change the magnesium ion dencity in the pcr system in order to optimize the pcr condition. at the end i selected the magnesium ion density as 1. 25 mm. the production of rt - pcr was inserted directionally into pgem ? z ( ampr ). the pgem ? z ( ampr ) was used to transform e coli jm109. i got a positive clone through culling and identificatin. the dna sequence inserted into pgem ? z ( ampr ) was sequenced and blasted with the cdna sequence of the # - mannanase mature peptide that got from genbank
分取誘導培養液中的菌體，用異硫氰酸胍法提取總rna ，總rna再經無rna酶的dna酶處理後用于rt ? pcr 。在pcr擴增目的基因時，通過優選擴增體系，使鎂離子濃度為1 . 25mm時rt ? pcr可順利地獲得目的基因，並能定向克隆到載體pgem ? 3z （ amp ~ r ）中。用克隆載體轉化宿主大腸桿菌jm109 ，通過篩選獲取陽性克隆子，對陽性克隆子進行酶切與pcr鑒定，並對載體中插入的目的基因進行測序。4, an intron sequence was also inserted upstream of gfpmut3 and its six reading frame could all be stopped, which could guarantee gfp translation in right reading frame
將藍色熒光蛋白基因bfp克隆到pet - 11c上，轉化bl21 （ de3 ）后實現了bfp在大腸桿菌中的誘導表達。In this study, we designed a pair of primers based on the sequence of the upstream and downstream of chicken il - 2 gene. about 600 bp chicken il - 2 cdna fragment was cloned from cona - stimulated chicken splenocytes by reverse transcription - polymerase chain reaction ( rt - pcr ) and was subcloned into puc18 vector. recombinant clone was demonstrated by restriction enzyme digestion and dna sequencing. next, we construct recombinant plasmid pproex ? t - il - 2. the cdna of chicken il - 2 gene was subcloned into bamh i / hind iii sites of vector. the recombinant plasmid pproex ? t - il - 2 was transformed into e. coli dh5a and the bacteria was induced with iptg. it was demonstrated by sds - page and western blot that a 18kda protein which was equal to chicken il - 2 protein in molecular weight was expressed in e. coli dh5a. the expression level was up to 30 % of the total bacterial proteins. the purified protein was used to prepare the antibody against chicken il - 2 protein
經酶切鑒定及dna序列測定，該基因為雞il - 2基因，其序列與sundick等報道的完全一致。在此基礎上，我們把雞il - 2基因亞克隆到大腸桿菌原核表達載體pproex ~ （ tm ） ht中，構建重組表達質粒並進行確證性序列測定，重組質粒測序結果表明將編碼雞il - 2成熟蛋白的基因正確地插入到原核表達載體pproex ~ （ tm ） ht的目的位點。重組質粒轉化受體菌dh5後用iptg於37進行誘導培養， sds - page和westernblot分析顯示，表達的雞il - 2融合蛋白分子量約為18kda ，表達的融合蛋白經薄層掃描發現目的蛋白表達量約占菌體蛋白的30 。900 bp promoter - directed gus expression was highly induced by sa and bth, while the 603 bp promoter, whether mutated or not, did not respond to sa and bth induction, which indicated that the element in response to sa and bth lied among 575 ~ 872 bp from transcription start site
全長900bp啟動子能夠應答sa和bth的誘導，而603bp長的啟動子無論突變與否對sa和bth均無應答，證明sa和bth的應答區域在克隆啟動子的轉錄起始位點上游575 872bp之間。The plasmids pci - mbl54 containing full length of mutations mbl cdna were propagated hi escherichia coli xl - 1 blue, then the extracted and purified pci - mbl54 were used to transfect dhfr ( - ) chinese - hamster ovary ( cho ) cells. after screeened with g418 and cloned, 4 g418 - resistent clones were randomly selected for detection of mrna expression by rt - pcr and molecular beacons. it was found that all of the 4 positive cell clones expresse mbl analogue as detected in transcription level
抽提、鑒定、純化重組質粒后，脂質體轉染法將重組質粒導入中國倉鼠卵巢細胞（ cho - dhfr ~ - ）中， g418選擇轉染子並克隆化培養，經rt - pcr和分子燈塔探針雜交鑒定其mrna轉錄，獲得4株穩定表達54位密碼突變型mbl的cho細胞。The hplap induced from the clone with two copies of hplap gene had higher enzyme activity. the clone with more or less copy was unfavourable to the hplap expression in p. pastoris
對部分克隆進行誘導表達發現只有2拷貝轉化子表達產物hplap活性最高，過高過低拷貝數均不利丁hplap的表達。分享友人