pUAST Drosophila Gene Express對花ion Vector (User-defined pro老厭moter)

Our pUAST vector system is a well-c道懂haracterized and highly effec站笑tive system for generating t紙區ransgenic flies and co樹嗎ntrolling transgene ex業學pression. This system 個船is derived from 農歌the commonly used D熱站rosophila P-element transpo笑吃son and can be u冷個sed for achieving 生男ubiquitous, tissue-specific or i熱化nducible transgene expression喝來.

The complete pUAST system cons鄉小ists of two vectors, both engine站市ered as E. coli plasmids. One vector re頻件ferred to as the pUAST pl購件asmid, contains 討亮two P-element terminal repeats呢工 bracketing the 的為region/gene to be transpose年舊d. The other vecto不知r, referred to as the helper p紅女lasmid or transposase plasmid, en志水codes the P transposase.

When the pUAST and the transposase p離暗lasmid are co-injected into targ做業et cells, the trans那山posase produced fro自那m the helper plasmid r笑影ecognizes the two P-element termin務服al repeats on the pUAST plasmid湖新, and inserts the 喝亮flanked region including the 友算two P-element terminal r花鄉epeats into the host genome. Ins很樹ertion occurs without any 朋吧significant bias with respect t個公o insertion site sequence.

The P-element is a class-II tran光月sposon, meaning that i開近t moves in a cut-and-paste mann懂那er, hopping from place to place withou下船t leaving copies behind.空女 (In contrast, class-I transposon多錢s move in a copy-道多and-paste manner.) The transpositio工謝n creates 8 bp direct repea聽訊ts at the integration site in the ge好醫nome.

The pUAST system is common子紅ly used to gene熱志rate transgenic flies by c城吃o-injecting the pUAST and女得 the helper plasmid encoding the P爸海 transposase into Droso很暗phila early embryos. P transposa厭線se-mediated recombination betwe得友en the two P-element terminal repeat吧刀s leads to germline recombination e術南vents which produce transgenic offs友時pring carrying the user’s gene of i關大nterest. The P transposase will裡制 only be expressed for a short t少那ime, and with loss 拍什of the helper plasmid, the integration作校 of the transposon in the host ge吃器nome becomes permanent. The mini whi鐘白te gene on the pUAST vector encodes fo個一r eye color and acts as a mark訊請er for the identif金藍ication of transgenic flie可書s which have undergone succe不下ssful recombination of the transgene黃厭. PCR or other molecular methods can a鐘購lso be used to identify transgen那器ic cells or animals.

The user-defined promoter version of t北黃he pUAST Drosoph日知ila gene expression vector allows 雜身users to select a promoter of th吧關eir choice from熱空 our Drosophila promoter dat河到abase for driving the expression 短中of their gene of interes知報t (GOI) depending upon their exp物通erimental goal. Our Drosophila promot愛鄉er database offers t跳話he following promoter choices: ubiqui線女tous promoters including a冷笑ctin 5C, polyubiquitin an靜為d alpha-1 tubulin 在物for driving ubiq匠河uitous expression of th腦制e GOI; tissue-specific promoters su關藍ch as Rh2 for driving GOI expres路窗sion, specifically in Drosophila 信行ocelli and inducible promoters suc在雜h as Mtn for achiev技會ing inducible ex煙相pression of the GO有厭I with the presence of Cu+.

For further information校購 about this vector sy筆喝stem, please refer to the papers be裡綠low.