在生命系統(tǒng)中，基因的功能可通過系統(tǒng)性敲低、敲除或過表達來確定，此實驗方法被稱為功能基因組學，目前是眾多生物學研究領域的關鍵發(fā)現(xiàn)工具，如藥物靶點鑒定、耐藥性、宿主-病原互作和生物通路分析等。因此，功能基因高通量篩選文庫應運而生，今天聚焦96/384孔板陣列核酸片段文庫，來看看這類文庫的高通量篩選流程及其多重應用方向。

陣列核酸片段文庫

將siRNA、sgRNA與自動化、生物分析、高內(nèi)涵數(shù)據(jù)捕獲和生物統(tǒng)計分析相結(jié)合，篩選表型讀數(shù)可在標準設備上進行，如酶標儀；也可選用多種終點分析法，如比色、熒光或發(fā)光測定來評估細胞增殖、蛋白分泌、報告基因活性和凋亡誘導等情況。

1.產(chǎn)品特點

●類型豐富：提供人、小鼠全基因組和各基因家族siRNA, sgRNA文庫

●效果保證：

①siRNA：每個基因提供4條siRNA，保證mRNA水平上每個基因的4條siRNA中有3條有≥75%沉默效率

②sgRNA：每個基因提供3條sgRNA，保證每條sgRNA都能成功編輯目標位點

●應用廣泛：適用于多種表型篩選，避免不同狀態(tài)細胞間的旁分泌作用

●定制靈活：可挑選感興趣基因定制

ON-TARGETplus siRNA顯著降低脫靶效應，同時保持高水平的靶基因敲低作用

在人的原代CD4+ T細胞中進行Cas9 RNP電轉(zhuǎn)，每個基因使用一條預制sgRNA，流式細胞術分析對應蛋白敲除效果

2.篩選操作流程

siRNA文庫反向轉(zhuǎn)染流程示意圖

3.應用方向

*這里以siRNA文庫為例，更多sgRNA文庫應用案例可自行檢索

4.優(yōu)寧維文庫好物推薦

*還可提供小鼠相關文庫，可聯(lián)系您身邊的優(yōu)寧維銷售進一步咨詢

Reference

1.Brass, Abraham L et al. “Identification of host proteins required for HIV infection through a functional genomic screen.” Science (New York, N.Y.) vol. 319,5865 (2008): 921-6. doi:10.1126/science.1152725

2.Brass, Abraham L et al. “The IFITM proteins mediate cellular resistance to influenza A H1N1 virus, West Nile virus, and dengue virus.” Cell vol. 139,7 (2009): 1243-54. doi:10.1016/j.cell.2009.12.017

3.Yang, Penghe et al. “PSMD14 stabilizes estrogen signaling and facilitates breast cancer progression via deubiquitinating ERα.” Oncogene, 10.1038/s41388-023-02905-1. 29 Nov. 2023, doi:10.1038/s41388-023-02905-1

4.Li, Wenyang et al. “Deubiquitinase USP20 promotes breast cancer metastasis by stabilizing SNAI2.” Genes & development vol. 34,19-20 (2020): 1310-1315. doi:10.1101/gad.339804.120

5.Jurga, Mateusz et al. “USP11 controls R-loops by regulating senataxin proteostasis.” Nature communications vol. 12,1 5156. 15 Sep. 2021, doi:10.1038/s41467-021-25459-w

6.Kasahara, Kousuke et al. “EGF receptor kinase suppresses ciliogenesis through activation of USP8 deubiquitinase.” Nature communications vol. 9,1 758. 22 Feb. 2018, doi:10.1038/s41467-018-03117-y

7.Lee, Jae Young et al. “The deubiquitinating enzyme, ubiquitin-specific peptidase 50, regulates inflammasome activation by targeting the ASC adaptor protein.” FEBS letters vol. 591,3 (2017): 479-490. doi:10.1002/1873-3468.12558

8.Scott, Helen L et al. “A dual druggable genome-wide siRNA and compound library screening approach identifies modulators of parkin recruitment to mitochondria.” The Journal of biological chemistry vol. 295,10 (2020): 3285-3300. doi:10.1074/jbc.RA119.009699

9.Naik, Nenavath Gopal et al. “Epigenetic factor siRNA screen during primary KSHV infection identifies novel host restriction factors for the lytic cycle of KSHV.” PLoS pathogens vol. 16,1 e1008268. 10 Jan. 2020, doi:10.1371/journal.ppat.1008268

10.de Wilde, Adriaan H et al. “A Kinome-Wide Small Interfering RNA Screen Identifies Proviral and Antiviral Host Factors in Severe Acute Respiratory Syndrome Coronavirus Replication, Including Double-Stranded RNA-Activated Protein Kinase and Early Secretory Pathway Proteins.” Journal of virology vol. 89,16 (2015): 8318-33. doi:10.1128/JVI.01029-15