By Kevan M. A. Gartland, Michael R. Davey
Agrobacterium Protocols bargains starting and skilled researchers the main finished choice of step by step protocols for the genetic manipulation of vegetation utilizing Agrobacterium. the themes diversity from the upkeep of bacterial tradition collections to points of the metabolism and body structure of reworked tissues and transgenic vegetation. Drawing at the paintings of major scientists from laboratories all over the world, Agrobacterium Protocols presents a wealth of suggestions for introducing particular DNA sequences into aim plant species and discusses the environmental implications of genetically engineered vegetation. Its targeted approaches will facilitate fast move of complicated suggestions to different laboratories and their exploitation in primary and utilized plant biology.
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Extra resources for Agrobacterium Protocols (Methods in Molecular Biology Vol 44)
32 g MgS04. 004 g MnCl, . 4 H,O. 8 with 6N KOH. Autoclave. The pH of this stock solution tends to be modified after autoclaving. The autoclaved solution is readjusted to the desired pH and autoclaved a second time. 6. 0: 500 mL neutral ATx2 stock solution, 1 g ammonium sulfate, 2 g glucose, 15 g Bacto-Agar (Difco, Detroit, MI). Final vol, 1 L. Ammonmm sulfate and glucose are prepared as 100X stock solutions and are autoclaved separately. Agar is also autoclaved separately in water. 7. 8: 500 mL acidic ATx2 stock solution, 1 g ammonium sulfate, 2 g glucose.
All the pGA binary vectors reported in this chapter carry the wild-type npt gene. 3/01 HXSHpK C Bg 1111 1 I, Fig. 1. A circular map of pGA628. 3), right border (BR), left border (BL), npt gene, and multiple cloning site. oriT, pRK2 origin of conjugative transfer; oriV, pRK2 origin of replication; tetracycline resistant gene; t$A, transacting replication factor gene; 5, transcription termination region of the gene 5 of pTiA6; 7’, transcription termination region of the gene 7 of pTiA6. Restriction enzyme sites: B, BarnHI; Bg, BgllI; C, Cl&; EcoRI; H, HindIII; Hp, Hpai; K, KpnI; P, PstI; S, SstI; Sa, MI; St, SstII; X, XbaI.
Totowa, 37 NJ Dion et al. 38 2. 1. Measurement of Bacterial Growth 1, Spectrophotometer to accomodate 1%mm diameter test tubes. 2. Erlenmeyer flasks (50~mL vol) fitted with a 18-mm diameter test tube as a side arm. These test tubes should be calibrated for measurements of optical density at 600 nm (OD6t,0). 3. Incubator with rotary shaker. 4. 010 g FeS04. 004 g MnCl, a4 H,O. 0 with 6N KOH and autoclave. A slight precipitate will develop on autoclaving, which may be removed by filtration on Whatman (Maidstone, UK) no.
Agrobacterium Protocols (Methods in Molecular Biology Vol 44) by Kevan M. A. Gartland, Michael R. Davey