• 西亚试剂::BAC End-Sequencing
  • 西亚试剂::BAC End-Sequencing
  • For every 4 mls of culture, dissolve the BAC DNA pellet in 40 µl of water. for example: Usually each BAC is grown in 20 mls LB/CM total, then is dispensed into one Autogen tube (4 mls in each of the 5 tubes). After miniprep, add 40 µl of water to each tube (200 µl total for each BAC).   Vortex the Autogen tube and let sit for at least 0.5 hour. Then pool the 5 samples into one for each BAC.  ...详细+
  • 西亚试剂::free shotgun sequencing library from BAC DNA
  • 西亚试剂::free shotgun sequencing library from BAC DNA
  • Miniprep of BAC DNA   Inoculate 4 - 8 tubes of 3ml/tube LB with 15ug/ml chloramphenicol with BAC clone and grow oveernight. Miniprep with AutoGen 740 according to the protocol described in our Web page. DNA pellet were resuspended in 265 ul TE buffer pH 8.0 or ddH2O.   NotI restriction digestion 265 ul miniprep DNA 30 ul of 10X New England BioLabs Buffer 3 5 ul of 10 units/ul Not 1 (New Eng...详细+
  • 西亚试剂::cDNA Production
  • 西亚试剂::cDNA Production
  •   cDNA Production Lab of Dr. Mark Barton Frank, Oklahoma Medical Research Foundation   Please read the excellent descriptions of handling RNA and of cDNA synthesis by J. Sambrook, E. F. Fritsch, and T.Maniatis in Molecular Cloning, A Laboratory Manual (Cold Spring Harbor Laboratory Press, Second Edition, New York, 1989). The methodology for the synthesis of the first strand of the cDNA summarized below is based primarily on this so...详细+
  • 西亚试剂::BAC DNA Isolation with AutoGen 740
  • 西亚试剂::BAC DNA Isolation with AutoGen 740
  • The BAC clone from glycerol culture sublibrary is innoculated into 3 ml of LB with 20µg/ml chloramphenicol and cultured at 37°C with shaking for 16hrs. The liquid bacterial culture is transferred into the tube specific for AutoGene 740 DNA Isolation System (Autogen, Inc.) For BAC DNA purification, we use alkaline lysis approach by set up " System 1" in the panel of the AutoGen 740. Then, follow the manual provided by manufacturer. Reag...详细+
  • 西亚试剂::IMMUNOSCREENING OF LAMBDA-GT11
  • 西亚试剂::IMMUNOSCREENING OF LAMBDA-GT11
  •   Adsroption of Anti-E. coli Antibodies Grow E. coli strain Y1090 in L broth at 37°C overnight. Transfer to 10 mM MgSO4 and store at 4°C for future use. These cells are good for about 1 week. Soak one 132 mm nitrocellulose membrane per Petri dish in 10mM IPTG (MW=238.31). Briefly blot the excess solution from the filters, and wrap them in cellophane. These should be moist and stored at 4°C. We typically adsorb serum with fiv...详细+
  • 西亚试剂::
  • 西亚试剂::
  • L Broth Agar 1 L of L Broth 15 g agar Autoclave L Broth Top Agar Make L Broth Agar except substitute 7.5 g agar/L. TMG (10 mM Tris-HCl, pH 7.5; 10 mM MgSO4; 0.1%     For 100 ml:            1 ml of 1M Tris-HCl pH 7            1 ml of 1M MgS04            0.1 g gelatin ...详细+
  • 西亚试剂::SCREENING A LAMBDA BACTERIOPHAGE
  • 西亚试剂::SCREENING A LAMBDA BACTERIOPHAGE
  • Plate a bacteriophage library at about 30,000 - 50,000 pfus per 150 mm Petri dish by adding an appropriate amount of bacteriophage to 500µl MgSO4-stored E. coli. Dilute bacteria, if needed (ex. VCS237 to A600 = 0.5). Incubate for 15 minutes at 37°C without shaking. Plate with 7 ml LB top agar on LB agar plates. Once the top agar has solidified, turn the plates upside down and incubate overnight at 37°C. The next day, cool the plates at...详细+
  • 西亚试剂::RFLP和RAPD技术原理和操作步骤
  • 西亚试剂::RFLP和RAPD技术原理和操作步骤
  • DNA分子水平上的多态性检测技术是进行基因组研究的基础。RFLP(Restriction Fragment Length Polymorphism,限制片段长度多态性)已被广泛用于基因组遗传图谱构建、基因定位以及生物进化和分类的研究。RFLP是根据不同品种(个体)基因组的限制性内切酶的酶切位点碱基发生突变,或酶切位点之间发生了碱基的插入、缺失,导致酶切片段大小发生了变化,这种...详细+
  • 西亚试剂::RFLP和RAPD技术原理和操作步骤
  • 西亚试剂::RFLP和RAPD技术原理和操作步骤
  • DNA分子水平上的多态性检测技术是进行基因组研究的基础。RFLP(Restriction Fragment Length Polymorphism,限制片段长度多态性)已被广泛用于基因组遗传图谱构建、基因定位以及生物进化和分类的研究。RFLP是根据不同品种(个体)基因组的限制性内切酶的酶切位点碱基发生突变,或酶切位点之间发生了碱基的插入、缺失,导致酶切片段大小发生了变化,这种...详细+