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241.
Specificity of substrate recognition by the EcoRI restriction endonuclease. 总被引:58,自引:17,他引:58 下载免费PDF全文
B Polisky P Greene D E Garfin B J McCarthy H M Goodman H W Boyer 《Proceedings of the National Academy of Sciences of the United States of America》1975,72(9):3310-3314
The substrate specificity of the EcoRI restriction endonuclease can be varied in vitro by changing the pH and the ionic environment of the reaction. Phosphodiester bond cleavage occurs at a DNA hexanucleotide sequence d(N-G-A-A-T-T-C-N)/d(N-C-T-T-A-A-G-N) when the ionic strength is high, 100 mM Tris-HCl, 50 mM NaCl, 5 mM MgCl2, and the pH is approximately 7.3. Lowering the ionic strength to 25 mM Tris-HCl, 2 mM MgCl2, and adjusting the pH to 8.5 reduces the recognition specificity of the EcoRI endonuclease to the tetranucleotide sequence, d(N-A-A-T-T-N)/d(N-T-T-A-A-N). The enzymatic activity responsible for this substrate recognition is referred to as EcoRI. Cleavage of pVH51 plasmid DNA under EcoRI conditions results in a number of partial digest fragments, some of which disappear slowly over a prolonged digestion period. This suggests that different recognition sites are cleaved at different rates. Comparison of DNA fragment patterns of modified and unmodified pVH51 DNA indicates that the canonical EcoRI sequence is the most rapidly cleaved site under EcoRI conditions. DNA modified in vivo by the EcoRI methylase is not cleaved by the EcoRI endonuclease under standard conditions, but is cleaved under EcoRI conditions at sites other than the standard EcoRI substrate. 相似文献
242.
Saumya Jayakumar Michael S. Middleton Eric J. Lawitz Parvez S. Mantry Stephen H. Caldwell Hays Arnold Anna Mae Diehl Reem Ghalib Magdy Elkhashab Manal F. Abdelmalek Kris V. Kowdley C. Stephen Djedjos Ren Xu Ling Han G. Mani Subramanian Robert P. Myers Zachary D. Goodman Nezam H. Afdhal Rohit Loomba 《Journal of hepatology》2019,70(1):133-141
243.
Zobair Younossi Maria Stepanova Arun J. Sanyal Stephen A. Harrison Vlad Ratziu Manal F. Abdelmalek Anna Mae Diehl Stephen Caldwell Mitchell L. Shiffman Raul Aguilar Schall Bryan McColgan G. Mani Subramanian Robert P. Myers Andrew Muir Nezam H. Afdhal Jaime Bosch Zachary Goodman 《Journal of hepatology》2018,68(6):1365-1370
244.
Immunologic status of hemophilia patients treated with cryoprecipitate or lyophilized concentrate 总被引:1,自引:0,他引:1
We evaluated 37 patients with moderate or severe hemophilia A and six patients with severe factor IX deficiency for clinical or laboratory evidence of immune abnormalities. Patients were assigned to one of four groups according to the type of clotting factor replacement. Twenty patients had received only cryoprecipitate during the two years preceding the evaluation (group I); 11 additional patients were treated predominantly with cryoprecipitate but had also received up to nine bottles of factor VIII concentrate (group II); six patients received factor VIII concentrate (group III); six patients received factor IX concentrate (group IV). There was no clinical or laboratory evidence of immunodeficiency among the 43 patients. The mean absolute number of Th cells was normal in all patient groups, but the mean absolute number of Ts cells was increased compared with controls, both in patients treated with cryoprecipitate and in patients treated with factor VIII or factor IX concentrate. There was no correlation between the Th/Ts ratio and patient age, alanine aminotransferase level, hepatitis serology, in vitro lymphocyte function, or amount of clotting factor administered. Our observations demonstrate that the volunteer or commercial origin of clotting factor replacement cannot fully explain the alterations in lymphocyte subset distribution previously described in patients with hemophilia A. 相似文献
245.
RD Vaithilingam SH Safii NA Baharuddin LP Karen‐Ng R Saub F Ariffin H Ramli A Sharifuddin MFH Hidayat R Raman YK Chan NA Rani RA Rahim N Shahruddin SC Cheong PM Bartold RB Zain 《Oral diseases》2015,21(1):e62-e69
Periodontal bio‐repositories, which allow banking of clinically validated human data and biological samples, provide an opportunity to derive biomarkers for periodontal diagnosis, prognosis and therapeutic activities which are expected to improve patient management. This article presents the establishing of the Malaysian Periodontal Database and Biobank System (MPDBS) which was initiated in 2011 with the aim to facilitate periodontal research. Partnerships were established with collaborating centres. Policies on specimen access, authorship and acknowledgement policies were agreed upon by all participating centres before the initiation of the periodontal biobank. Ethical approval for the collection of samples and data were obtained from institutional ethics review boards. A broad‐based approach for informed consent was used, which covered areas related to quality of life impacts, genetics and molecular aspects of periodontal disease. Sample collection and processing was performed using a standardized protocol. Biobanking resources such as equipment and freezers were shared with the Malaysian Oral Cancer Database and Tissue Bank System (MOCDTBS). In the development of the MPDBS, challenges that were previously faced by the MOCDTBS were considered. Future challenges in terms of ethical and legal issues will be faced when international collaborations necessitate the transportation of specimens across borders. 相似文献
246.
247.
Kirwan LD MacLusky NJ Shapiro HM Abramson BL Thomas SG Goodman JM 《The Journal of clinical endocrinology and metabolism》2004,89(4):1618-1629
Previous studies have shown that conjugated estrogens and continuous medroxyprogesterone increases heart disease risk in healthy women. Little is known about the effects of the natural ovarian hormones estradiol and progesterone on cardiovascular function at rest and exercise. The purpose of this study was to investigate the short- and longer-term effects of a cyclic format of hormone replacement therapy (HRT) (1 mg estradiol daily with cyclic micronized progesterone, 200 mg for 10 d/month) on cardiovascular function at rest and during exercise in healthy, postmenopausal women. A double-blind, cross-over study was conducted in 31 patients. Peak oxygen uptake and ventilatory threshold in addition to submaximal cardiac output were determined. Peripheral measures of resting and peak ischemic blood flows were also determined. Measurements were made at baseline, after 4 h of estrogen/placebo exposure, and subsequently after 1, 2, and 3 months. The sequence of data collection was repeated after 6-wk washout. Oral estradiol with cyclic micronized progesterone increases peak ischemic peripheral blood flow chronically but fails to improve exercise tolerance and peak oxygen uptake. Similarly, submaximal central cardiovascular function is unaffected by HRT. This suggests that estradiol has a beneficial effect on peripheral blood flow, but this benefit offers little advantage in terms of peak exercise performance after 3 months of HRT. 相似文献
248.
249.
N. Gerber D. H. Ilson A. J. Wu Y. Y. Janjigian D. P. Kelsen J. Zheng Z. Zhang M. S. Bains N. Rizk V. W. Rusch K. A. Goodman 《Diseases of the esophagus》2014,27(3):235-241
This study looks at toxicity and survival data when chemoradiation (CRT) is delivered using intensity‐modulated radiation therapy (IMRT) after induction chemotherapy. Forty‐one patients with esophageal adenocarcinoma treated with IMRT from March 2007 to May 2009 at Memorial Sloan‐Kettering Cancer Center were analyzed. All patients received induction chemotherapy prior to CRT. Thirty‐nine percent (n = 16) of patients underwent surgical resection less than 4 months after completing CRT. Patients were predominantly male (78%), with a median age of 68 years (range 32–85 years). The majority of acute treatment‐related toxicity was hematologic or gastrointestinal, with 17% of patients having grade 3+ hematologic toxicity and 12% of patients having grade 3+ gastrointestinal toxicity. Only two patients developed grade 2–3 pneumonitis (5%) and 5 patients experienced post‐operative pulmonary complications (29%). Eight patients (20%) required a treatment break. With a median follow up of 41 months for surviving patients, 2‐year overall survival was 61%, and the cumulative incidences of local failure (LF) and distant metastases were 40% and 51%, respectively. This rate of LF was reduced to 13% in patients who underwent surgical resection. Surgery and younger age were significant predictors of decreased time to LF on univariate analysis. Induction chemotherapy followed by CRT using IMRT in the treatment of esophageal cancer is well tolerated and is not associated with an elevated risk of postoperative pulmonary complications. The use of IMRT may allow for integration of more intensified systemic therapy or radiation dose escalation for esophageal adenocarcinoma, ultimately improving outcomes for patients with this aggressive disease. 相似文献
250.
Amyloidosis is a rare but devastating condition caused by deposition of misfolded proteins as aggregates in the extracellular tissues of the body, leading to impairment of organ function. High clinical suspicion is required to facilitate early diagnosis. Correct identification of the causal amyloid protein is absolutely crucial for clinical management in order to avoid misdiagnosis and inappropriate, potentially harmful treatment, to assess prognosis, and to offer genetic counselling if relevant. This review summarises the current evidence on which the diagnosis and subtyping of amyloidosis is based, outlines the limitations of various diagnostic techniques, particularly in an Australian and New Zealand context, and discusses optimal strategies for the diagnostic approach to these patients. Recommendations are provided for when particularly to suspect amyloidosis, what investigations are required, as well as an approach to accurate subtyping of amyloidosis. 相似文献