An efficient system of genetic transformation and plant regeneration via somatic embryogenesis was established in crownvetch (Coronilla varia L.) by infecting the segments of cotyledons and hypocotyls of 15d-old seedlings with Agrobacterium rhizogenes strain 15834. Hairy roots were produced directly from the wounded surface of the explants or via calluses on hormone-free Murashige and Skoog (MS) medium after infection by A. rhizogenes. Transformed roots grew rapidly either on solid or liquid MS medium, and exhibited typical hairy root phenotypes. The highest transformation frequency (87.4%) was achieved by preculturing cotyledons for 2d and pre-treating the A. rhizogenes with suitable concentration of acetosyringone at logarithmic phase (OD600 = 0.8). The embryogenic calluses with 100% induction frequency were induced from hairy roots on MS medium containing 0.2mg/L 2,4-D, 0.5mg/L NAA and 0.5mg/L KT. Globular-, heart-, torpedo-, and cotyledon shaped somatic embryos were produced orderly and developed into plantlets when transferred the embryogenic calluses on MS medium supplemented with 0.5mg/L KT, 0.2mg/L IBA and 300mg/L proline. The transformed plants did not show differences in morphology except abundant lateral root branches compared to the non-transformed plants. However, the contents of 3-nitropropanic acid in hairy roots and leaves of one of 5 transformed clones were 57.68% and 58.17% in roots and leaves of untransformed plants, respectively. Opine paper electrophoresis revealed the integration and expression of TR-DNA. PCR analysis confirmed that the TL-DNA including 654 bp rol B sequence was inserted into the genome of transformed hairy roots and their regenerated plants.
Fabaceae ; genetics ; growth & development ; physiology ; Plant Roots ; genetics ; growth & development ; physiology ; Plants, Genetically Modified ; genetics ; growth & development ; Regeneration ; Rhizobium ; genetics ; Tissue Culture Techniques ; Transformation, Genetic

OBJECTIVETo investigate the changes of perioperative serum levels of interleukin-6 (IL-6), C-reactive protein (CRP), and cortisol, as well as gastric intramucosal pH (pHi) and plasma lactate, aiming to compare systemic changes and tissue perfusion during colorectal and orthopaedic surgical procedures.

METHODSTwenty patients were randomly assigned to two groups, 10 cases of operation on vertebral canal, 10 cases of colorectal radical operation. Venous blood was drawn at 1 day before operation, 2, 4, and 6 hours following skin incision, and 1 day after operation, in order to measure serum IL-6, CRP, and cortisol. pHi and plasma lactate were also measured at the same time points.

RESULTSSerum concentrations of IL-6 and cortisol increased gradually following operation, reaching the peak value at 6 hours from the beginning of operation. CRP was not detectable until the first day after operation. Peak concentration of IL-6 had positive relationship with CRP. These variables changed more significantly in colorectal group than that in orthopaedic group (P < 0.05). pHi decreased gradually, reaching the lowest level at 4 hours from the beginning of operation, and to more extent in colorectal group than that in orthopaedic group (P < 0.05).

CONCLUSIONIL-6 may reflect tissue damage more sensitively than CRP. Colorectal surgery might induce systemic disorder to more extent, in terms of immuno-endocrinal aspect as well as tissue perfusion, reflected with pHi.

Adult ; Aged ; C-Reactive Protein ; metabolism ; Colorectal Neoplasms ; blood ; physiopathology ; surgery ; Female ; Gastric Acidity Determination ; Gastric Mucosa ; metabolism ; Humans ; Hydrocortisone ; blood ; Hydrogen-Ion Concentration ; Interleukin-6 ; blood ; Intervertebral Disc Displacement ; blood ; physiopathology ; surgery ; Lactic Acid ; blood ; Male ; Middle Aged ; Perioperative Care ; methods ; Spinal Stenosis ; blood ; physiopathology ; surgery

BACKGROUNDIn recent years the proportion of lung adenocarcinoma (adCA) which occurs in lung cancer patients has increased. Using laser capture microdissection (LCM) combined with liquid chip-mass spectrometry technology, we aimed to screen lung cancer biomarkers by studying the proteins in the tissues of adCA.

METHODSWe used LCM and magnetic bead based weak cation exchange (MB-WCX) to separate and purify the homogeneous adCA cells and normal cells from six cases of fresh adCA and matched normal lung tissues. The proteins were analyzed and identified by matrix assisted laser desorption/ionization time-of-fight mass spectrometry (MALDI-OF-MS). We screened for the best pattern using a radial basic function neural network algorithm.

RESULTSAbout 2.895 × 10(6) and 1.584 × 10(6) cells were satisfactorily obtained by LCM from six cases of fresh lung adCA and matched normal lung tissues, respectively. The homogeneities of cell population were estimated to be over 95% as determined by microscopic visualization. Comparing the differentially expressed proteins between the lung adCA and the matched normal lung group, 221 and 239 protein peaks, respectively, were found in the mass-to-charge ration (M/Z) between 800 Da and 10 000 Da. According to t test, the expression of two protein peaks at 7521.5 M/Z and 5079.3 M/Z had the largest difference between tissues. They were more weakly expressed in the lung adCA compared to the matched normal group. The two protein peaks could accurately separate the lung adCA from the matched normal lung group by the sample distribution chart. A discriminatory pattern which can separate the lung adCA from the matched normal lung tissue consisting of three proteins at 3358.1 M/Z, 5079.3 M/Z and 7521.5 M/Z was established by a radial basic function neural network algorithm with a sensitivity of 100% and a specificity of 100%.

CONCLUSIONSDifferential proteins in lung adCA were screened using LCM combined with liquid chip-mass spectrometry technology, and a biomarker model was established. It is possible that this technology is going to become a powerful tool in screening and early diagnosis of lung adCA.

Adenocarcinoma ; metabolism ; Aged ; Female ; Humans ; In Vitro Techniques ; Lung Neoplasms ; metabolism ; Male ; Microdissection ; methods ; Middle Aged ; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization ; methods