A set of simple-sequence repeat (SSR) markers covering the Prunus genome

A set of 109 microsatellite primer pairs recently developed for peach and cherry have been studied in the almond x peach F(2) progeny previously used to construct a saturated Prunus map containing mainly restriction fragment length polymorphism markers. All but one gave amplification products, and 87 (80%) segregated in the progeny and detected 96 loci. The resulting Prunus map contains a total of 342 markers covering a total distance of 522 cM. The approximate position of nine additional simple sequence repeats (SSRs) was established by comparison with other almond and peach maps. SSRs were placed in all the eight linkage groups of this map, and their distribution was relatively even, providing a genome-wide coverage with an average density of 5.4 cM/SSR. Twenty-four single-locus SSRs, highly polymorphic in peach, and each falling within 24 evenly spaced approximately 25-cM regions covering the whole Prunus genome, are proposed as a 'genotyping set' useful as a reference for fingerprinting, pedigree and genetic analysis of this species.     

Expressed sequence tag profiling identifies developmental and anatomic partitioning of gene expression in the mouse prostate

Background  

The prostate gland is an organ with highly specialized functional attributes that serves to enhance the fertility of mammalian species. Much of the information pertaining to normal and pathological conditions affecting the prostate has been obtained through extensive developmental, biochemical and genetic analyses of rodent species. Although important insights can be obtained through detailed anatomical and histological assessments of mouse and rat models, further mechanistic explanations are greatly aided through studies of gene and protein expression.     

Reproduction in captive common marmosets (Callithrix jacchus)

Though sexual maturation may begin at around one year of age, first successful reproduction of the common marmoset (Callithrix jacchus) is likely to be later, and it is generally recommended that animals not be mated before 1.5 years of age. The average gestation period is estimated to be 143 to 144 days. A crown-rump length measurement taken by use of ultrasonography during the linear, rapid, prenatal growth phase (between approx. days 60 and 95) can be compared against standard growth curves to estimate delivery date to within 3 to 4 days, on average. Marmosets produce more young per delivery than does any other anthropoid primate, and have more variation in litter size. Many long-established colonies report that triplets are the most common litter size, and there is documented association between higher maternal body weight and higher ovulation numbers. Higher litter sizes generally do not generate higher numbers of viable young. Marmosets are unusual among primates in having a postpartum ovulation that typically results in conception and successful delivery; reported median inter-birth intervals range from 154 to 162 days. However, pregnancy losses are quite common; one study of a large breeding colony indicated 50 percent loss between conception and term delivery. The average life span for breeding females is around six years; the range of reported average lifetime number of litters for a breeding pair is 3.45 to 4.0. Our purpose is to provide an overview of reproduction in the common marmoset, including basic reproductive life history, lactation and weaning, social housing requirements, and common problems encountered in the captive breeding of this species. A brief comparison between marmoset and tamarin reproduction also will be provided.     

An analysis of two refolding routes for a C-terminally truncated human collagenase-3 expressed in Escherichia coli

We describe here the expression of a C-terminally truncated form of human procollagenase-3 in Escherichia coli. The protein was found almost exclusively in inclusion bodies that were solubilized and refolded by two separate methods and then purified on Ni-NTA agarose. The purified proenzyme could be activated with either trypsin or APMA and active enzyme could be purified on a peptidic hydroxamate affinity column. Competitive elution from the affinity matrix yielded a highly purified preparation.     

Improved growth of cultured brain microvascular endothelial cells on glass coated with a biological matrix

An improved method for culturing primary rat brain capillary endothelial cells on glass has been developed, using a corneal extracellular matrix coat. Since the collagen-coated plastic attachment surface conventionally used for primary cultures of brain microvascular endothelium gives a high level of background fluorescence in microfluorimetric studies, an alternative attachment surface was tested involving no plastic element. Five substrata combinations were examined and a new combination of glass and corneal endothelial extracellular matrix coat was found to provide excellent cell adhesion, culture growth and purity. Other established substrata combinations tested for comparison, either involved plastic, or used glass with collagen or carbodiimide and collagen coating but the last two gave poor endothelial cell adhesion and growth. Our method using this new attachment surface combination results in stable and pure endothelial cultures, as verified by immunocytochemistry, which are suitable for fluorimetric investigations.     

Laser microdissection of conifer stem tissues: Isolation and analysis of high quality RNA,terpene synthase enzyme activity and terpenoid metabolites from resin ducts and cambial zone tissue of white spruce (<Emphasis Type="Italic">Picea glauca</Emphasis>)

Background  

Laser microdissection (LMD) has been established for isolation of individual tissue types from herbaceous plants. However, there are few reports of cell- and tissue-specific analysis in woody perennials. While microdissected tissues are commonly analyzed for gene expression, reports of protein, enzyme activity and metabolite analysis are limited due in part to an inability to amplify these molecules. Conifer stem tissues are organized in regular patterns with xylem, phloem and cortex development controlled by the activity of the cambial zone (CZ). Defense responses of conifer stems against insects and pathogens involve increased accumulation of terpenoids in cortical resin ducts (CRDs) and de novo formation of traumatic resin ducts from CZ initials. These tissues are difficult to isolate for tissue-specific molecular and biochemical characterization and are thus good targets for application of LMD.     

AMPKα2 deficiency uncovers time dependency in the regulation of contraction-induced palmitate and glucose uptake in mouse muscle

AMP-activated protein kinase (AMPK) is a fuel sensor in skeletal muscle with multiple downstream signaling targets that may be triggered by increases in intracellular Ca(2+) concentration ([Ca(2+)]). The purpose of this study was to determine whether increases in intracellular [Ca(2+)] induced by caffeine act solely via AMPKα(2) and whether AMPKα(2) is essential to increase glucose uptake, fatty acid (FA) uptake, and FA oxidation in contracting skeletal muscle. Hindlimbs from wild-type (WT) or AMPKα(2) dominant-negative (DN) transgene mice were perfused during rest (n = 11), treatment with 3 mM caffeine (n = 10), or muscle contraction (n = 11). Time-dependent effects on glucose and FA uptake were uncovered throughout the 20-min muscle contraction perfusion period (P < 0.05). Glucose uptake rates did not increase in DN mice during muscle contraction until the last 5 min of the protocol (P < 0.05). FA uptake rates were elevated at the onset of muscle contraction and diminished by the end of the protocol in DN mice (P < 0.05). FA oxidation rates were abolished in the DN mice during muscle contraction (P < 0.05). The DN transgene had no effect on caffeine-induced FA uptake and oxidation (P > 0.05). Glucose uptake rates were blunted in caffeine-treated DN mice (P < 0.05). The DN transgene resulted in a greater use of intramuscular triglycerides as a fuel source during muscle contraction. The DN transgene did not alter caffeine- or contraction-mediated changes in the phosphorylation of Ca(2+)/calmodulin-dependent protein kinase I or ERK1/2 (P > 0.05). These data suggest that AMPKα(2) is involved in the regulation of substrate uptake in a time-dependent manner in contracting muscle but is not necessary for regulation of FA uptake and oxidation during caffeine treatment.