Optimization of PCR conditions to amplify microsatellite loci in the bunchgrass lizard (Sceloporus slevini) genomic DNA

<p>Abstract</p> <p>Background</p> <p>Microsatellites, also called Simple Sequence Repeats (SSRs), repetitions of nucleotide motifs of 1-5 bases, are currently the markers of choice due to their abundant distribution in the genomes, and suitability for high-throughput analysis. A total of five different primer pairs were optimized for polymerase chain reaction (PCR) to amplify microsatellite loci in total genomic DNA of bunchgrass lizards (<it>Sceloporus slevini</it>) collected from three sites in southeastern Arizona; the Sonoita Plain, Chiricahua Mountains and Huachuca Mountains.</p> <p>Findings</p> <p>The primers used for current investigation were originally designed for the Eastern Fence Lizard (<it>Sceloporus undulatus</it>). Five primer pairs were selected based on annealing temperatures for optimizing the PCR conditions to amplify with bunchgrass lizards. Different concentrations of DNA and annealing temperature were optimized. While keeping other reagents constant, a DNA concentration, 37.5 ng in the final reaction volume and PCR conditions of an initial denaturation of 94°C for five minutes, an annealing temperature of 55°C and final extension of 72°C for four minutes gave the best amplification for all the primer pairs.</p> <p>Conclusions</p> <p>Modifying the standard protocol for annealing temperatures and final extension time increases the success of cross amplification of specific microsatellite loci in the bunchgrass lizard. A loading volume of 5 ul DNA at a concentration of 10 ng/ul and a 2% agarose for gel electrophoresis were observed the best for cross amplification of selected five primer pairs on bunch grass lizard.</p> <p>Trial Registration</p> <p>The research was conducted with Arizona Game and Fish Department scientific collecting permits SP565256, SP657407 & SP749119 to Dr. Christian A d'Orgeix.</p

Chromosomal locations of the maize (Zea mays L.) HtP and rt genes that confer resistance to Exserohilum turcicum

We used 125 microsatellite markers to genotype the maize (Zea mays L.) near isogenic lines (NIL) L30HtPHtPRtRt and L30htphtpRtRt and the L40htphtprtrt line which contrast regarding the presence of the recently described dominant HtP and the recessive rt genes that confer resistance to Exserohilum turcicum. Five microsatellite markers revealed polymorphisms between the NIL and were considered candidate linked markers for the HtP resistance gene. Linkage was confirmed by bulked segregant sample (BSS) analysis of 32 susceptible and 34 resistant plants from a BC1F1 population derived from the cross (L30HtPHtPRtRt x L40htphtprtrt) x L40htphtprtrt. The bnlg198 and dupssr25 markers, both located on maize chromosome 2L (bin 2.08), were polymorphic between bulks. Linkage distances were estimated based on co-segregation data of the 32 susceptible plants and indicated distances of 28.7 centimorgans (cM) between HtP and bnlg198 and 23.5 cM between HtP and dupssr25. The same set of susceptible plants was also genotyped with markers polymorphic between L30HtPHtPRtRt and L40htphtprtrt in order to find markers linked to the rt gene. Marker bnlg197, from chromosome 3L (bin 3.06), was found linked to rt at a distance of 9.7 cM. This is the first report on the chromosomal locations of these newly described genes

Effectiveness of depression and anxiety prevention in adolescents with high familial risk: Study protocol for a randomized controlled trial

Contains fulltext : 121611.pdf (publisher's version ) (Open Access)Background: Depression and anxiety disorders during adolescence can have detrimental consequences. Both disorders are related to negative outcome in various areas during adolescence and are also predictive of depression and anxiety disorders later in life. Especially parental psychopathology and being female are risk factors that increase the probability of developing one of these disorders during adolescence. Research has shown that prevention programs have promising results, especially for adolescents who have these risk factors. Therefore, in this study, we will focus on the effectiveness of a prevention program ‘A jump forward’ that has been developed for adolescent girls with a familial risk of depression and/or anxiety. Methods/Design: We designed a randomized controlled trial to test the effectiveness of an indicated and selective prevention program aimed at depression and anxiety in adolescent girls. Adolescents aged between 11 and 15 years old with depressive and/or anxiety symptoms and with parents who show indicators of parental psychopathology will be randomly assigned to the experimental (N = 80) or control groups (N = 80). Participants in the experimental group will follow a preventive intervention, consisting of six sessions of 90 minutes each. All participants will complete baseline, intervention phase 1 (after session 2), intervention phase 2 (after session 4), post-intervention, 6 month follow-up, and 12 month follow-up assessments. Furthermore, parents will be asked to complete assessments at baseline, post-intervention, and 12-month follow-up. Primary outcome will be depressive symptoms. Secondary outcomes will be anxiety symptoms, suicidal ideation, response style, negative cognitive errors, parental emotional support and parental control, parental psychopathology, parenting stress and adolescents’ depression and anxiety symptoms according to the parents. Discussion: This paper described the study designed to evaluate a program for preventing depression and/or anxiety in high-risk adolescents over a 12-month follow-up period. If the program showed to be effective in reducing symptoms of depression and anxiety and preventing adolescents from developing clinical levels of these disorders, our results would be relevant to practice. Thus, the intervention could be used on a large scale. Moreover, this study aims to contribute to the evidence-based prevention of depression and anxiety of adolescents.7 p