The synthesis of a gene coding for horseradish peroxidase (HRP, isozyme c; EC 1.11.1.7) is described using a polymerase chain reaction (PCR)-mediated gene synthesis approach developed in our laboratory. In this approach, all the oligonucleotides making up the gene are ligated in a single step by using the two outer oligonucleotides as PCR primers and the crude ligation mixture as the target. Th...

The traditional markerless gene deletion technique based on overlap extension PCR has been used for generating gene deletions in multidrug-resistant Acinetobacter baumannii. However, the method is time-consuming because it requires restriction digestion of the PCR products in DNA cloning and the construction of new vectors containing a suitable antibiotic resistance cassette for the selection o...

OBJECTIVE We attempted to reveal the changes of the human telomerase reverse transcriptase (hTERT) alternative splicing pattern in gastric carcinogenesis. METHODS Three alternative splicing sites (alpha, beta, gamma) were selected and designed PCR primer. The expression of 8 hTERT alternative splicing variants (ASVs) in normal gastric mucosa, precancerous lesions and gastric cancer were detec...

Ultra-deep RNA sequencing has become a powerful approach for genome-wide analysis of pre-mRNA alternative splicing. We develop MATS (multivariate analysis of transcript splicing), a bayesian statistical framework for flexible hypothesis testing of differential alternative splicing patterns on RNA-Seq data. MATS uses a multivariate uniform prior to model the between-sample correlation in exon sp...

With the goal of identifying splicing alterations in myotonic dystrophy 1 (DM1) tissues that may yield insights into targets or mechanisms, we have surveyed mis-splicing events in three systems using a RT-PCR screening and validation platform. First, a transgenic mouse model expressing CUG-repeats identified splicing alterations shared with other mouse models of DM1. Second, using cell cultures...

For the purpose of splicing widening bridges without interrupting traffic in expressway reconstruction and extension projects, different types anti-differential disturbance concrete (ADC) were developed. This paper experimentally investigated mechanical performance ADC under influence traffic–vehicle coupling vibration. The experimental device, with one end fixed other vertically vibrated, was ...

In the last several years, the use of double-stranded DNA templates together with thermostable-polymerase PCR has essentially replaced the use single-stranded DNA templates using the thermolabile polymerase for in vitro mutagenesis. Numerous PCR methods are now available, such as overlap-extension PCR, megaprimer PCR, and inverse PCR. All these PCR methods are reliable, effective, and convenien...

PURPOSE Prior studies have demonstrated tumor-specific alternative splicing events in various solid tumor types. The role of alternative splicing in the development and progression of head and neck squamous cell carcinoma (HNSCC) is unclear. Our study queried exon-level expression to implicate splice variants in HNSCC tumors. EXPERIMENTAL DESIGN We performed a comparative genome-wide analysis...

Alternative splicing provides a critical and flexible layer of regulation intervening in many biological processes to regulate the diversity of proteins and impact cell phenotype. To identify alternative splicing differences that distinguish epithelial from mesenchymal tissues, we have investigated hundreds of cassette exons using a high-throughput reverse transcription-PCR (RT-PCR) platform. E...

3 Running ASpli 2 3.1 To BIN or not to BIN . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2 3.2 Read Counting . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3 3.2.1 Mapping file and bam loading . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 4 3.2.2 Overlap features and read ali...