Notes: In this paper, proteins were expressed from PCR-generated template DNA using TNT^® Quick for PCR DNA, the TNT^® T7 Wheat Germ Extract System and the E.coli T7 S30 Extract System. (2603)

Notes: The authors used the SV Total RNA Isolation System to purify total RNA from 1 ml of whole blood anticoagulated with EDTA for RT-PCR studies of prostate-specific antigens. (2470)

Notes: The Wizard® Genomic DNA Purification Kit was used to isolate genomic DNA from wildtype and engineered mutants of B. bronchiseptica. The isolated genomic DNA was used in PCR to analyze the size of the dnt gene. The engineered deletion mutants produced a 1.7kb PCR product and the wildtype gene produced a 2.3kb product. (2293)

Notes: PAI-1 is an inhibitor of tissue-type plasminogen activator (t-PA) and has been shown to have neuroprotective activity through the TGF-β pathway. The authors performed RTPCR experiments to study PAI-1 expression in cultured neurons and astrocytes. To confirm specificity of PAI-1 amplified products, the products were cloned into pGEMT vectors and sequenced. Wildtype and PAI-1 deficient astrocytes were transfected  (using the cationic lipid reagent, Transfast Transfection Reagent) with a luciferase reporter gene driven by the TGF-β1 response element (CAGA-luc) to determine if PAI-1^-/- cells could transduce TGF-β1 signal. Luciferase activity was quantitated using the Luciferase Assay Kit. (2608)

Notes: The authors determined allele frequencies for the Penta E and Penta D loci in 269 unrelated Austrian Caucasians using the PowerPlex^® 16 System. DNA was purified from blood samples using a Qiagen kit, and 1ng of DNA was amplified using a GeneAmp^® PCR System 9600 as directed by the manufacturer. Amplified products were detected using an ABI PRISM^® 310 Genetic Analyzer. (3851)

Notes: The authors investigated expression of BRCA1 from alternative transcripts possessing different 5′ UTRs. RT-PCR using Promega's AMV reverse transcriptase was performed to generate cDNAs from total RNA isolated from human tissue. Promega's Taq DNA Polymerase was used for the PCR reaction. The authors also prepared two mRNAs that contained one or the other of the 2 alternative 5′ UTRs (ex1a or ex1b) fused with the luciferase coding region. To generate the corresponding DNA for these constructs, the luciferase coding region was amplified from Promega's pGEM® Vector and ligated to PCR-amplified ex1a or exlb. These ligation products served as the template amplifying two cDNA constructs (exla-luc or ex1b-luc). Ten additional cDNAs were made containing a variety of changes to the 5′UTR region of BRCA1. In vitro translation experiments were performed to determine how the composition of the 5′UTR affected protein expression levels (using Promega's RNasin to protect transcribed mRNA; T7 polymerase buffer; and rabbit reticulocyte and wheat germ extracts for translation). The authors conclude that secondary structures associated with elements in the longer 5′UTR reduced translation rates and could be responsible for the reduced expression of BRCA1 often associated with spontaneous ovarian and breast cancers. (2441)

Notes: The Access RT-PCR System was used to generate nuclear gene fragments from Chlamydomonas reinhardtii for use as probes for RNA blot hybridization assays and microarrays. RT-PCR reaction products were cloned into the pGEM^®-T Easy Vector for sequence verification and to allow easier manipulation. Details of the generation of microarray slides printed on GAPII glass slides (Corning) are provided. (2694)

Notes: T4 Polynucleotide Kinase was used to phosphorylate primers before ligating them into a phagemid vector to allow cloning of Sfi I-restricted DNA fragments. Phagemid was prepared from XL1-Blue cells using the Wizard® Minipreps DNA Purification System. Inserts were PCR amplified from the phagemid and purified using the Wizard® PCR Preps DNA Purification System. In separate experiments, the cDNA for the melanin concentrating hormone receptor 1 cDNA was cloned from total melanocyte RNA using MMLV Reverse Transcriptase. In vitro translation of the cDNA was performed using the TnT T7 Coupled Reticulocyte Lysate System and Canine Pancreatic Microsomal Membranes. (2602)

Notes: MMLV reverse transcriptase and RNasin Ribonuclease Inhibitor were used to produce cDNA from total RNA isolated from transfected and untransfected human breast cancer cells. The coding region of LKB1 was amplified by PCR using Taq DNA Polymerase and buffer from Promega. (2601)

Notes: Total herpesviral RNA was isolated from HeLa cells, BC-1 cells, 293 cells and human heart tissue using the RNAgents^® Total RNA Isolation System. The isolated RNA was used for RT-PCR.  First-strand cDNA synthesis was accomplished using Promega's M-MLV Reverse Transcriptase and Random Hexamer Primers. (2563)

Notes: The PowerPlex^® 16 System was used to generate population data for 196 unrelated individuals from northern, central and southern Tunisia. Blood was collected on FTA^® paper, and DNA was extracted and amplified as directed by the manufacturer. Amplified products were detected using the ABI PRISM^® 310 Genetic Analyzer. (3835)

Notes: To analyze whether the presence or absence of calcium in Yersinia enterocolitica growth media affected secretion of Yersinia outer proteins (Yops), these researchers used mutational analysis on four gene products that work together to regulate yopQ expression. The various Y. enterocolitica strains were grown in TSB medium supplemented with 5mM CaCl₂ or EGTA (+/-Ca²⁺). After a 2-hour incubation followed by a 2-hour induction, the cultures were centrifuged and the supernatant was separated from the cell pellet. After resuspension in cell lysis buffer, the pellet was subjected to French press and 100µl aliquots were used with the SV Total RNA Isolation System. The eluted total RNA was digested with DNase, phenol/chloroform extracted and ethanol precipitated. Once resuspended, 0.6µg of Yersinia RNA was used in two-step RT-PCR to amplify the yopQ gene and the amplimer was analyzed on an ethidium bromide-stained 2% agarose gel. (3071)

Notes: The authors performed a population study using the PowerPlex^® 16 System. (2277)

Notes: In this study the PowerPlex^® 1.2 System and the FFFL Multiplex (F13A01, FESFPS, F13B, LPL) were used in a validation study of short tandem repeat systems for parentage testing. (2274)

Notes: In this paper, extracted total RNA was treated with RQ1 RNase-Free DNase prior to RT-PCR analysis. (2365)

Notes: The RNAgents^® Total RNA Isolation System was used to isolate viral RNA from the pellet resulting from centrifugation of 20ml of allantoic fluid or tissue culture supernatants. The isolated RNA was used for RT-PCR. (2575)

Notes: The PowerPlex^® 16 System was used in a concordance study involving more than 500 population database samples that included African Americans, Bahamians, and Southwestern Hispanics. Samples were typed using PowerPlex^® 16 and the Profiler Plus™ and COfiler™ kits. The results show that the primers used in these systems are reliable for typing reference samples destined for use in CODIS. (2253)

Notes: The SV Total RNA Isolation System was used to isolated RNA from various dissected mouse brain areas. Ten microliters of the isolated RNA were used for RT-PCR analysis of gene expression patterns. (2186)

Notes: The SV Total RNA Isolation System was used to isolated RNA from the syncytiotrophoblast of human term placentas. The isolated RNA was used for RT-PCR with M-MLV Reverse Transcriptase used for the RT reaction. The 110 kDa PKD2 protein was expressed in vitro with the TNT^® T7 Coupled Reticulocyte Lysate System with and without Canine Pancreatic Microsomal Membranes. The channel activity of the expressed protein was examined and the functional channel could be inhibited by known inhibitors. Expressing the luciferase control with the membranes did not produce the same effect. (2187)

Notes: The cDNA for the TERE1 gene was amplified by PCR and directly subcloned into the pTARGET™ Mammalian Expression Vector. The 338-amino acid (36.8kDa) TERE1 protein was stably expressed in three human bladder transitional cell carcinomas (J82, T24 and 1376 TCC) and the human embryonic kidney cell line, HEK 293. Stable transfectants were selected with the antibiotic G-418. Overexpression of the protein caused 80-90% inhibition of cellular proliferation in two of the transitional cell carcinoma cell lines and inhibition of aneuploidy in the third. Transfections were accomplished in 24-well plates with the Tfx™-20 Transfection Reagent. Excellent details of the 1 hour transfection protocol are presented. (2596)

Notes: The SV Total RNA Isolation System was used to isolate total RNA from primary human coronary artery endothelial cells. The isolated RNA was used to make gene array ³³P-labeled targets on nylon cDNA microarray filters using Oligo(dT) and M-MLV Reverse Transcriptase, RNase H-. The same material was used for first-strand synthesis in RT-PCR. (2697)

Notes: Taq Polymerase was used to amplify mitochondrial sequences from various laser-capture microdissected rat DNA samples. A 15.7 kilobase PCR fragment was cloned into the pGEM^®-T Easy Vector. The resultant clone was used to identify sequence breakpoints. (3234)

Notes: The cDNA for the short isoform of the caspase-2 mRNA was amplified and cloned directly into the pTARGET™ Mammalian Expression Vector. The protein was stably expressed in U937 human leukemic cells by selection with the antibiotic G-418.  Stable clones were identified by PCR of the genomic DNA of selected colonies with primers to the T7 promoter of the pTARGET™ Vector and the downstream primer of the caspase-2 message. Overexpression of the protein interfered with etoposide-mediated apoptosis. (2597)

Notes: The SV Total RNA Isolation System was used to isolated RNA from autopsy samples of human brain with FTDP-17 disorder. The isolated RNA was used for semi-quantitative RT-PCR (2162)

Notes: This study reports Austrian and Croatian population data for the loci contained in the PowerPlex^® 1.2 System. (2276)