ABSTRACT
BACKGROUND: Having laboratory technicians prepare soft-tissue casts and implant abutments with or without concomitant removable temporary prostheses during the restorative phase of single-tooth replacement is an accepted practice. It can, however, result in functional and esthetic intraoral discrepancies. CASE DESCRIPTION: Single-tooth implants can be restored with crowns (like those for natural teeth) fabricated at a dental laboratory on casts obtained from final impressions of prepared implant abutments. In the case reported, the restorative dentist restored the patient's single-tooth implant after taking a transfer impression. He constructed a cast simulating the peri-implant soft tissue with final impression material and prepared the abutment on this model. His dental assistant then fabricated a fixed provisional restoration on the prepared abutment. At the patient's next visit, the dentist torqued the prepared abutment onto the implant, took a final impression and inserted the provisional restoration. A crown was made conventionally at the dental laboratory and cemented in place at the following visit. CLINICAL IMPLICATIONS: This alternative method for restoring single-tooth implants enhances esthetics by more accurately simulating marginal gingival architecture. It also improves function by preloading the implant through fixed temporization after the dentist, rather than the laboratory technician, prepares the abutment to the dentist's preferred contours.
Subject(s)
Crowns , Dental Implants, Single-Tooth , Dental Restoration, Permanent/methods , Dental Abutments , Dental Implantation, Endosseous , Dental Impression Technique , Dental Prosthesis Design , Female , Humans , Middle Aged , Models, DentalABSTRACT
We have synthesized more than 30 different deoxyribonucleosides and triphosphates with modifications either in the base or the phosphate moiety as analogs of 2'-dGTP for DNA sequencing applications. All the modified nucleoside triphosphates were tested as substrates for DNA polymerases, including Sequenase T7 DNA polymerase or Thermo Sequenase DNA polymerase. Two of the analogs, 7-ethyl-7-deaza-dGTP and 7-hydroxymethyl-7-deaza-dGTP meet our requirements as better sequencing reagents.
Subject(s)
Guanine Nucleotides/chemical synthesis , Guanosine Triphosphate/analogs & derivatives , Sequence Analysis, DNA/methods , Guanine Nucleotides/chemistryABSTRACT
The use of Cyanine dye (Cy5 and Cy5.5) labeled dideoxy terminators with Thermo Sequenase DNA polymerase in DNA sequencing provides uniform band intensity, improved sequence read-length, and accuracy. It also greatly improves the ability to detect single base heterozygotes with dye-terminator sequencing method.
Subject(s)
Fluorescent Dyes/chemistry , Sequence Analysis, DNA/methods , Base Sequence , DNA , Taq Polymerase/chemistrySubject(s)
Crowns , Gingival Hemorrhage/prevention & control , Gingivoplasty , Hemostatics/therapeutic use , Phosphoric Acids/therapeutic use , Cementation , Epinephrine/therapeutic use , Gingival Hemorrhage/etiology , Gingivoplasty/adverse effects , Gingivoplasty/methods , Hemostasis, Surgical/methods , Hemostatic Techniques , Humans , RetreatmentABSTRACT
A combination of thermostable enzymes has been developed that produces higher quality cycle sequences. Thermo Sequenase DNA polymerase is a thermostable enzyme engineered to catalyze the incorporation of ddNTPs with an efficiency several thousandfold better than other thermostable DNA polymerases. Since the enzyme also catalyzes pyrophosphorolysis at dideoxy termini, a thermostable inorganic pyrophosphatase is needed to remove the pyrophosphate produced during sequencing reactions. Thermoplasma acidophilum inorganic pyrophosphatase (TAP) is thermostable and effective for converting pyrophosphate to orthophosphate. The use of the combination of Thermo Sequenase polymerase and TAP for cycle sequencing yields sequence data with uniform band intensities, allowing the determination of longer, more accurate sequence reads. Uniform band intensities also facilitate interpretation of sequence anomalies and the presence of mixed templates. Sequencing PCR products of DNA amplified from heterozygous diploid individuals results in signals of equal intensity from each allele.
Subject(s)
DNA-Directed DNA Polymerase/metabolism , DNA/chemistry , Pyrophosphatases/metabolism , Sequence Analysis, DNA/methods , Thermoplasma/enzymology , Cloning, Molecular , DNA-Directed DNA Polymerase/chemistry , DNA-Directed DNA Polymerase/genetics , Dideoxynucleosides/metabolism , Diphosphates/metabolism , Heterozygote , Humans , Inorganic Pyrophosphatase , Molecular Sequence Data , Mutagenesis , Taq PolymeraseABSTRACT
The incorporation of fluorescently labeled dideoxynucleotides by T7 DNA polymerase is optimized by the use of Mn2+, fluorescein analogs and four 2'-deoxyribonucleoside 5'-O-(1-thiotriphosphates) (dNTP alpha S's). The one-tube extension protocol was tested on single-stranded templates, as well as PCR fragments which were made single-stranded by digestion with T7 gene 6 exonuclease. Dye primer sequencing using four dNTP alpha S's was shown to give uniform termination patterns which were comparable to four dNTPs. Efficiency of the polymerase also appeared to improve with the dNTP alpha S's. A mathematical model was developed to predict the pattern of termination based on enzyme activity and ratios of ddNTP/dNTPs. This method can be used to optimize sequencing reactions and to estimate enzyme discrimination constants of chain terminators.
