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In order to verify the accuracy of the qPCR system, known genotype samples and random samples are employed.
Both polymerase chain reaction restriction fragment length polymorphism (PCR RFLP) and high resolution melting (HRM) methods were developed to genotype samples.
In order to reduce the time and effort required to produce a single high resolution HLA genotype, samples were typed in parallel by Sanger sequencing and probe hybridization to obtain G level typing resolution.
Objectives: An HCV genotyping protocol using HCV TMA linked with LiPA (TMA LiPA) was developed and tested for ability to genotype samples that had previously failed genotyping by PCR-based methods in conjunction with LiPA.
In addition, we excluded 60 duplicate genotype samples and removed 9 subjects with ethnic backgrounds other than African-origin (Black) or European-origin (White).
For determining the epidemiological transmission reservoir of the unassigned genotype samples, the patient epidemiological information including the age, gender, ethnicity, place of birth, route of transmission, plasma sampling date were collected from the Integrated Treatment Centre, Department of Health.
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Each genotype sampled from the coalescent is copied N0/10 times into the initial population.
We selected binary markers (Supplementary Table 2) from the International Society of Genetic Genealogy10 database and from whole Y-chromosome sequencing and genotyped samples either by direct Sanger sequencing or RFLP assays.
We genotyped samples for HOXB13 G84E using the MassARRAY® system.
Genotyping samples were processed as described previously [42].
Genotyped samples were amplified with Easy-A high fidelity taq (Stratagene, La Jolla, California) and TA-cloned into pCRII (Invitrogen, Carlsbad, California).
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