Bias in topoisomerase (TOPO)-cloning of multitemplate PCR products using locked nucleic acid (LNA)-substituted primers

Liu, Yuan; Döring, Jessica; Hurek, Thomas

doi:10.1016/j.mimet.2012.10.002

Cited by 2 publications

(1 citation statement)

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“…A locked nucleic acid (LNA) is a nucleic acid analogue containing a methylene bridge that connects the 2’-oxygen of ribose with the 4’-carbon [23]. The introduction of LNA into the DNA oligomer improves the hybridization affinity for complementary sequences, and the LNA-based probe shows strong mismatch discriminatory power [24–26]. In this study, the G145R mutant as a minor strain as well as a major strain was detected in chronically infected children with HBV due to breakthrough infection using LNA-based probe real-time PCR.…”

Section: Introductionmentioning

confidence: 99%

Evaluation of the G145R Mutant of the Hepatitis B Virus as a Minor Strain in Mother-to-Child Transmission

et al. 2016

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The role of the hepatitis B virus (HBV) mutant G145R, with a single change in amino acid 145 of the surface protein, as a minor population remains unknown in mother-to-child transmission. The minor strain as well as the major strain of the G145R mutant were evaluated in three cohorts using a locked nucleic acid probe-based real-time PCR. The breakthrough cohort consisted of children who were born to HBV carrier mothers and became HBV carriers despite immnoprophylaxis (n = 25). The control cohort consisted of HBV carriers who had no history of receiving the hepatitis B vaccine, hepatitis B immunoglobulin or antiviral treatment (n = 126). The pregnant cohort comprised pregnant women with chronic HBV infection (n = 31). In the breakthrough cohort, 6 showed positive PCR results (major, 2; minor, 4). In the control cohort, 13 showed positive PCR results (major, 0; minor, 13). HBeAg-positive patients were prone to have the G145R mutant as a minor population. Deep sequencing was performed in a total of 32 children (PCR positive, n = 13; negative, n = 19). In the breakthrough cohort, the frequency of the G145R mutant ranged from 0.54% to 6.58%. In the control cohort, the frequency of the G145R mutant ranged from 0.42% to 4.1%. Of the 31 pregnant women, 4 showed positive PCR results (major, n = 0; minor, n = 4). All of the pregnant women were positive for HBeAg and showed a high viral load. Three babies born to 3 pregnant women with the G145R mutant were evaluated. After the completion of immunoprophylaxis, 2 infants became negative for HBsAg. The remaining infant became negative for HBsAg after the first dose of HB vaccine. G145R was detected in one-fourth of the children with immunoprophylaxis failure. However, the pre-existence of the G145R mutant as a minor population in pregnant women does not always cause breakthrough infection in infants.

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Section: Introductionmentioning

confidence: 99%