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To identify a wide spectrum of drug resistance conferring mutations, we generated a high complexity library of mutagenized BCR-ABL cDNA in a retroviral vector and introduced this into cells by retroviral transduction.
With the software version RTA/OLB v1.8, a complex library without base composition biases can be loaded with 600,000-700,000 per mm, while a low complexity library should be loaded in the range of 500,000-600,000 per mm.
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These are high complexity libraries (over 1010 different clones each) derived from B cells of healthy donors [5].
The larger genome size of this species meant that reduced complexity libraries were constructed using the restriction enzyme Alu I.
To overcome the read length limitation, research groups have developed complex library preparation processes, which can be time-consuming, difficult, and result in low complexity libraries.
Low complexity libraries with less than 30% of mappable reads or samples with less than 800,000 mappable reads (6 in total) where excluded from further analysis.
Analysis of reduced complexity libraries starts by mapping quality-filtered reads to a reference genome using software such as ELAND or BWA.
Differences between low and high complexity libraries, are caused by an increased background signal (lower base qualities) and cluster tracking issues (N bases) which occur when many reads have the same base in a cycle and thus end up on the same image.
The 10 k Membranome phage-Ab collection is a "specialized", low-complexity library of clones which bind receptors expressed in their native form on the cell surface.
Agarose gel electrophoresis of the muscle reverse library revealed the presence of 5 clear bands, which indicated a low-complexity library.
Although these sequences were isolated from a higher-complexity library, these aptamer beacons did not yield functional sequences with better signaling capabilities.
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