Illumina Corporation

Paul Belitz

Robert Ezra Langlois

John S Vieceli

Stephen Tanner

M Shane Bowen

Alex Nemiroski

Hui Han

Kevan Samiee

Danny Yuan Chan

Bala Murali K Venkatesan

Xiaoyu Chen

Memory efficient methods determine corrected color values from image data acquired by a nucleic acid sequencer during a base calling cycle. Such methods may: (a) obtain an image of a substrate (e.g., a portion of a flow cell) including a plurality of sites where nucleic acid bases are read; (b) measure color values of the plurality of sites from the image of the substrate; (c) store the color values in a processor buffer of the sequencer's one or more processors; (d) retrieve partially phase-corrected color values of the plurality of sites, where the partially phase-corrected color values were stored in the sequencer's memory during an immediately preceding base calling cycle; (e) determine a prephasing correction; and (f) determine the corrected color values. In various implementations, these operations are all performed during a single base calling cycle. In certain embodiments, the methods additionally include using the corrected color values to make base calls for the plurality of sites. Sequencers may be designed or configured to implement such methods.

Phasing correction

Techniques are described for dynamically correcting image distortion during imaging of a patterned sample having repeating spots. Different sets of image distortion correction coefficients may be calculated for different regions of a sample during a first imaging cycle of a multicycle imaging run and subsequently applied in real time to image data generated during subsequent cycles. In one implementation, image distortion correction coefficients may be calculated for an image of a patterned sample having repeated spots by: estimating an affine transform of the image; sharpening the image; and iteratively searching for an optimal set of distortion correction coefficients for the sharpened image, where iteratively searching for the optimal set of distortion correction coefficients for the sharpened image includes calculating a mean chastity for spot locations in the image, and where the estimated affine transform is applied during each iteration of the search.

Optical distortion correction for imaged samples

Fiducial markers are provided on patterned arrays of the type that may be used for molecular analysis, such as sequencing. The fiducials may have configurations that enhance their detection in image or detection data, that facilitate or improve processing, that provide encoding of useful information, and so forth. Examples of the fiducials may include an 'always on' type that respond to multiple frequencies of radiation used during processing and detection so as to return signals during successive cycles of imaging.

System and method with fiducials responding to multiple excitation frequencies

Fiducial markers are provided on patterned arrays of the type that may be used for molecular analysis, such as sequencing. The fiducials may have configurations and layouts that enhance their detection in image or detection data, that facilitate or improve processing, that provide encoding of useful information, and so forth. Examples of the fiducials may include offset layouts that may be useful in detecting the fiducials in different types and approaches in imaging, and that may help to distinguish regions of the array from one another in image data.

System and method with fiducials having offset layouts

Fiducial markers are provided on patterned arrays of the type that may be used for molecular analysis, such as sequencing. The fiducials may have configurations that enhance their detection in image or detection data, that facilitate or improve processing, that provide encoding of useful information, and so forth. Examples of the fiducials may include a non-closed shape that may encode information, allow for bubbles to escape during manufacture, and provide additional advantages over closed shape fiducials.

System and method with fiducials of non-closed shapes

Fiducial markers are provided on patterned arrays of the type that may be used for molecular analysis, such as sequencing. The fiducial markers may have configurations that enhance their detection in image or detection data, that facilitate or improve processing, that provide encoding of useful information, and so forth. Examples of the fiducial markers may include features and materials that are provided on or in the support of a patterned array and that return at least a portion of incident light by reflection. The fiducial markers may form gratings or other encoding configurations that assist in image processing, alignment, or other aspects of processing of the patterned array.

System and method with reflective fiducials for locating or registering locations receiving biological samples in successive cycles of fluorescent imaging

Fiducial markers are provided on a patterned array of the type that may be used for molecular analysis, such as sequencing. The fiducial markers may have configurations and layouts that enhance their detection in image or detection data, that facilitate or improve processing, that provide encoding of useful information, and so forth. Examples of the fiducial markers may include non-rectilinear layouts that may provide for more robust location of both the fiducial markers and sites of the patterned array.

System and method with fiducials in non-rectilinear layouts

Methods and systems for analysis of image data generated from various reference points. Particularly, the methods and systems provided are useful for real time analysis of image and sequence data generated during DNA sequencing methodologies.

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San Diego, CA, United States of America

Paul Belitz