Methods and compositions for simultaneously sequencing a nucleic acid template sequence and copy sequence

A composition, comprising:

• a nucleic acid molecule comprising a first construct strand and a second construct strand,
• wherein:
• the first construct strand comprises, from 5′ to 3′, a first sequencing primer region, a first template sequence, a first nucleic acid linker, a copy of the first sequencing primer region, and a single copy of the first template sequence;
• and
• the second construct strand comprises, from 5′ to 3′, a second template sequence, wherein the second template sequence is a reverse complement of the first template sequence, a second sequencing primer region,
• wherein the second sequencing primer region is a reverse complement of the first sequencing primer region, a second nucleic acid linker, and
• wherein the second nucleic acid linker is a reverse complement of the first nucleic acid linker, a single copy of the second template sequence, and a copy of the second sequencing primer region.

A composition, comprising:

• a nucleic acid molecule comprising a first construct strand and a second construct strand, wherein:
• the first construct strand comprises, from 5′ to 3′, a first sequencing primer region, a first template sequence, a first nucleic acid linker, a copy of the first sequencing primer region,
• wherein substantially all cytosine bases in the copy of the first sequencing primer region are methylated, and a copy of the first template sequence,
• wherein substantially all cytosine bases in the copy of the first template sequence are methylated;
• and
• the second construct strand comprises, from 5′ to 3′, a second template sequence, wherein the second template sequence is a reverse complement of the first template sequence, a second sequencing primer region, wherein the second sequencing primer region is a reverse complement of the first sequencing primer region, a second nucleic acid linker,
• wherein the second nucleic acid linker is a reverse complement of the first nucleic acid linker, a single copy of the second template sequence,
• wherein substantially all cytosine bases in the copy of the second template sequence are methylated, and a copy of the second sequencing primer region, and
• wherein substantially all cytosine bases in the copy of the second sequencing primer region are methylated.