Reverse complementary sequences in FASTQ data

[Yunyu]

Hi,

I am trying to analyze our fastq.gz data along with the GeckoV2 A+B library which provided online. Our fastq data contains reverse complementary sequences. 

I've tried using the default regular expression CACC(.{20}) and click the sequence is reverse complementary to extract the sgRNA info from our fastq file. However, it shows the warning message as "The entered FASTQ regular expression did only work for 5 to 5% of sequences within your FASTQ files.This is an indication for a wrong FASTQ Regular Expression or a bad quality of your sequencing library." 

I've also tried using the custom regular expression as (.{20}GGCG to extract sgRNA sequence in our fastq file and align with GeckoV2 A+B library. It seems worked out in the beginning. However, on the review data page, the sgRNA extraction ratio is 90%, but less than 1% sequences were aligned to the library.  

Could you help with the situation? Thank you very much for the help!

Best,

Yunyu