I have collected a such sentence:
It's not what you do once in a while, it's what you do day in and day out that makes the difference.
Well,I know what the sentence say,but It have two subject???
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What's at issue here is not two subjects but two entire independent clauses, each with its own subject and predicate.
It's not what you do once in a while.
It's what you do day in and day out that makes the difference.
When start learning to write your teachers insist on your following certain "rules". One of those "rules" is the prohibition of comma splicing: joining two independent clauses with a comma instead of a conjunction like and or but.
But this isn't a real rule: it's a teaching rule, for children, who tend to run everything together when they write. As you get older, and particularly as you read more and more, you find that rules like this are frequently broken by Real Writers when they have good reasons for doing so.
This is a case where there is a good reason for employing a comma splice. What you have here is two contrasting independent clauses which have the same structure—and which in fact share the same final complement. This is the full thought which underlies the sentence, laid out so you see the parallels:
It's not what you do once in a while [that makes the difference],
it's what you do day in and day out that makes the difference.
The author expresses this as a single sentence to make sure you understand that the first clause is incomplete: it's not a simple declaration, it's an it cleft construction, which isn't finished until it's completed with the relative clause at the end.